Abstracts From the th Annual Meeting of the European Council for

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					Abstracts From the
12th Annual Meeting of
the European Council for
Cardiovascular Research (ECCR)


La Colle sur Loup
Nice, France
12–14 October 2007
  Abstracts From the 12th Annual Meeting of the ECCR

ECCR Oral Presentations                                                                              littermates. Intravenous injection of 2 nmol/kg CGRP, unlike AM, decreased arterial
                                                                                                     pressure equally by 18 5 mmHg in tg and wt animals. The concomitant increase in heart
                                                                                                     rate evoked by CGRP however was 3.7 times higher in tg mice than in wt animals. The
                                                                                                     effects of CGRP in tg and wt mice, unlike a decrease in arterial pressure in response to
                                                                                     01.01           20 nmol/kg AM, were suppressed by 2 mol/kg CGRP(8 –37) antagonist. Propranolol, in
ABNORMALITIES OF HEART MORPHOLOGY AT AN EARLY STAGE OF                                               contrast to hexamethonium, blocked the CGRP evoked increase in heart rate in both tg and
RENAL DISEASE—THE ROLE OF OXIDATIVE STRESS                                                           wt animals. This was consistent with the immunohistochemical localization of the V5-CLR
                                                                                                     in the superior cervical ganglion of tg mice. In conclusion, hypotension evoked by CGRP
Piecha G1, Koleganova N1, Geldyyev A1, Berger I1, Ritz E2, Gross ML1                                 in tg and wt mice was comparable, and CGRP was more potent than AM. Moreover, the
1
  Institute of Pathology, University of Heidelberg, Heidelberg, Germany, 2Department of              stronger positive chronotropic action of CGRP in tg as compared to wt mice appears to be
Medicine, University of Heidelberg, Heidelberg, Germany                                              mediated by CLR/RAMP1 CGRP receptors localized in post-ganglionic sympathetic neurons
                                                                                                     of tg mice.
Even minor reduction of GFR causes not only accelerated atherogenesis, but also cardiac
remodeling. Oxidative stress is one of the responsible mechanisms. The current study in
the ApoE -/- mouse was designed to investigate whether nephron reduction by
uninephrectomy (UNX) causes cardiac remodeling and whether this is prevented by
antioxidative treatment, compared to an ACE inhibitor. We randomized the ApoE -/- mice
to undergo UNX or sham operation and subsequent treatment with either Tempol, Ebselen,                                                                     01.04
Trandolapril, or a combination of Tempol plus Trandolapril. After 12 weeks the experiment
was terminated by perfusion fixation. Quantitative morphologic analysis of the myocardium
                                                                                                     INCREASED CONTRACTILE RESPONSE TO -ADRENERGIC STIMULATION
was performed. Untreated UNX animals had lower capillary length density (2709 407                    OF RAT HEART OVEREXPRESSING SR CaATPase
mm/mm3) and higher volume fraction of interstitial tissue (4.5 1.2%) in the myocardium
compared to sham op (3706 571, 3.1 0.7, respectively). These changes did not develop                 Wei W, Paul M, Vetter R
in UNX animals treated with Tempol (3776 534, 2.7 0.7), Ebselen (3778 524,                                     ¨                                                 ´
                                                                                                     Institut fur Klinische Pharmakologie und Toxikologie, Charite, Berlin, Germany
3.5 0.6), Trandolapril (3568 488, 3.4 1.1), or Tempol Trandolapril (3659 338,
3.1 0.5). In untreated UNX compared to sham op the expression of nitrotyrosine, TGF- 1,              To examine the effect of overexpression of the sarcoplasmic reticulum (SR) CaATPase
VEGF, and collagen I was more marked, and this was ameliorated by Tempol, Ebselen,                   SERCA2 in response to increasing concentrations of the -adrenergic agonist isoproterenol
Trandolapril, and Tempol       Trandolapril. Even minor reduction of renal function by               (Iso), we compared the contractile function of isovolumetrically contracting Langendorff-
uninephrectomy causes remodeling of the heart in the ApoE -/- mouse and this was                     perfused hearts of male transgenic rats overexpressing SR Ca ATPase (TG) with those of
ameliorated both by antioxidants and by ACE inhibitors.                                              transgene-negative littermates (NTG). Analyzing cardiac function of TG and NTG (n 6)
                                                                                                     under basal conditions, hearts of TG and NTG contracted with a comparable spontaneous
                                                                                                     rate (HR), developed similar left ventricular peak systolic pressure (LVPSP) and did not
                                                                                                     differ in maximum rates of pressure rise ( dP/dtmax) and decay (-dP/dtmax). In presence of
                                                     01.02
                                                                                                     Iso, values of HR, LVPSP, dP/dtmax, and -dP/dtmax showed a concentration dependent
ADENOSINE SWITCHES MACROPHAGES FROM A PRO-INFLAMMATORY TO                                            increase in both groups. The positive chronotropic effects of Iso did not differ between
AN ANGIOGENIC PHENOTYPE                                                                              them. However, the maximum relative increases in LVPSP, dP/dtmax and -dP/dtmax were
                                                                                                     markedly higher in TG as compared to NTG and the concentration-response curves were
Ernens I, Velot E, Vausort M, Devaux Y, Wagner DR                                                    shifted to the left. At 100 nM Iso, values of LVPSP, dP/dtmax and -dP/dtmax showed an
          ´
CRP-Sante, Luxembourg, Luxembourg                                                                    increase of 55, 97 and 133% in TG vs. 36, 76 and 96% in NTG (p 0.05 each),
                                                                                                     respectively. Calculated from the respective concentration-response-dependencies, the
Purpose: Post-infarct left ventricular remodeling is a leading cause of heart failure.               most potent lowering of the EC50 value was observed for -dP/dtmax (2.7 nM in TG vs. 3.2
Angiogenesis may positively affect ventricular remodeling. Here we report the character-             nM Iso in NTG, p 0.05). We conclude that functional overexpression of SR CaATPases
ization of the angiogenic switch induced by adenosine in human macrophages isolated                  strengthened the inotropic and lusitropic responses of ex vivo transgenic rat hearts to
from healthy volunteers and patients with acute MI. Methods: Primary monocytes isolated                -adrenergic stimulation. In the latter, the gain of function due to the action of the
from human peripheral blood mononuclear cells were differentiated by 50 ng/mL                          -agonist appears to be highest for the process of relaxation.
macrophage-colony stimulating factor for 7 days. Macrophages were preincubated for 15
minutes with adenosine (10 M) or adenosine analogs (0.1 to 10 M) before LPS
treatment (100 ng/mL for 24 hours). ELISA and real-time quantitative PCR were used to
measure levels of TNF- , Matrix Metalloproteinase (MMP)-9, and Vascular Endothelial
Growth Factor (VEGF). Results: In LPS-stimulated macrophages, adenosine induced a 83%
decrease of TNF- secretion (p 0.005). This decrease was replicated by the A3 agonist
                                                                                                                                                            01.05
Cl-IB-MECA, suggesting involvement of the A3 receptor. Adenosine increased VEGF
secretion both alone (3.8-fold, p 0.01) and in synergy with LPS (28-fold, p 0.0002).                 CEREBROVASCULAR EFFECTS OF AN ORAL GLUCOSE TOLERANCE TEST
Adenosine also increased MMP-9 expression. The effects of adenosine on VEGF and
MMP-9 were replicated by the A2a agonist C141 and blocked by the A2a (SCH58261) and                  Patel S1, Bathula R1, Page C1, Tillin T1, Chaturvedi N1, Godsland I1, Potter J2,
A2b (MRS1754) antagonists, suggesting involvement of the A2a/A2b receptors. The effects              Panerai RB3, Hughes AD1
                                                                                                     1
of adenosine on TNF- and VEGF expression were stronger in macrophages from acute MI                    International Centre for Circulatory Health, Imperial College London; 2School of
patients than healthy volunteers. Conclusion: Adenosine induces an angiogenic switch in              Medicine, Health Policy and Practice; University of East Anglia; 3Department of Medical
human macrophages characterized by decreased TNF- secretion and enhanced MMP-9                       Physics, Leicester University, UK.
and VEGF expression. This switch is more robust in patients with acute MI. Thus,
adenosine regulating agents may represent a new approach to stimulate angiogenesis.                  Background: Diabetes and insulin resistance increase stroke risk, strikingly so in
                                                                                                     Indo-Asians. The pathophysiology behind this implicates changes in cerebral autoregula-
                                                                                                     tion. However, whether this risk is attributable to acute vascular effects of hyperglycaemia,
                                                                                                     or chronic diabetic vascular change, is unknown. This study investigated changes in
                                                        01.03
                                                                                                     cerebral autoregulation as a consequence of acute hyperglycaemia, and compared
CALCITONIN GENE-RELATED PEPTIDE EVOKES SUSTAINED HIGH                                                responses between Europeans and Indo-Asians. Methods: Double blinded, placebo
AMPLITUDE TACHYCARDIA IN CALCITONIN RECEPTOR-LIKE RECEPTOR                                           controlled crossover study of healthy Indo-Asian (n 6) and European (n 5) male
TRANSGENIC MICE THROUGH STIMULATION OF MYOCARDIAL                                                    volunteers aged 18 –30 years. An oral glucose tolerance test (OGTT) induced hypergly-
SYMPATHETIC ACTIVITY                                                                                 caemia whilst subjects underwent metabolic profiling and repeated assessment of their
                                                                                                     cerebral circulation, in particular dynamic cerebral autoregulatory index (DCAI), using
Kunz TH1, Scott M2, Ittner L1, Fischer JA1, Born W1, Vogel J2                                        transcranial Doppler ultrasonography over 2 hours. Results: Overall, the DCAI decreased
1
  Research Laboratory, Orthopedic University Hospital Balgrist, Zurich, Switzerland,                 from 6.5 to 5.3 between 60 and 90mins post glucose load (p 0.05). However, these
2
  Institute of Veterinary Physiology, Zurich, Switzerland                                            marked changes were only observed in Europeans, no change in DCAI occurred in
                                                                                                     Indo-Asians. Pulsatility Index, a marker of cerebrovascular resistance, increased from 0.86
Calcitonin gene-related peptide (CGRP) and adrenomedullin (AM) are potent vasodilators               to 0.95 at 30mins (p 0.02) in all subjects, and within ethnic groups. Cerebral blood flow
and exert positive chronotropic and inotropic effects on the heart. Receptors for CGRP and           velocity dropped during the OGTT in Indo-Asians (44.9 cm/s to 40.3 cm/s between 0 and
AM are calcitonin receptor-like receptor (CLR)/receptor-activity-modifying protein (RAMP)1           30 mins; p 0.001) but not in Europeans. Conclusions: Acute hyperglycemia impairs
and CLR/RAMP2 heterodimers, respectively. To delineate distinct cardiovascular effects of            cerebrovascular autoregulation in Europeans but not Indo Asians. Conversely, flow velocity
CGRP and AM we overexpressed in mice (tg) a V5-tagged rat-CLR (V5-CLR). A smooth                     declined only in Indo-Asians. This suggests that cerebrovascular changes in response to
muscle a-actin promoter was used to direct the transgene into blood vessels. Interestingly,          acute hyperglycaemia differ by ethnicity, and require further detailed exploration to
basal arterial pressure and heart rate were indistinguishable in tg mice and in control (wt)         determine the mechanisms for stroke risk in relation to diabetes.



                                                                                               800
                                                                                                                         2007 ECCR Oral Presentations                                    801

                                                         01.06                                 reversible after the cessation of L-NAME, but persistent arterial structural alterations and
INHIBITION OF CEREBROVASCULAR ERK1/2 ACTIVATION REDUCES LATE                                   enhanced formation of EDCF may maintain elevated blood pressure even after the
CEREBRAL ISCHEMIA AND RECEPTOR UPREGULATION AFTER                                              restoration of NO-synthase activity. (VEGA 1/3429/0, APVT grant 51– 02704, GA CR
SUBARACHNOID HEMORRHAGE                                                                        305/03/0769 and IGA MH CR NR 7786 –3/2004 grant)

Ansar S, Edvinsson L
Glostrup Research Institute, Glostrup, Denmark and Clinical Sciences, Division of
Experimental Vascular Research, Lund University, Lund, Sweden

Introduction: Late cerebral ischemia that occurs after subarachnoid hemorrhage (SAH)                                                                    02.03
often results in death or severe disability. The pathophysiological mechanism responsible
for the late cerebral ischemia that occurs after SAH still remain elusive and no specific
                                                                                               REACTIVE OXYGEN SPECIES HAVE A CENTRAL ROLE IN FLOW (SHEAR
treatment exists. Our hypothesis suggests that cerebral ischemia leads to pathophysio-         STRESS)-INDUCED REMODELING IN RAT MESENTERIC RESISTANCE
logical receptor changes in the vascular smooth muscle cells via activation of extracellular   ARTERIES
signal-regulated kinase ERK1/2. Presently we wanted to find out the therapeutic window
for an ERK1/2 inhibitor and thereby investigate its clinical relevance. Methods: SAH was       Belin de Chantemele E, Vessieres E, Guihot AL, Chanoine S, Dumont O, Loufrani L,
induced by injecting 250 l blood into the prechiasmatic cistern. Six, 12 and 24 h after        Henrion D
the induced SAH the raf inhibitor SB-386023-b was injected intracisternally. Two days          UMR-CNRS 6214 INSERM U771, Angers, France
after the SAH the cerebral arteries were harvested and contractile responses to
endothelin-1 (ET-1), 5-carboxamidotryptamine (5-CT) and angiotensin II (Ang II) were           Resistance arteries have a key role in the control of local blood flow. They are able to adapt
investigated in myographs. In addition, the mRNA and protein levels of ET, 5-HT1 and AT1       or remodel in response to chronic increases in blood flow. Remodeling involves a rise in
receptors were investigated by real time PCR and immunohistochemistry. Results:                diameter and a higher endothelium-mediated dilation. Alteration in endothelium function
Treatment with the ERK1/2 inhibitor SB-386023-b at 6 h after the SAH decreased the             observed in cardiovascular and metabolic diseases is associated with excess reactive
elevated maximum contraction elicited by application of ET-1, 5-CT and Ang II in cerebral      oxygen species (ROS). We investigated the role of ROS in the adaptation of resistance
arteries considerably compared to SAH. The ERK1/2 inhibition prevented the upregulated
                                                                                               arteries to a chronic increase in blood flow in vivo. We used the mesenteric circulation in
ETB, 5-HT1B and AT1 receptor mRNA and protein levels seen after SAH.Conclusion This
study clearly suggests that ERK1/2 inhibition reduce late cerebral ischemia after SAH and      which arcading arterioles allow local arterial ligation. Rat resistance arteries were
may be an important approach towards new stroke therapies in man.                              submitted to chronic increases in blood flow by ligating sequentially second order arteries.
                                                                                               Arteries were thus submitted to high (HF) or normal flow (NF) for 3 weeks. Rats were
                                                                                               treated with the superoxide dismutase mimetic tempol or water. Arterial diameter and
                                                       02.01                                   endothelium-mediated dilation increased significantly in HF arteries in control rats. In
A NOVEL LYMPHOCYTE-DERIVED VASOACTIVE PLASMA ANGIOTENSIN                                       tempol-treated rats HF arterial diameter did not increase although endothelium-mediated
PEPTIDE                                                                                        dilation was increased compared to NF. HF remodeling was associated with eNOS
                                                                                               overexpression without change in eNOS phosphorylation. HF remodeling was also
Jankowski V, Toelle M, van der Giet M, Zidek W, Jankowski J                                    associated with gp91 and p67 overexpression. In HF arteries, acute or chronic tempol
Charite, Berlin, Germany                                                                       increased endothelium-dependent dilation. ROS production was higher in HF than in NF
                                                                                               arteries. Conclusion: In high flow-induced remodeling of resistance arteries the rise in
Angiotensin peptides play a central role in cardiovascular physiology and pathology.           diameter involves ROS overproduction, which in turn reduced endothelium-mediated
Among these peptides, angiotensin II (Ang II) has been investigated most intensively.          dilation. HF-induced rise in diameter was totally prevented by tempol, which prevented
However, further angiotensin peptides such as Ang 1–7, Ang III and Ang IV also contribute      ROS production. Nevertheless, this treatment improved NO-dependent dilation in remod-
to vascular regulation, and may elicit additional, different or even opposite effects to Ang   eled arteries
II. Here, we describe a novel Ang II-related, strong vasoconstrictive substance in plasma
from healthy humans and end-stage renal failure patients. Chromatographic purification
and structural analysis by MALDI-TOF / TOF revealed an angiotensin octapeptide with the
sequence Ala-Arg-Val-Tyr-Ile-His-Pro–Phe, which differs from Ang II in Ala1 instead of
Asp1. Des[Asp1]-[Ala1]-Ang II, in the following named Angiotensin A (Ang A), is most likely
generated enzymatically. In the presence of mononuclear leukocytes, Ang II is converted
to Ang A by decarboxylation of Asp1. Ang A has the same affinity to the AT1 receptor as                                                              02.04
Ang II, but a higher affinity to the AT2 receptor. In healthy subjects, Ang A concentrations   IS THE “ANGIOGENIC QUANTUM” A MICRO-NETWORK? SYNCHROTRON
were less than 20% of the Ang II concentrations, but the ratio Ang A / Ang II was higher       RADIATION BASED MICRO-COMPUTED TOMOGRAPHY FOR 3D
in end-stage renal failure patients. Ang A is a novel human, strong vasoconstrictive
angiotensin-derived peptide, most likely generated by enzymatic transformation through
                                                                                               EVALUATION OF CEREBRAL CAPILLARY NETWORKS IN MICE
mononuclear leukocyte-derived aspartate decarboxylase. Plasma Ang A concentration is           OVEREXPRESSING VEGF165 IN THE BRAIN
increased in end-stage renal failure. Due to its stronger agonism at the AT2 receptor, Ang
A may modulate the harmful effects of Ang II.                                                  Heinzer S1, Kuhn G2, Krucker T3, Meyer E3, Ulmann-Schuler A3, Stampanoni M4,
                                                                                               Gassmann M2, Marti HH5, Muller R1, Vogel J2
                                                                                               1
                                                                                                 Institute for Biomedical Engineering, Zurich, Switzerland, 2Institute of Veterinary
                                                        02.02                                  Physiology, Zurich, Switzerland, 3Insitute of Zoology, Zurich, Switzerland, 4Swiss Light
THE RESTORATION OF VASCULAR RESPONSES MEDIATED BY NO AND                                       Source, Paul Scherrer Institut, Villingen, Switzerland, 5Department of Physiology and
ENDOTHELIUM-DERIVED CONSTRICTING FACTOR AFTER CESSATION OF                                     Pathophysiology, Heidelberg, Germany
L-NAME ADMINISTRATION
                                                                                               The formation of a functional vascular network requires both, the generation of normal,
Paulis L1, Zicha J2, Kunes J2, Hojna S2, Kojsova S3, Pechanova O3, Simko F1                    healthy vessels and their arrangement into an effective network architecture. While our
1
  Institute of Pathophysiology, School of Medicine, Bratislava, Slovakia, 2Institute of        knowledge about the development of single vessels significantly increased during the last
Physiology, Acad Sci Czech Rep and CRC, Prague, Czech Republic, 3Institute of Normal           years, mechanisms responsible for arrangement of vessel elements into a functional
and Pathological Physiology, Slovak Acad Sci, Bratislava, Slovakia                             network are poorly understood, probably due to the lack of suitable methods for
                                                                                               quantification of the microvascular network structure. In transgenic mice overexpressing
The level of restoration of normal vascular responses in the regression of already             six-fold higher VEGF165 levels locally in the brain (tg), blood flow capacity is not increased
established hypertension may be important for blood pressure and risk reduction. We            despite 2- to 3-fold higher density of normal and perfused capillaries. We used vascular
aimed to investigate, the restoration of vascular responses mediated by NO and EDCF,           corrosion casting and hierarchical micro-computed tomography combined with a newly
altered by L-NAME administration. Four groups of Wistar rats were investigated: 5-week         developed network analysis tool to analyze vascular architecture in gray- and white-matter
L-NAME, spontaneous recovery (L-NAME for 5 weeks followed by 3-week recovery) and
                                                                                               structures of the brain of tg mice. Our results indicate that VEGF165 overexpression leads
age-matched controls. L-NAME caused systolic and diastolic blood pressure elevation
                                                                                               to formation of complete additional micro-networks connected to higher order vessels,
accompanied by reduced inner femoral artery diameter, augmented EDCF-signaling and
impaired NO-signaling (indicated by decreased NO-synthase activity in the aorta, impaired      rather than insertion of single capillaries into the existing vessel structure. This implies that
ACh-induced relaxations and abolished sensitivity of ACh-induced relaxations to L-NAME-        the smallest “angiogenic quantum”, i.e. the final, stable result of angiogenesis and
preincubation) in femoral as well as small mesenteric arteries. During spontaneous             subsequent remodeling, is not a single vessel, but rather a complete micro-network.
recovery complete restoration of NO-signaling took place. However, the inner diameter          High-resolution 3D imaging combined with network analysis can substantially improve our
remained unaffected and augmented EDCF-signaling and blood pressure were only                  understanding of vascular architecture, beneficial for the development of therapeutic
partially restored. We conclude that L-NAME administration leads to disruption of              angiogenesis as a clinical tool for applications such as wound healing or treatment of
NO-signaling in vessels with large and small diameter. This impaired NO-signaling is fully     ischemic diseases.
802          Hypertension Vol 50, No 4 October 2007


                                                                                    02.05     remodelling. Cell culture experiments with adipocytes further demonstrated the role of fat
CHRONIC ESTRADIOL ADMINISTRATION IN VIVO PROMOTES THE                                         tissue in the activation of proteolysis. 3T3-L1 preadipocytes were differentiated to
PRO-INFLAMMATORY RESPONSE OF MACROPHAGES TO TLR-4                                             adipocytes. After incubation with TNF-alpha (10 ng/ml, for 24 hours), cells expressed
                                                                                              significantly higher mRNA levels of cathepsin B (180%; p 0,05), cathepsin D (164%;
ACTIVATION. INVOLVEMENT OF THE PI(3) KINASE PATHWAY
                                                                                              p 0,05) as well as MMP2 (118%; ns) and MMP3 (10fold; p 0,01) as compared to control
                                                                                              cells. Treatment with telmisartan significantly (p 0,05) attenuated the expression of these
Calippe B.1, Douin-Echinard V.1, Laffargue M.2, Laurell H.1, Rana-Poussine V.1, Pipy B.3,
                                                                                              proteases. We conclude that enhanced secretion of proteases by adventitial adipocytes
Bayard F.1, Arnal JF1, Gourdy P.1
1                                                                                             contributes to outward aortic remodelling in obesity.
                             ´           ´
  INSERM U858, Institut de Medecine Moleculaire de Rangueil, IFR31, Toulouse,
Haute-Garonne, France, 2INSERM U563, Departement Lipoproteines et Mediateurs
                                           ´                   ´         ´
Lipidiques, IFR30, CHU Toulouse Purpan, Toulouse, Haute-Garonne, France, 3UPRESA
2405, IFR31, CHU Toulouse Rangueil, Toulouse, Haute-Garonne, France

Although estrogens are recognized to influence immune responses, their effect on                                                                     03.01
macrophage functions in vivo remains to be clarified. Indeed, short-term exposure to          SUSTAINED BLOOD PRESSURE REDUCTION BY BAROREFLEX
17 -estradiol (E2) in vitro has been reported to decrease the production of pro-              HYPERTENSION THERAPY WITH A CHRONICALLY IMPLANTED SYSTEM:
inflammatory cytokines by macrophages, as confirmed here in the model of murine               LONGTERM DATA FROM THE RHEOS DEBUT-HT STUDY IN PATIENTS
resident peritoneal macrophages in response to LPS. In striking contrast, chronic
administration of E2 to ovariectomized mice leads to a marked increase in the production
                                                                                              WITH RESISTANT HYPERTENSION
of IL-1 , IL-6, IL-12p40 and iNOS expression by these cells in the same conditions of
                                                                                              Scheffers I1, Schmidli J2, Kroon A1, Tordoir J1, Mohaupt M2, Allemann Y2, Jordan J3,
activation. This pro-inflammatory effect of E2 in vivo is mediated by estrogen receptor
                                                                                              Engeli S3, Liebeskind U3, Luft F3, Eckert S4, Hansky B4, Cody R5, Peters T5, de Leeuw
and results, at least partly, from the inhibitory effect of E2 on the PI(3)K/Akt pathway in
                                                                                              P1
macrophages since: 1) the PI(3)K/Akt pathway prevents the pro-inflammatory cytokine           1
                                                                                                University Hospital Maastricht and Cardiovascular Research Institute Maastricht,
gene activation in response to TLR-4 activation; 2) chronic E2 treatment inhibits the
                                                                                              Maastricht, Netherlands, 2University Hospital Inselspital Bern, Bern, Switzerland,
LPS-induced activation of the PI(3)K/Akt pathway; 3) the pro-inflammatory effect of E2 is     3
                                                                                                Humbold University Charite Campus Buch, Berlin, Germany, 4Heart and Diabetes, Bad
                                                                                                                          ´
abolished by wortmannin, a PI(3)K inhibitor. In conclusion, these results indicate that
                                                                                              Oeynhausen, Germany, 5CVRx Inc., Maple Grove, USA
short-term in vitro effects do not predict the long-term in vivo effect of estrogens on
peritoneal macrophage functions. This unanticipated pro-inflammatory action of E2 on
                                                                                              Objective: The Device Based Therapy of Hypertension (DEBuT-HT) study uses an
macrophages should now be integrated in terms of physiology and taken into account in
                                                                                              implantable pulse generator and electrodes implanted bilaterally at the carotid sinus to
many pathologies influenced by sex hormones.
                                                                                              electrically activate the carotid baroreflex chronically. The long-term ( 1 year) blood
                                                                                              pressure (BP) data in a cohort of patients who underwent chronic therapy were assessed
                                                       02.06                                  to analyse if therapeutic responsiveness was sustained. Design and Method: Of 30
                                                                                              patients currently enrolled in DEBuT-HT, a cohort of 13 patients (6f / 7m, age: 51 9 yrs,
A NOVEL OPENER OF SMALL AND INTERMEDIATE CALCIUM-ACTIVATED
                                                                                              BMI: 32 7 kg/m2) from three European centers have completed 1year of chronic
K CHANNELS RELEASES NO AND CAUSES EDHF TYPE VASODILATATION                                    therapy with the device, activated one month after successful surgical implantation
IN RAT MESENTERIC ARTERIES                                                                    (Baseline). This cohort’s duration of device based treatment is currently 21.9 5 months.
                                                                                              A drop in systolic BP of at least 20 mmHg was achieved in 9 of 13 (69%) patients at 3
Simonsen U1, Krøjgaard C1, Stankevicius E1, Schjørring O1, Litvin K1, Hughes A2               months and in 10 of 13 (77%) patients at 1 year. In 6 (46%) patients 8 impulse generators
1
  University of Aarhus, Aarhus, Denmark, 2Imperial College, London, United Kingdom            have been exchanged without complications or unexpected adverse effects. The study
                                                                                              continues to enroll patients. The currently enrolled patient’s overall duration on device
The present study addressed whether NS309 (6,7-dichloro-1H-indole-2,3-dione 3-oxime)          therapy is 17 8 months. Conclusions: The Rheos system is a novel implantable device
acts on calcium-activated K channels and leads to release of nitric oxide (NO) and causes     under investigation for the chronic treatment of resistant hypertension. These first long
endothelium-derived hyperpolarizing factor (EDHF) type relaxation. Quantitative PCR           term results indicate that the response to therapy with the Rheos system is sustained and
(QPCR) and patch clamp studies were performed in human umbilical vein endothelial cells       support that such a device-based approach to blood pressure control may be a future
and vasorelaxation and release of NO studied in rat mesenteric arteries. QPCR showed          therapeutic option in patients with drug resistant hypertension.
expression of small (SKCa) and intermediate (IKCa) conductance calcium-activated K
channels in primary human umbilical vein endothelial cells, while expression of large-
conductance (BKCa) calcium-activated K channels was below the detection limit. In whole
cell patch clamp studies, NS309 (1–1000 nM) induced calcium-dependent increases in
conductance which were inhibited by apamin, a blocker of SKCa channels, charybdotoxin
and TRAM-34, blockers of IKCa channels. In segments of superior mesenteric artery, NS309                                                             03.02
increased NO concentration 12 2 nM (n 4) and evoked endothelium and concentration-            THE -LACTAM ANTIBIOTIC, CEFTRIAXON, DRAMATICALLY REDUCES
dependent relaxation. These responses were inhibited in the presence of a combination of      ACUTE STROKE MORTALITY BY STIMULATING GLUTAMATE
apamin and charybdotoxin. In pressurized mesenteric small arteries contracted with            TRANSPORTER ACTIVITY
U46619, NS309 evoked endothelium-dependent vasodilatation that was inhibited by
apamin, charybdotoxin, and TRAM-34. A combination of ADMA, apamin, and charybdo-              Namsolleck P1, Thone-Reineke C1, Krikov M1, Neumann C1, Schmerbach K1, Steckelings
                                                                                                                 ¨
toxin abolished NS309 vasodilatation. NS309 is an opener of SKCa and IKCa channels and        U1, Funke-Kaiser H1, Schefe J1, Hortnagl H1, Godes M1, Muller S2, Villringer A2, Unger
                                                                                                                                ¨                        ¨
leads to endothelium-dependent vasorelaxation associated with release of NO in rat            T 1

superior mesenteric artery, while EDHF-type vasodilatation is involved in mesenteric small    1
                                                                                                                                               ´            ¨
                                                                                                CCR, Center for Cardiovascular Research, Charite - Universitatsmedizin Berlin, Berlin,
arteries.                                                                                     Germany, 2Clinic and Polyclinic for Neurology, Charite - Universitatsmedizin Berlin,
                                                                                                                                                   ´            ¨
                                                                                              Berlin, Germany
                                                       02.07                                  Recently, -lactam antibiotics were shown to reduce neuronal damage in amyotrophic
ENHANCED PROTEASE PRODUCTION BY ADIPOCYTES CONTRIBUTES TO                                     lateral sclerosis reportedly through an increased expression of the glutamate transporter
AORTIC REMODELLING IN EXPERIMENTAL OBESITY                                                    GLT1. Since excessive release of glutamate into the interstitium is also causative for
                                                                                              neuronal damage in stroke, we tested the effectiveness of the -lactam antibiotic,
   ¨
Kruger F, Foryst-Ludwig A, Clemenz M, Sommerfeld M, Kemnitz UR, Kintscher U, Unger            ceftriaxon, on neurological outcome, infarct size, and mortality in stroke in rats.
T, Kaschina E                                                                                 Subsequently, molecular pathways leading to neuroprotection were analyzed. Male
CCR/Center for Cardiovascular Research, Berlin, Germany                                       normotensive rats were subjected to right middle cerebral artery occlusion (MCAO) for
                                                                                              90min with subsequent reperfusion. Animals received ceftriaxon (200 mg/kg bodyweight
There is a strong correlation between vascular remodelling and obesity, but the               i.p. ) as a single injection 90min after MCAO. Neurological outcome and infarct size were
mechanisms behind have not yet been clearly defined. We investigated proteolytic              estimated 48h after MCAO. Furthermore, GLT1 as well as neurotrophin (TrkB and BDNF)
processes in aorta using a mice obesity model and cell culture experiments in adipocytes.     expression in various brain regions was determined. The influence of ceftriaxon treatment
Mice (n 8) were fed with a high-fat diet (59% kcal from fat) for 12 weeks. Aortic tissues     on GLT1-promoter activity and glutamate uptake was investigated in primary rat astrocyte
were studied using histology, immunohistology and immunoblotting. Animals without diet        culture. Ceftriaxon treatment significantly improved neurological outcome and led to a
served as controls. In the fat-diet group, histological analysis revealed an aortic           dramatic reduction of acute mortality (48h) from 34.5% (in vehicle-treated rats) to 0% (in
remodelling: aortic dilatation, elastin fragmentation, cystic medial degradation and fatty    ceftriaxon-treated rats). Ceftriaxon did not alter GLT1 expression in the brain nor in primary
degeneration of the tunica media. Protein expression of proteases, cathepsin B, cathepsin     astrocytes but stimulated glutamate uptake. Furthermore, ceftriaxon induced expression of
D and MMP-3, as well as inflammatory cytokines, TNF-alpha and IL-1beta was strongly           BDNF and TrkB in the cortex. Our data provide evidence that ceftriaxon causes a significant
up-regulated in the diet group compared to controls. Immunohistochemical staining             reduction in acute stroke mortality possibly by stimulating glutamate transporter activity
showed a co-localisation of cathepsins and TNF-alpha with adipocytes in the adventitia,       and neutrophin expression in the brain. Thus -lactams may represent a novel therapeutic
suggesting that mainly these cells contribute to inflammatory processes following vascular    approach for the first line acute therapy of stroke.
                                                                                                                         2007 ECCR Oral Presentations                                  803

                                                        03.03                                   was a significant decrease in blood pressure level with the number of alleles 2. In silico
ORALLY ACTIVE AMINOPEPTIDASE A INHIBITORS REDUCE BLOOD                                          analyses suggested the presence of several transcription factor binding sites (TFBS).
PRESSURE BY BLOCKING THE BRAIN RENIN-ANGIOTENSIN SYSTEM                                         EMSAs demonstrated that the genetic variant interacts with nuclear proteins from HepG2
                                                                                                cells in an allele-specific pattern, being differentially altered by stimulation with cAMP or
ACTIVITY: A NEW STRATEGY FOR THE TREATMENT OF HYPERTENSION
                                                                                                PMA. SuperShifts showed that the region constitutes a complex enhancer module, whose
                                                                                                binding protein constituents were identified as AP-1 binding proteins, members of the
Marc Y1, Bodineau L1, Frugiere A1, Inguimbert N2, Fassot C1, Roques BP2,
                                                                                                C/EBP and IRF families. Reporter gene assays revealed that the region is a functional
Llorens-Cortes C1
1                                                                                               cis-active transcriptional unit, identifying it as a true enhancer region of the hIGF1 gene.
  UMR S 691, Inserm, College de France, Paris VI, Paris, France, 2Inserm U640, Paris,
                                                                                                Alleles displayed a significantly different, cell type-specific activity with partly dramatic
France
                                                                                                consequences for gene expression. Cotransfections proved a high specificity for the
                                                                                                functional recruitment of nuclear proteins to the enhancer. We characterized the molecular
Overactivity of the brain renin-angiotensin system (RAS) has been involved in the
                                                                                                functionality of an enhancer region of the hIGF1 promoter and identified allele-specific
development of hypertension (HTA) in several animal models, such as DOCA-salt rats, a
                                                                                                proteins responsible for the alteration in transcriptional activity introduced by a genetic
hypertension salt-dependent model. We previously reported that in the murine brain, AngII
                                                                                                promoter variant, which was highly associated with essential hypertension as well as blood
is converted to AngIII by aminopeptidase A (APA). AngIII is one of the main effector peptides
                                                                                                pressure in NT
of the central RAS in the control of blood pressure (BP). Therefore the inhibition of brain
APA, but not peripheral APA, with EC33, a specific and selective APA inhibitor leads to a
decrease in BP. Thus APA represents a potential central candidate target for the treatment
of HTA. If APA inhibitors are to be used as central antihypertensive agents, they must be
able, after oral administration, to block brain APA activity. This was achieved with RB150,
an orally active prodrug obtained by dimerization of EC33 through a disulfide bond. Thus,                                                                04.02
RB150 po administered in conscious DOCA-salt rats, crossed intestinal, hepatic and              DO DIFFERENCES IN LOWER LIMB BLOOD FLOW EXPLAIN THE ETHNIC
blood-brain barriers and inhibited brain APA activity until a value similar to that measured    DISPARITY IN PERIPHERAL ARTERIAL DISEASE INCIDENCE BETWEEN
in the brain of normotensive rats. This resulted in DOCA-salt rats but not in normotensive      SOUTH ASIANS AND EUROPEANS?
rats in a marked dose-dependent reduction in BP (ED50: 0,5mg/kg) in less than two hours
for up to several hours, without changing heart rate. In addition, this treatment increases     Alakakone A1, Tillin T1, Chambers J1, Malik I1, Coady E1, Raynor S1, Mayet J1, Wright
diuresis, which by limiting the fluid compartment, contributes to decrease BP. Thus,            A1, Kooner J1, Shore A2, Thom S1, Chaturvedi N1, Hughes A1
                                                                                                1
RB150 could constitute the prototype of a new class of central antihypertensive agents.           International Centre for Circulatory Health, St Mary’s Hospital, Hammersmith Hospital &
                                                                                                Imperial College London, London, United Kingdom, 2Peninsula Medical School, Exeter,
                                                                                                United Kingdom
                                                         03.04
THE ANGIOTENSIN AT2 RECEPTOR AGONIST COMPOUND 21 IMPROVES                                       Background: South Asians have higher rates of cardiovascular disease in comparison with
                                                                                                Europeans. Paradoxically, they experience lower levels of peripheral arterial disease.
HEART FUNCTION AFTER MYOCARDIAL INFARCTION IN THE RAT BY                                        Objectives: To determine whether differences in lower limb haemodynamics, in relation
PREVENTING APOPTOSIS AND INFLAMMATION                                                           to vessel size, flow patterns and shear stress could explain this ethnic disparity. Methods:
                                                                                                We studied a West London population-based sample of 41 European and 43 South Asian
Grzesiak A1, Kaschina E1, Foryst-Ludwig A1, Sommerfeld M1, Kemnitz UR1, Curato C1,              men without clinical evidence of coronary artery disease. Left common femoral artery
Timm M1, Hallberg A2, Li J1, Steckelings UM1, Unger Th1                                         (CFA) flow velocity and diameter were measured using ultrasound. Volumetric flow and
1
                                                                              ´
  Center for Cardiovascular Research (CCR) / Institute of Pharmacology, Charite -               shear stress were estimated using Womersley’s equation for pulsatile flow. Data are mean
Universitatsmedizin, Berlin, Germany, 2Dept. of Medicinal Chemistry, Organic
          ¨                                                                                        SD or geometric mean (95% CIs). Results: South Asians were significantly smaller in
Pharmaceutical Chemistry, Uppsala University, Uppsala, Sweden                                   size and had smaller common femoral lumen diameters than Europeans (7.59 0.98 vs.
                                                                                                8.71 1.73 mm, p 0.001); this ethnic difference persisted on adjustment for body size.
This study is the first to examine the effect of direct AT2 receptor stimulation on             Total blood flow to the leg was significantly reduced in South Asians, (100.5 (87.9,114.7)
post-infarct cardiac function by means of the non-peptide angiotensin AT2-receptor              vs. 123.5 (97.5,156.5) ml/min, p 0.04). Time-averaged wall shear stress was slightly
agonist Compound 21 (C21). MI was induced in Wistar rats by permanent ligation of the           higher in South Asian men (2.67 (2.26, 3.16) vs. 2.31 (1.94, 2.75) dyne/cm2, p 0.23).
left coronary artery. Treatment with C21 (0.03mg/kg/day i.p.) started 24 h after MI and was     Europeans experienced greater peak reverse flow velocity (-0.19 0.07 vs. -0.16 0.06
continued for 7 days. Heamodynamic measurements were performed via transthoracic                m/sec, p 0.04) and peak reverse shear (-53.8 14.2 vs. -48.5 15.0 dyne/cm2,
Doppler echocardiography (1d and 7d post MI) and intracardiac Millar catheter (7 d post         p 0.28). Conclusions: South Asians have significantly reduced CFA blood flow, possibly
MI). Cardiac tissues were studied using immunoblotting and real time RT-PCR. C21                accounting for the smaller lumen diameters. Whilst there were no significant ethnic group
significantly improved systolic and diastolic ventricular function as compared to the           differences in time-averaged wall shear stress, Europeans experienced greater reverse
placebo treated MI group: EF 69 3 vs 51 3.3%, FS 40.4 2.5 vs 27.5 2%, E/A 2 1.3                 flow velocities. Reverse flow is less athero-protective than forward flow and this may
vs 6.5 0.5, DT 22 1 vs 14 1 ms, LVEDP 11.2 2 vs 19.6 2.5 mmHg, cardiac                          explain the increased risk of peripheral atherosclerosis in Europeans.
contractility index 98 3.6 vs 66 3.3 1/s, maxdP/dt 6803 312 vs 3726 168 mmHg/s.
Regarding putative underlying mechanisms, C21 diminished MI-induced Fas-ligand and
caspase-3 expression in the peri-infarct zone indicating an anti-apoptotic effect. Phospho-
p42/44 and Phospho-p38 MAPK - both involved in the regulation of cell survival - were
strongly reduced after MI, but almost completely rescued by C21 treatment. Furthermore,
                                                                                                                                                       04.03
C21 decreased MI-induced serum MCP-1 (1.8-fold) and myeloperoxidase (2.3-fold) as well
as cardiac IL-6, IL 1-beta and IL-2 pointing to an anti-inflammatory effect. We conclude        EPROSARTAN REDUCES THE RESPONSE TO COLD PRESSOR TESTING IN
that direct AT2 receptor stimulation may be a novel therapeutic approach to improve             HEALTHY HUMANS
post-infarct systolic and diastolic function by anti-apoptotic and anti-inflammatory
mechanisms.                                                                                     Vase H, Lauridsen TG, Bech JN, Pedersen EB
                                                                                                Department of Medical Research, Holstebro Hospital, Holstebro, Denmark

                                                                                                Considerable interactions exist between the sympathetic nervous system (SNS) and the
                                                         04.01                                  renin-angiotensin-aldosterone system. Angiotensin II infusion facilitates activity in the SNS.
ALTERATIONS OF THE TRANSCRIPTIONAL ACTIVITY OF THE HUMAN IGF1                                   In animals studies angiotensin II receptor (AT1) antagonists have been shown to inhibit the
PROMOTER CAUSED BY A SNP ARE ASSOCIATED WITH HYPERTENSION                                       activity of the SNS. In humans studies conflicting results exist regarding this effect of AT1
AND BLOOD PRESSURE                                                                              antagonism. We tested the hypothesis that the AT1 antagonist eprosartan would inhibit the
                                                                                                activation of the SNS induced by a cold pressor test (CPT). The effect of eprosartan was
Telgmann R1, Brand E2, Dordelmann C1, Nicaud V3, Ru mann C1, Cambien C3, Tiret L3,
                              ¨                         ¨                                       measured before, during and after a CPT in a randomized, placebo controlled, double
Paul M4, Brand-Herrmann S-M1                                                                    blinded, crossover study in 17 healthy volunteers. Eprosartan reduced the increase in heart
1
  Leibniz-Institute for Arteriosclerosis Research, Molecular genetics of Cardiovascular         rate HR by 3.9 (39 %, P 0.05) beats/min. The initial increase in mean arterial pressure
Disease, Munster, Germany, 2Internal Medicine D, University Clinic Munster, Munster,
             ¨                                                           ¨        ¨             (MAP) was unaffected by eprosartan, however the increase in MAP after 2 minutes of CPT
Germany, 3Inserm U525, Paris, France, 4Charite, University Medicine Berlin, Berlin,
                                                  ´                                             was reduced by 4.7 mm Hg (30 %, P 0.05) by eprosartan. Eprosartan caused a more
Germany                                                                                         pronounced reduction in free water clearance (CH20) by 1.3 ml/min (51 %, P 0.01) while
                                                                                                fractional excretion of sodium (FENa), glomerular filtration rate, plasma vasopressin (AVP)
IGF1 is a pleioptropic growth factor exerting important functions in the cardiovascular         and urinary excretion of aquaporin-2 (AQP-2) were unchanged. The increase in plasma
system. A well-characterized case-control study for essential hypertension (PEGASE:             noradrenaline was unaffected by eprosartan. We conclude that eprosartan reduces the
n 745 hypertensives [HT]; n 769 normotensives [NT]) was genotyped for a common                  cardiovascular response to a CPT, indicating a reduced activation of the SNS. This was
hIGF1 promoter variant (major allele [1], minor allele [2]). There was a significant            accompanied by a more pronounced reduction in CH20 without changes in FENa, plasma AVP
difference in genotype frequency between HT and NT (P 0.019), with an odds ratio for            or urinary AQP-2 excretion, suggesting an increase in non-AQP-2 mediated water
hypertension of 0.73 ([95%CI 0.58 – 0.91], P 0.006) associated with allele 2. In NT there       reabsorption.
804           Hypertension Vol 50, No 4 October 2007


                                                                                       04.04     vasoreactivity to Ang1–7 in patients on a low sodium diet or in patients having a renal
RECIPROCAL CONSOMIC STRAINS TO EVALUATE PHENOTYPIC                                               artery stenosis. Conclusions: Ang1–7 induces renal vasodilation in hypertensive patients.
IMPORTANCE OF CHROMOSOME 13 IN LYON HYPERTENSIVE RATS                                            Simulation of an activated RAS results in similar Ang1–7-induced renal vasodilation.
                                                                                                 Hence, our results do not support the concept of Ang1–7 being an endogenous antagonist
Gilibert S1, Kwitek AE2, Nussberger J3, Sassard J1, Bataillard A1                                of AngII but rather point towards a separate role which is independent from AngII.
1
  Universite de Lyon, Lyon, France, 2Medical College of Wisconsin, Milwaukee, WI, USA,
           ´
3
  Centre Hospitalier Universitaire Vaudois, Lausanne, Switzerland
                                                                                                                                                                                      W1.03
Lyon hypertensive (LH) rats exhibit spontaneous salt sensitive hypertension, marked              HEPARIN IMPAIRS GLYCOCALYX BARRIER PROPERTIES AND
proteinuria and signs of metabolic syndrome: overweight, hyperlipidemia and increased            ATTENUATES SHEAR DEPENDENT VASODILATATION IN MICE
insulin/glucose ratio. A full genome scan showed the existence, on chromosome 13, of
Quantitative Trait Loci (QTLs) associated with blood pressure (BP) and kidney renin              VanTeeffelen JWGE, Brands J, Vink H
concentration. In order to determine their influence, we generated reciprocal consomic           Maastricht University, Maastricht, Netherlands
strains: LH-13BN i.e. LH rats in which chromosome 13 has been fully substituted by a
Brown Norway (BN) chromosome 13; conversely, BN-13LH i.e. BN normotensive rat with a             The glycocalyx is a hydrated mesh of polysaccharides and adsorbed plasma proteins
chromosome 13 from LH rats. The characterisation of these strains included radio-                which forms the true interface between the flowing blood and the endothelium. We
telemetric measurement of BP during normal and elevated salt intake as well as the               hypothesized in the present study that competitive binding of heparin to glycocalyx
determination of renal, metabolic and morphological parameters. In comparison to LH              associated proteins would affect its barrier properties and the mechanotransduction of
parents, LH-13BN rats showed increased body weight and markedly decreased mean BP                shear stress to the endothelium. In anaesthetized C57Bl/6 mice, clearance of 70 kDa
(-13 mmHg) and of proteinuria and lipids. Differences between BN-13LH and BN rats are            dextrans from the circulation was increased (P 0.05 versus saline) 1 hour after 1.25 U of
much smaller than those observed between LH-13BN and LH rats. Plasma renin activity is           heparin, as well as after glycocalyx degradation with 40 U of hyaluronidase (amount
not affected by the substitution of chromosome 13. In conclusion, the present work               cleared in 30 min after saline: 11 5%, after heparin: 45 8%, after hyaluronidase:
demonstrates that the QTLs found on chromosome 13 are of functional importance. It               30 3%). Clearance of 40 kDa dextrans increased (P 0.05 versus saline) to a lesser
shows that chromosome 13 is a major determinant of BP level in LH rats and confirms that         extent after both treatments (saline: 46 3%; heparin: 60 5%; hyaluronidase: 60 2%).
this effect is independent of renin activity. Therefore, these chromosome 13 consomic rats       The dilator response of second-order arterioles in cremaster muscle during reactive
may well be used to generate overlapping congenic rats in order to approach the genes            hyperaemia was reduced for up to 90 min after heparin as reflected by a decrease
possibly involved in alterations shown by LH rats.                                               (P 0.008) in t50 of dilator response, and this effect was associated with a diminished nitric
                                                                                                 oxide bioavailability. Hyaluronidase resulted in reductions (P 0.05) in baseline and peak
                                                                                                 reactive hyperaemic diameter, while, despite an increase in wall shear rate at the
                                                        04.05                                    beginning of reactive hyperaemia, t50 of dilatation was not affected. In conclusion, our data
                                                                                                 in mice show that a heparin challenge is associated with increased vascular leakage of
RENAL TUBULOINTERSTITIAL, VASCULAR AND GLOMERULAR FIBROSIS                                       dextrans and impaired arteriolar vasodilatation during reactive hyperaemia. Our data
ARE DIFFERENTLY REGULATED BY RENIN ANGIOTENSIN AND                                               suggest that protein-heparan sulphate interactions are important for a functional
ENDOTHELIN-1 SYSTEMS IN A MODEL OF ANGII-DEPENDENT                                               glycocalyx.
HYPERTENSION
Seccia TM, Maniero C, Belloni AS, Pessina AC, Rossi GP                                                                                                W1.04
                                `
DMCS-Clinica Medica 4, Universita di Padova, Padova, Italy                                       HUMAN GLOMERULAR ENDOTHELIAL GLYCOCALYX CONTRIBUTES TO THE
                                                                                                 PERMEABILITY BARRIER TO PROTEIN IN VITRO
Activation of the renin angiotensin system (RAS) and endothelin(ET)-1 system promotes
renal damage via not well known mechanisms. We investigated the effects of RAS and/or            Singh A1, Satchell SC1, McKenzie EA3, Neal CR2, Tooke JE4, Mathieson PW1
ET-1 blockade on the vascular, tubular and glomerular in the kidney of a model of severe         1
                                                                                                   Academic Renal Unit, University of Bristol,, Bristol, United Kingdom, 2Microvascular
Ang II-dependent hypertension, (mRen2)27 transgenic rat [TGRen2]. 5-week-old TGR rats            Research Labs, University of Bristol,, Bristol, United Kingdom, 3Faculty of Life Sciences,
were randomly assigned to one of the following treatments: 1) bosentan (n 5), 2)                 University of Manchester,, Manchester, United Kingdom, 4Department of Diabetes and
BMS-182874, a selective antagonist of ETA subtype receptor (BMS; n 4) 3) irbesartan              Vascular Medicine, Peninsula Medical School,, Exeter, United Kingdom
(n 5) 4) irbesartan combined to BMS (n 4); 5) nifedipine (n 5); 6) placebo (n 8). After
4 weeks collagen was measured on kidney 5 m-sections stained with Sirius-red using               Glycocalyx coats the luminal surface of the glomerular capillaries and is composed of
Leica photomicroscopy. Type I and III collagens were identified under polarization               glycoproteins including proteoglycans. Human Heparanase (HPSE) degrades heparan
microscopy. Blood pressure (BP) and plasma aldosterone levels were measured by tail-cuff         sulphate glycosaminoglycans and is upregulated in proteinuric states. In this study, we
method and RIA. A significant decrease in BP and plasma aldosterone was observed in              characterise the structure of the human glomerular endothelial cell (GEnC) glycocalyx, and
irbesartan-treated rats. Vascular fibrosis was significantly reduced after bosentan, BMS         examine its functional relevance especially after treatment with recombinant HPSE. We
and nifedipine treatments, with no change in collagen I/III ratio. A significant reduction in    used conditionally immortalised GEnC developed in our laboratory. The ultrastructure of
tubulointerstitial fibrosis was measured after bosentan and irbesartan, whereas glomerular       glycocalyx was analysed by electron microscopy (EM) and confocal microscopy using
fibrosis was reduced after BMS-treatment. Conclusions. 1) Hypertension and renal                 fluoroscein-wheat germ agglutinin (WGA). Removal of glycocalyx by neuraminidase,
fibrosis are regulated by different mechanisms. 2) In contrast to tubulointerstitial fibrosis,   bacterial Heparinase III and HPSE was assessed using WGA and an anti-heparan sulphate
which is modulated by both RAS and ET-1 system, glomerular and vascular fibrosis are             antibody. Barrier properties of GEnC monolayers were studied by trans-endothelial
mostly regulated by ET-1 system.                                                                 electrical resistance (TEER) and passage of labelled albumin. EM revealed a 200nm thick
                                                                                                 glycocalyx over the plasma membrane of GEnC which was further demonstrated by WGA.
                                                                                                 Neuraminidase treatment removed the majority of glycocalyx which functionally corre-
                                                         04.06                                   sponded to reduced TEER by 59% and increased albumin flux by 207%. Heparinase III and
ANGIOTENSIN 1–7 INDUCES RENAL VASODILATION IN HYPERTENSIVE                                       HPSE specifically cleaved HS: this caused no change in TEER, but increased the albumin
PATIENTS INDEPENDENT OF AN ACTIVATED RENIN-ANGIOTENSIN                                           passage across the monolayers by 40% and 39% respectively. These results are
SYSTEM                                                                                           consistent with an important role for the GEnC glycocalyx in restriction of glomerular
                                                                                                 protein passage in vivo and suggest ways in which HPSE levels may be linked to
Mostard GJM, Houben AJHM, Kroon AA, van Engelshoven JMA, de Leeuw PW                             proteinuria in clinical disease.
University Hospital Maastricht, Maastricht, Netherlands

Introduction: Animal studies suggest a counterregulatory role of Angiotensin 1–7                                                                           W2.03
(Ang1–7), acting as an endogenous antagonist of Angiotensin II (AngII). The aim of the           IDENTIFICATION OF PROTEASE INHIBITORS KININOGEN AND TIMP-1 AS
present study was to evaluate the vascular effects of Ang1–7 in the human hypertensive           NOVEL MARKERS OF EARLY-PHASE CEREBRAL ARTERIOGENESIS
kidney in patients with a varying degree of activation of their renin-angiotensin system
(RAS). Design & Methods: In 77 hypertensive patients, scheduled for renal angiography,           Hillmeister P1, Lehmann K1, Bondke A1, Witt H2, Duelsner A1, Gotthold C1, Gruber C3,
selective renal blood flow (RBF) was measured using the 133Xenon washout technique               Ruiz-Noppinger P2, Buschmann IR1
                                                                                                 1
before and during intrarenal infusion of Ang1–7 (three consecutive doses: 0.27– 0.9 –2.7                                                               ´          ¨
                                                                                                   CCR – Center for Cardiovascular Research, Charite – Universitatsmedizin Berlin, Dept.
ng/kg/min). Local RAS stimulation was mimicked by co-infusion of AngII ( 0.3 ng/kg/min),         of Internal Medicine/ Cardiology, Berlin, Germany, 2CCR – Center for Cardiovascular
prior to and in parallel with Ang1–7 infusion (n 34). In addition, analyses of Ang1–7            Research, Max-Planck-Institute for Molecular Genetics, Berlin, Germany, 3Charite –
                                                                                                                                                                                  ´
reactivity were made in a subgroup of patients on low sodium diet, and in a subgroup of                   ¨
                                                                                                 Universitatsmedizin Berlin, Institute of Integrative Neuroanatomy, Department of
patients with renal artery stenosis (both conditions of activated RAS). Finally, renal           Anatomy, Berlin, Germany
angiography was performed. Results: Ang1–7 caused vasodilation in all groups. Maximal
dilation after Ang1–7 was not significantly different in the group with than in the group        This study aims at a comprehensive understanding of the transcriptional activation during
without pre-constriction with AngII. Subanalyses revealed no differences in renal                early phase collateral vessel growth in a rat model of cerebral adaptive arteriogenesis.
                                                                                                                         2007 ECCR Oral Presentations                                  805

While cerebral arteriogenesis constitutes a promising therapeutic concept for cerebrovas-                                                              W3.04
cular ischemia, transcriptional profiles important for therapeutic target identification have   ROS GENERATION IN RELATION TO ADD1 GLY460TRP POLYMORPHISM
not yet been studied. Using a 3 vessel occlusion (3-VO) rat model of non-ischemic cerebral
hypoperfusion, collateral material from the posterior cerebral artery region (PCA) was          Barlassina C1, Ruello A1, Tripodi G2, Azimonti S2, Torielli L2, Del Vecchio L1, Cusi D1
isolated without contamination of adjacent tissue. We detected differential expression          1
                                                                                                  University of Milano, Dept Sciences & Biomedical Technologies, Milano, Italy, 2Prassis,
particularly 24h post 3-VO with 164 genes significantly deregulated (91 upregulated and         Sigma-Tau Research Institute, Settimo Milanese, Milano, Italy
73 downregulated). Expression patterns contain gene transcripts predominantly involved in
proliferation, inflammation and migration, with three protease inhibitors T-kininogen, Alpha    Background: We showed that human skin fibroblasts bearing one 460Trp allele of
2-macroglobulin and TIMP-1 strongly upregulated. These results were confirmed by                a-adducin (ADD1), the Na sensitive one, display compensatory (?) over-activity of the
quantitative real time RT-PCR. In-situ hybridisation 24h post 3-VO localized expression of      antioxidant system (ECCR 2004). Aim: to extend the observation on the relationship
TIMP-1 and T-kininogen to collateral arteries, while immunhistological staining for PCNA        between ADD1 Gly460Trp polymorphism and ROS system in cells with ADD1 cDNAs
demonstrated proliferative activation, thus resulting in a significant diameter increase        transfected with the normotensive (Gly) or hypertensive (Trp) allele. Methods: Human HK2
(30%) of the PCA as detected 7 days post 3-VO, confirming the arteriogenic phenotype.           cells stably transfected with Gly460-Ser586 or Trp460-Cys586 human ADD1 cDNA.
Furthermore morphological activation of the endothelium was detected by scanning                Intracellular ROS measured with 2’7’-DCFH-DA as a fluorescent probe in HK2 at
electron microscopy. Based on this expression analysis, putative targets for novel              confluence after 24h serum deprivation. Measurements in basal conditions, with 500 M
therapeutic strategies to stimulate cerebral arteriogenesis may be identified.                  Angiotensin II (AII), 6 M Insulin (INS) for 20’, with 10 M DPI (NADPH OX inhibitor) for 30’.
                                                                                                Results: Mean basal ( 49%; p 0.023) ROS production was higher in 460Trp clones and
                                                                                                remained higher after AII ( 35%; p 0.011). However AII stimulated ROS production
                                                                                                equally Gly460 and 460Trp clones. INS stimulated substantially ROS production ( 160%
                                                                                                p 0.013) in 460Trp clones. ROS inhibition by DPI was not different, indicating that most
                                                                                                of the difference in ROS production was not mediated by NADPH oxidase. and NADPH
                                                                                     W2.04
                                                                                                oxidase mediated ROS production. Conclusion: The hypertensive allele is associated with
PHARMACOLOGICAL PROPERTIES OF ENDOTHELIUM-DEPENDENT                                             larger basal and insulin induced ROS production. This may provide the clinical connection
VASODILATATION IN RAT MESENTERIC COLLATERAL ARTERIES                                            to hypertensive patients carrying the 460Trp allele, who are Na-sensitive and more prone
                                                                                                to develop cardiovascular complications.
Hilgers RHP, DeMey JGR
Cardiovascular Research Institute Maastricht, Maastricht, Netherlands
                                                                                                                                                                                      05.01
Background: Wall shear stress (WSS) is a major determinant of arterial structure and            STATUS OF RAT CLONING
function, yet, collateral arteries are present at several strategic locations of the vascular
system. In the rat mesentery, findings by Smiesko et al. (1989) suggest that in collateral      Popova E, Bader M, Krivokharchenko A
arteries (COLL) WSS might be oscillatory and is on average 75% smaller than in the arterial               ¨
                                                                                                Max-Delbruck-Center, Berlin, Germany
segments where they interconnect upstream fourth-order mesenteric arteries (MA4).
Hypothesis: Endothelium-dependent vasodilator mechanisms are upregulated in pre-                Cloning technology would allow targeted genetic alterations in the rat, a species which is
existing collateral arteries. Methods: Endothelium-dependent and –independent isometric         very instrumental for cardiovascular research but yet not accessible for such methods due
tension responses were compared in COLL and MA4, isolated from the rat mesenteric               to the lack of germ-line competent embryonic stem cells. The present study was
arterial bed. Results: Lumen diameter (10%) and maximal contractile responses (30%)             performed to examine the developmental ability of nuclear transfer (NT) embryos derived
were significantly smaller in COLL. Effects on ACh-induced relaxation during agonist or         from embryonic blastomeres (embryonic cell NT) or cumulus cells (somatic cell NT). We
K -induced contraction did not differ between COLL and MA4 for (1) the NO-synthase              developed an efficient method for the production of rat embryos by somatic cell NT
inhibitor L-NAME, (2) the COX inhibitor indomethacin, (3) non-selective blockade of             including enucleation without chemical treatment and electrofusion. The efficiency of
calcium-activated K (KCa) channels (by 40 mmol/L K ), and (4) selective blockade of SKCa-       embryo reconstruction was significantly higher (75.9%) after slitting the zona pellucida and
(by UCL1684) and IKCa (by TRAM-34), alone or in combination. Conclusion: The low and            than pressing the oocyte with a holding pipette compared to the conventional protocol
oscillatory WSS in pre-existing collateral arteries does not result in alterations of the       using aspiration of the metaphase-plate with an enucleation pipette (41.8%). When
contributions of NO, EDHF and (possibly) myo-endothelial coupling to pharmacological            karyoplasts from blastomeres were used for the reconstruction of embryos, highest in vitro
stimulation of endothelium-dependent vasodilatation. Other mechanisms might be respon-          cleavage rates were obtained with nuclei in an early phase of the cell cycle transferred into
sible for the maintenance of the patency and vasomotor responses of pre-existing                enucleated pre-activated oocytes or zygotes. However, further in vitro development of
collateral arteries.                                                                            reconstructed embryos produced from blastomere nuclei or cumulus cells was arrested at
                                                                                                early cleavage stages under all conditions tested in this study. Immediate transfer of
                                                                                                reconstructed embryos with nuclei from 2-cell embryos at an early stage of the cell cycle
                                                                                                in pre-activated enucleated oocytes, but not of somatic cell NT embryos, into recipients
                                                                                                resulted in production of live offspring with a general efficiency of 0.4 –2.2 %. Future
                                                      W3.03                                     efforts will focus on optimizing the production of somatic and embryonic cell NT rat
                                                                                                embryos.
FUNCTIONAL ANALYSES OF MOLECULAR HAPLOTYPES OF THE HUMAN
OSTEOPROTEGERIN GENE PROMOTER

Hagedorn C1, Telgmann R1, Brand E2, Doerdelmann C1, Nicaud V3, Kurtz S4,
Schoenfelder J4, Baumgart P5, Kleine-Katthoefer P6, Cambien F3, Paul M4,
Brand-Herrmann SM1
1
  Leibniz-Institute for Atherosclerosis Research, Muenster, Germany, 2University Clinic of                                                                                            05.02
Muenster, Muenster, Germany, 3Inserm U525, Faculte de Medecine Pitie-Salpetriere,
                                                      ´       ´          ´      ˆ `             WNK1, A NEW PLAYER IN VASCULAR REACTIVITY ?
Paris, France, 4Charite, University Medicine Berlin, Berlin, Germany, 5Clemens hospital,
                        ´
                                                    6
Clinic of Internal Medicine I, Muenster, Germany, St. Franziskus hospital, Medical              Bergaya S1, Faure S2, Achard JM2, Jeunemaitre X1, Hadchouel J1
                                                                                                1
Clinic III, Muenster, Germany                                                                     Inserm U772 - College de France, Paris, France, 2Department of Physiology - Limoges
                                                                                                University Hospital, Limoges, France
Osteoprotegerin (OPG), a member of the TNF receptor superfamily, plays an emerging role
in cardiovascular disease (CVD); some studies reported associations between OPG SNPs            WNK1 is a serine-threonine kinase of which mutations cause Familial Hyperkaliemic
and CVD phenotypes including plasma OPG levels. By use of OPG gene scanning in 190              Hypertension, a rare inherited form of hypertension. WNK1 gene encodes for two proteins:
chromosomes from high-risk MI patients, we identified 2 exonic, 3 intronic and 5 variants       a short kidney-specific isoform, and a full-length isoform (L-WNK1) expressed ubiquitously
in the 5’-regulatory gene portion (T-957C, A-943G, G-897A, T-861G, T-156C). After               and notably within the vascular system. The vascular role of L-WNK1 has not been yet
resequencing of 64 additional chromosomes, we identified three common distal molecular          investigated. We used heterozygous mice bearing a constitutive inactivation of L-WNK1
haplotypes by subcloning procedures (OPG-wt, OPG-Haplo2 [-943G], OPG-Haplo4 [-957C,             (L-WNK1 /-) and their littermates controls (L-WNK1 / ) in order to perform in vitro
-943G, -897A and -861G]), which we introduced into the reporter gene vector pGL3prom.           pharmacological studies on thoracic aorta rings mounted on a wire-myograph system. Our
Transient transfection analyses in SaOs2 under basic and stimulatory conditions with            data show that phenylephrine-induced contractions and acetylcholine-induced relaxations
TNF (2 ng/mL; 6hrs), 17 -Estradiol (E2) (10-8M; 24hrs), and both, revealed that                 are significantly reduced in L-WNK1 /- mice compared to their controls L-WNK1 /
stimulation with E2 and E2/TNF resulted in a mildly but significantly lesser transcriptional    (P 0.0001 and P 0.0005, respectively) while potassium chloride contractions remain
activity of OPG-Haplo2 (P 0.023 and P 0.028, respectively). Irrespective of stimulation,        unaffected (P 0.41). Blockade of calcium-activated potassium channels (KCa), using the
OPG-Haplo4 consistently displayed a significantly higher activity compared to OPG-wt            non-selective antagonist TEA, restores phenylephrine and acetylcholine responses in
(P 4.88x10-6); both results were confirmed in kinetic analyses over a 14h period. We            L-WNK1 /- to a level similar to L-WNK1 / (P 0.76 and P 0.78, respectively).
characterized the functionality of molecular OPG promoter haplotypes under different            Furthermore, quantitative PCR performed on thoracic aorta evidences that the beta subunit
stimulatory conditions and suggest that gene expression and transcriptome analyses are          of the big-conductance potassium channels (BKCa) expression is significantly reduced in
now warranted to show their differential respective impacts on CVD phenoptypes.                 L-WNK1 /- compared to L-WNK1 / (49% decrease, P 0.01), while the SK2 isoform
806          Hypertension Vol 50, No 4 October 2007


of the small-conductance potassium channels (SKCa) expression is increased (85%               administered before the development of overt hypertension, is able to prevent vascular
increase, P 0.04) and the expression of SK1 and SK3 isoforms of SKCa channels and the         remodelling and mechanical alterations typical for this condition. This was tested in the
intermediate-conductance potassium channels (IKCa) remain unchanged (P 0.4). Our              present study. Materials and methods: Twenty-four 4 weeks old rats were included in the
results suggest that L-WNK1 plays an important role in vascular G-protein-coupled             study, 12 SHR and 12 normotensive Wistar-Kyoto (WKY) rats. Half were implanted with a
receptors-mediated contractions and relaxations, through the involvement of calcium-          subcutaneous osmotic minipump that delivered 60 U/day of heparin for a period of 8
activated potassium channels.                                                                 weeks. Mesenteric resistance arteries were harvested and mounted on a pressure
                                                                                              myograph. Pressure/diameter (P/D) curves from 20 to 130 mmHg were constructed in the
                                                                                              absence of tone and wall-to-lumen ratio (W/L) was determined at a transmural pressure
                                                                                              of 80 mmHg. Results: There was a significant (p 0.003) upward shift of the P/D curve
                                                                                              in treated SHR (HepSHR) compared to control SHR (CntSHR). No significant difference was
                                                                                              detected between treated (HepWKY) and control WKY, although a trend was observed. Wall
                                                        05.03                                 to lumen ratio was significantly smaller in treated SHR compared to the control group
DOWN REGULATION OF L-TYPE CALCIUM CHANNEL EXPRESSION IN RAT                                   (HepSHR 0,061 0,012 vs CntSHR 0,082 0,012; p 0.02) and similar to control WKY
SMALL MESENTERIC ARTERIES WITH THE SPECIFIC SIRNA                                             (cntWKY 0,067 0,025; p NS). No significant difference was found between treated and
TRANSFECTION IN VIVO                                                                          untreated WKY. Conclusion: Chronic treatment at young age with a low dose of heparin
                                                                                              prevents the development of resistance artery structural changes in SHR
Matchkov VV, Larsen P, Briggs Boedtkjer DM, Kold-Petersen H, Nilsson H, Aalkjaer C
Institute of Physiology and Biophysics, University of Aarhus, Aarhus, Denmark

Ca2 entry via L-type Ca2 channels (Cav1.2) is a key factor in regulation of excitation-                                                                 05.05
contraction coupling in smooth muscle cells (SMCs). Previous gene deletion studies have       A POSITIVE FAMILY HISTORY IDENTIFIES SUBJECTS PRONE TO DEVELOP
provided insight into the critical role of pore-forming 1C subunit in regulation of blood     RECURRENT THROMBOTIC EVENTS AFTER A FIRST PREMATURE
pressure level. Homozygous knockout is, however, lethal and this limitation can be            THROMBOTIC EVENT
overcome by transient downregulation with siRNA. The branches of mesenteric vascular
bed in anesthetised Wistar rats were transfected with siRNA directed against 1CCav1.2 or      Mulders T1, Meijer Z1, van der Donk C1, Kroon AA1, Ferreira I2, Stehouwer CDA1,
with control scrambled siRNA. The effect of transfection was analyzed after 3 and 10 days     Pinto-Sietsma SJ1
using quantitative PCR and immunohistochemistry. The functional effects of transfection       1
                                                                                                Academic Hospital Maastricht, Maastricht, Netherlands, 2University Maastricht,
were studied using isometric myography. Specific transfection downregulates the mRNA
                                                                                              Maastricht, Netherlands
level of Cav1.2 by 93 2% within 3 days. Surprisingly, normalized internal diameter of
Cav1.2-siRNA transfected arteries was significantly reduced by 33 10% (n 7) vs.
scrambled siRNA transfected arteries. The transfection did not change the sensitivity to      Introduction: Cardiovascular disease (CVD) consists of atherosclerotic and thrombotic
K -depolarization but the maximal response was significantly reduced by 72 10% (n 6).         factors. The importance of thrombotic factors in CVD is still a matter of debate, since
Cav1.2-siRNA transfected arteries were less sensitive to agonist stimulation and had          several studies have not convincingly shown a positive relation. We therefore wondered
reduced responses to caffeine. The responses to K -depolarization and agonist-                whether these factors might play a much larger role in families with CVD. Hypothesis: In
stimulation in arteries transfected with Cav1.2-siRNA were upregulated in comparison with     families with CVD recurrent thrombotic or atherosclerotic events occur among individuals
the controls after 10 days. This upregulation was not, however, nifedipine-sensitive. Using   with a first premature event of the same nature. Methods: We retrospectively investigated
for first time in vivo transfection of arteries with siRNA, we demonstrated the importance    187 subjects with premature CVD (men or women with a CV event before the age of 50
of Cav1.2 for vascular reactivity and its tight coupling with other cellular Ca2 handling     or 55 respectively). A thrombotic event was defined as a myocardial infarction, acute
processes.                                                                                    coronary syndrome (ACS), stroke or transient ischemic attack without any carotid
                                                                                              abnormalities. An atherosclerotic event was defined as all CVD without thrombotic events.
                                                                                              A positive family history was defined as a first-degree family member with CVD before the
                                                       05.04                                  age of 55 for men or 60 for women. The data was analyzed with multiple logistic
CHRONIC TREATMENT WITH HEPARIN PREVENTS RESISTANCE ARTERY                                     regression. Results: Among subjects with a first premature thrombotic event, a positive
REMODELLING IN YOUNG SPONTANEOUSLY HYPERTENSIVE RATS                                          family history was associated with recurrent thrombotic event(s) [age and sex-adjusted
                                                                                              odds ratio (OR): 5.8; 95% confidence interval (CI): 1.6 –21.7], whereas among subjects
Boari GEM1, Schiffers PMH2, Agabiti Rosei E1, De Mey JG2                                      with a first premature atherosclerotic event there was no association with recurrent
1
  Department of Internal Medicine - University of Brescia, Brescia, Italy, 2Department of     atherosclerotic event(s) (OR: 1.5; 0.7–3.2). Conclusion: A positive family history in
Pharmacology and Toxicology - University of Maastricht, Maastricht, Netherlands               subjects with a first premature thrombotic event increases the likelihood of recurrent
                                                                                              thrombotic event(s), whereas it does not in subjects with a premature atherosclerotic
Background: There is strong evidence that chronic heparin treatment lowers blood              event. These findings suggest that a genetic defect within the thrombotic system may play
pressure in spontaneously hypertensive rats (SHR). It is not known whether this treatment,    a role in families with premature thrombotic disease.
                                                                                                                    2007 ECCR Poster Presentations                                   807

ECCR Poster Presentations                                                                     related cleavage fragment of PARP. Moreover, TGF- 1 stimulation of fibroblasts signifi-
                                                                                              cantly elevated collagen type III and OPN protein synthesis (15 min: 1.2-fold, both and 2 h:
                                                                                              1.4-fold/1.8-fold) compared to controls. These results indicate that in cardiac fibroblasts
                                                        PA.01                                 cultured from the infarct region, synthesis of ECMP is enhanced compared to controls.
                                                                                              Gene expression and protein synthesis of OPN is clearly upregulated. Thus, OPN and
CHRONIC CARDIOTROPHIN-1 ADMINISTRATION PROMOTES A DISTINCT
                                                                                              TGF- 1 may play a key role in collagen deposition during the early phases of myocardial
PATTERN OF CARDIAC REMODELLING IN NORMOTENSIVE AND                                            remodeling post-MI.
SPONTANEOUSLY HYPERTENSIVE RATS

 ˜        ´         ´          ˜
Inigo C, Lopez N, Dıez J, Fortuno MA                                                                                                                  PA.04
Division of Cardiovascular Sciences, Centre for Applied Medical Research, University of
                                                                                              LEFT ATRIAL VOLUME AS A BIOMARKER OF DIASTOLIC DYSFUNCTION IN
Navarra, Pamplona/Navarra, Spain
                                                                                              HYPERTENSIVE PATIENTS
We have previously reported that cardiotrophin-1 (CT-1), an IL-6 family member cytokine,
induces a different morphological and molecular pattern of hypertrophy in cardiac             Gianni M, Maresca AM, Nicolini E, Mongiardi C, Monza M, Dentali F, Grandi AM
myocytes isolated from normotensive and genetically hypertensive rats (SHR). To                        `
                                                                                              Universita Degli Studi dell’Insubria, Varese, Italy
investigate whether this effect is also present in vivo, recombinant rat CT-1 (20 g/kg) or
vehicle was daily administrated IP to male Wistar (n 3/group) and SHR rats (4 –5/group)       Background and Aim: Left ventricular (LV) diastolic dysfunction is common in hyperten-
along 21 days. Left ventricular chamber diameter (LVCD), left ventricular wall thickness      sive patients. Aim of the study was to assess the possible role of left atrial volume (LAV)
(LVWT) and cellular size were assayed by automatic image analysis. Myocardial protein         in the diagnosis of diastolic dysfunction in essential hypertension. Methods: We enrolled
expression was measured by Western Blot. In Wistar rats, CT-1 increased LVCD (45%,            43 consecutive hypertensive patients (32 men; age 52 7 years) with no history of atrial
p 0.011), decreased LVWT (10%, p 0.04) and promoted longitudinal enlargement of               fibrillation or valvular heart disease. By means of echocardiography we evaluated LAV (by
cardiomyocytes (20%, p 0.019) compared to vehicle. These changes were associated              biplane area-length method) indexed for body surface area, LV mass and LV diastolic
with the increase of all contractile proteins determined except MLC-2v and -skeletic          function (using Doppler transmitral flow: E/A ratio, deceleration time; Tissue Doppler
actin. On the contrary, in SHR, CT-1 decreased LVCD (14%, p 0.047), increased LVWT            Imaging of mitral and septal annulus motion, isovolumetric relaxation time). Results: LV
(15%, p 0.036) and induced cardiomyocyte growth predominantly in the transversal axis         hypertrophy (LVH) was found in 31 pts (13 with concentric LVH, 12 with eccentric LVH, 6
(10%, p 0.016). In this case, CT-1 increased the expression of all contractile proteins       with concentric remodelling). Ejection fraction was normal in all. Twenty-eight patients had
assayed, including MLCV-2 (45 %, P 0.016) and -skeletic actin (50%, P 0.046). In              diastolic dysfunction (19 abnormal relaxation, 9 patients pseudonormal pattern). LVH was
conclusion, chronic CT-1 administration induces dissimilar cardiac morphological and          greater in patients with diastolic dysfunction (120.5 21.5 g/m2 vs 95.7 15.6 g/m2 ,
molecular changes in rats depending on their phenotype: eccentric remodelling in              p 0.001). The indexed LAV was significantly smaller in patients with normal diastolic
normotensive Wistar rats and concentric hypertrophy in SHR.                                   function than in patients with diastolic dysfunction (25.2 8 ml/m2 vs 34.4 9 ml/ m2; p
                                                                                              0.002). In a multivariate analysis, indexed LAV was associated with diastolic function grade
                                                                                              (p 0.01), independently of age and LV mass. Conclusion: In essential hypertension LAV
                                                        PA.02                                 appears to be related to LV diastolic function and therefore could be used as a marker of
                                                                                              diastolic dysfunction. Prospective studies on larger number of subjects are needed in order
A ROLE OF CARDIOTROPHIN-1 IN ALDOSTERONE-INDUCED CARDIAC
                                                                                              to confirm our preliminary findings.
HYPERTROPHY

Lopez N1, Martın B2, Martınez L1, Lahera V2, Dıez J1, Cachofeiro V2, Fortuno MA1
  ´             ´         ´                   ´                           ˜
1                                                                                                                                                                                   PA.05
  Centre for Applied Medical Research, Pamplona, Spain, 2Universidad Complutense,
Madrid, Spain                                                                                 OXIDATIVE STRESS IN THE HEART OF INTRAUTERINE GROWTH
                                                                                              RETARDED RATS
Aldosterone (ALD) induces left ventricular hypertrophy (LVH) and myocardial inflammation.
Cardiotrophin-1 (CT-1) is an inflammatory cytokine that increases cardiomyocyte size. The     Koleganova N1, Piecha G1, Ariel I4, Gross ML2, Bursztyn M5
                                                                                              1
present study was designed to investigate the association between ALD and CT-1 in vitro         University of Heidelberg, Department of Internal Medicine, Heidelberg, Germany,
                                                                                              2
and in Wistar rats receiving ALD (21 days, s.c.) at three doses (10, 100 and 1000               University of Heidelberg, Institute of Pathology, Heidelberg, Germany, 3Department of
  g/Kg/day). LVH was evaluated by planimetry using an image analysis system to measure        Nephrology, Endicrinology, and Metabolic Diseases, Medical University of Silesia,
left ventricular wall area (LVWA), wall thickness (LVWT), and cardiomyocyte length and        Katowice, Poland, 4Department of Pathology, Hadassah-Hebrew University Medical
width. Cardiac expression of ANP, c-fos, c-myc and CT-1 was quantified by real time           Center, Mount-Scopus, Jerusalem, Israel, 5Department of Medicine, Hadassah-Hebrew
RT-PCR and Western blotting. In HL-1 cardiomyocytes, ALD treatment (10-9-10-6 M)              University Medical Center, Mount-Scopus, Jerusalem, Israel
dose-dependently induced CT-1 expression via the mineralocorticoid receptor and
p38MAPK. Moreover, ALD-induced increase in hypertrophic genes ANP and c-fos                   Intrauterine growth retardation (IUGR) leads to increased neonatal mortality and morbidity
expression (p 0.01) was inhibited (p 0.01) by antibodies specific against CT-1 receptor.      and, in the long term, to hypertension, ischemic heart disease, cerebrovascular disease
In vivo, only the groups treated with ALD-100 and ALD-1000 exhibited higher blood             and diabetes mellitus, which are leading causes of mortality and morbidity in adults.
pressure than the control group (p 0.01). The three ALD-treated groups showed                 Female Wistar rats were treated with subcutaneous insulin pellet (to induce IUGR). The
increased cardiac index (p 0.01) and CT-1 expression (p 0.01). ANP, c-fos, and c-myc          hearts were obtained from the offspring on their last gestational or at 16 weeks of age.
expression as well as cardiomyocyte length and width were also enhanced (p 0.01) in the       Blood pressure was measured using tail plethysmography. Oxidative stress markers were
three groups of ALD-treated rats. LVWA and LVWT were enlarged only in ALD-1000                assessed using immunohistochemistry. Fetuses of hyperinsulinemic dams were smaller
animals (p 0.01). In summary, ALD induces cardiac expression of the hypertrophic              than those of normal dams (5.1 0.4 g vs. 5.6 0.1 g, respectively; p 0.05). The
cytokine CT-1 and this protein seems to mediate the hypertrophic effects that the             heart/body weight ratio was lower in IUGR females and not different in IUGR males
mineralocorticoid directly exerts on cardiomyocytes, being independent from its hemody-       compared to controls. The IUGR males (but not females) had significantly higher blood
namic effect.                                                                                 pressure compared to controls (148 10 vs. 118 15 mmHg, p 0.001) at week 16. IUGR
                                                                                              newborns had lower expression of eNOS, iNOS, and superoxide dismutase compared to
                                                                                              controls. In 16-week-old IUGR offspring lower expression of nNOS and higher expression
                                                        PA.03                                 of TGF-beta was observed. Higher expression of phosphorylated p38 and phosphorylated
EFFECTS OF ISCHEMIA AND REPERFUSION ON CARDIAC FIBROBLASTS                                    ERK1/2 was observed only in the male IUGR offspring. IUGR born male rats had higher
                                                                                              blood pressure and increased oxidative stress in myocardium in adulthood. Pro-
Faramarzi S, von Bauer R, Ulbrich C, Paul M, Grimm D                                          hypertrophic, maladaptive signalling was observed in these animals.
      ´          ¨
Charite Universitatsmedizin, Berlin, Germany

Myocardial infarction (MI) is accompanied by a progressive accumulation of extracellular                                                             PA.06
matrix proteins (ECMP). They are produced predominantly by cardiac fibroblasts. We            CHRONIC AC-SDKP ADMINISTRATION REDUCES PERIVASCULAR FIBROSIS
studied the effect of ischemia and reperfusion on isolated cardiac fibroblasts. We cultured   AND TGF- IN THE HEART OF DIABETIC RATS
fibroblasts from left ventricles of ischemic (n 7) and non-ischemic (n 7) hemoperfused
porcine working hearts. The non-infarcted fibroblasts continued to produce the normal low     Castoldi G1, di Gioia C2, Perego L1, Mancini M2, Bombardi C1, Travaglini C2, Zerbini G3,
amounts of ECMP. Cardiac fibroblasts originating from the infarct region exhibited            Stella A1
                                                                                              1
enhanced synthesis of various ECMP. Collagen type I, III, Osteopontin (OPN), laminin and        Clinica Nefrologica,Az. Osp. San Gerardo. Dip Medicina Clinica e Prev. Univ.
fibronectin proteins were significantly increased in fibroblasts of ischemic regions          Milano-Bicocca, Monza, Italy, 2Dip. Medicina Sperimentale, Univ. La Sapienza, Rome,
compared to fibroblasts derived from the remote region or control myocardium. OPN mRNA        Italy, 3Unita’ di Fisiopatologia Renale del Diabete. Ist. Scient. San Raffaele, Milan, Italy
and protein was significantly upregulated in fibroblasts derived from the infarct area. 0.1
  M and 1 M (Angiotensin) Ang II significantly increased collagen, fibronectin, laminin,      Introduction: N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) is a tetrapeptide normally
OPN and TGF- 1 synthesis in cardiac fibroblasts. Ang II induced the cleavage of 116-kDa       present in plasma and hydrolized by ACE, with potential antifibrotic effect. Aim: To
poly(ADP-ribose)polymerase (PARP) which resulted in an increase in the 85-kDa apoptosis-      evaluate the effect of chronic Ac-SDKP administration on perivascular fibrosis and TGF
808          Hypertension Vol 50, No 4 October 2007


expression in heart of diabetic rats. Methods: Ac-SDKP (1mg/kg/die s.c.,n 6) or saline                                                                                                   PA.09
(n 7) were administered for two months by osmotic minipumps to 13 diabetic                       SOY DIET ATTENUATES CARDIAC HYPERTROPHY
(streptozotocin injection) Sprague-Dawley rats. Five rats (control group) underwent buffer
injection. Systolic blood pressure (SBP) and blood glucose level were measured during the        Westphal C1, Schubert C1, Prelle K2, Fliegner D1, Penkalla A1, Regitz-Zagrosek V1
whole experimental period. At the end of the experimental period rats were sacrificed,           1
                                                                                                   Charite – Universitatsmedizin Berlin CCR/GiM, Berlin, Germany, 2Schering AG, Berlin,
                                                                                                         ´            ¨
blood sample was collected to measure Ac-SDKP (EIA), and the hearts were excised. For            Germany
histomorphological analysis, coronal sections (5 m thick) of the left ventricle were
obtained and stained with Sirius Red. Perivascular fibrosis (%) was determined by                Background: Women have a more favourable adaptation to pressure overload than men.
quantitative morphometry with an automated image analysis system (MetaMorph. 6.2,                Exogenous Estrogen (E2) has been shown to reduce myocardial hypertrophy in animal
UIC). TGF levels in the heart were measured by ELISA (Promega). Results: Results are             models of pressure overload (PO). The effect of phytoestrogens, that are usually present
shown in the table: Conclusion: Chronic Ac-SDKP administration reduces perivascular              in soy-based animal food, on cardiac hypertrophy has not yet been studied. Aim: We
fibrosis and TGF expression in the hearts of diabetic rats, suggesting that Ac-SDKP might        investigated the influence of different estrogen receptor modulators and soy– based diet on
play a role in the benefical effects of the ACE-inhibitors in diabetic cardiomyopathy.           pressure overload induced myocardial hypertrophy in ovariectomized mice. Methods:
                                                                                                 Female C57Bl6 mice were kept on soy–free or soy– based diet from the age of three
                               Control          Diabetes            Diabetes Ac-SDKP
                               (n 5)             (n 7)                     (n 6)                 weeks. One week after ovariectomy at the age of seven weeks, release pellets with
                                                                                                 Estradiol, Estrogen receptor(ER) alpha–agonist or Placebo were implanted. At the age of
SBP (mmHg)                   114.1    5.4     125.6    8.3             123.3    10.4             nine weeks, we performed transverse aortic constriction (TAC) or Sham surgery.
Blood glucose (mg/dl)         81.2    3.4     421.1    26.8*           392.6    28.8*            Echocardiographic measurements were done nine weeks after TAC. Organs were
Ac-SDKP (nmol/l)               2.51    0.1      1.81    0.1              3.94    0.4*°
                                                                                                 harvested and frozen until use. Results: There was a significant increase in LVW/TL in all
Perivasc.fibrosis %           29.6    1.1      42.7    4.5*             28.5    3.1°
TGB pg/mg.prot.)              80.3    6.8     113.3    9.6*             80.5    8.7°             TAC operated animals compared to Sham nine weeks after TAC (Table). Ejection fraction
                                                                                                 (EF) decreased in all TAC groups, except the Estradiol treated and the soy– based diet
   *p 0.05 vs control; °p 0.05 vs diabetes; (ANOVA).                                             group. Conclusion: Pressure overload by TAC induced significant cardiac hypertrophy.
                                                                                                 Treatment with Estradiol or soy– based diet prevented cardiac hypertrophy and progres-
                                                                                                 sion to heart failure. Table: Echocardiographic calculations
                                                                                                 Group                                       diet             LVW/TL (mg/mm)            EF (%)

                                                                                                 Sham-TAC Placebo                         Soy–free                    6.1                 45.8
                                                          PA.07                                  TAC Placebo                              Soy–free                   12.7                 34.1
CHRONIC TREATMENT OF RATS BY ANGIOTENSIN II (ANG II) INCREASES                                   TAC Estradiol                            Soy–free                   10.5                 39.3
PDE ACTIVITIES IN RAT LEFT CARDIAC VENTRICLES                                                    TAC ER                                   Soy–free                   14.2                 42.2
                                                                                                 TAC Placebo                             Soy–based                    9.0                 41.1
Mokni W, Allison W, Etienne-Selloum N, Schini-Kerth V, Lugnier C
Institut Gilbert Laustriat CNRS UMR 7175, Illkirch, France
                                                                                                                                                          PA.10
The renin-angiotensin system actively participates to the development of heart failure. The      IDENTIFICATION OF LEFT VENTRICULAR PROTEINS MODULATED IN AN
objective was to evaluate the implication of PDEs, which regulate cardiac function, in heart     EXPERIMENTAL MODEL OF HEART FAILURE
failure induced by chronic treatment of rats with Ang II. Four male rats of 12-week age
were treated during 14 days by Ang II (0.4 mg/kg/d) using osmotic mini-pumps. Control            Cieniewski-Bernard C1, Mulder P2, Thuillez C2, Amouyel P1, Richard V2, Pinet F1
group (6 rats) received saline solution. Left ventricles were removed, frozen, powdered          1
                                                                                                   Inserm U744, Lille, France, 2Inserm U644, Rouen, France
individually in liquid nitrogen. The cAMP- and cGMP-PDE activities were measured, and
the contribution of PDE1 to PDE5 was assessed by using their respective PDE inhibitors.          One of the major determinants of the severity of heart failure is the extent of left ventricular
Ang II induced the development of cardiac hypertrophy (P 0.017) and arterial hyperten-           (LV) remodelling after myocardial infarction. Indeed, in patients, the severity of LV
sion ( 54 mm Hg, P 0.001), without tachycardia. The cAMP-PDE activity was 2.6 times              remodelling cannot be fully predicted. Recent developments in the field of proteomic
higher than the cGMP-PDE activity in control rats. These activities were mainly and              technology allow to analyse the entire cellular proteome, leading to the identification of
respectively due to PDE4 (specific for cAMP) and PDE2 (which hydrolyzes cAMP and                 proteins involved in a pathological condition and so to determine markers of the pathology.
cGMP). In Ang II-treated rats, PDE4 activity was increased ( 20%, P 0.0004). A specific          The purpose of our study is to identify, using a proteomic approach, proteins that are
inhibitor of PKA, H89 (1 M), did not affect the cAMP-PDE activity in control rats, but it        differentially expressed in viable left ventricle in an animal model in which myocardial
decreased the activity in Ang II-treated rats (-22%, P 0.005). The cGMP-PDE1 ( 167%,             infarction was induced by left coronary artery ligation. Left ventricle was removed from
P 0.012), PDE2 ( 85%, P 0.0001) and PDE5 ( 100%, P 0.002) activities were                        heart failure and from sham animals two months after coronary ligation. Proteins extracted
increased significantly. These results show that cardiac hypertrophy induced by Ang II is        from left ventricle were separated using two-dimensional gel electrophoresis ; silver
mainly associated with increased PDE2 and PDE4 activities. Increased PDE4 activity could         stained gels were than analysed using the bioinformatic software Platinum®6.0. Around
result from a PKA-dependent phosphorylation. These findings suggest that cAMP and                1200 polypeptidic spots were well resolved on 2D-gels, and more than twenty different
cGMP are implicated in the cardiac effects induced by Ang II.                                    proteins are differentially expressed between sham and heart failure animals. We have
                                                                                                 identified these proteins using mass spectrometry analysis. These proteins belong to
                                                                                                 various classes of proteins, including heat shock proteins, proteins implicated in protection
                                                                                                 against oxidative stress and metabolic enzymes. This work should provide important
                                                                                                 insights into the cellular mechanisms involved in left ventricular remodelling which occur
                                                                                         PA.08   after myocardial infarction, and could allow to identify new biomarkers.
ANGIOTENSIN RECEPTORS IN INFLAMMATION AFTER MYOCARDIAL
INFARCTION
                                                                                                                                                        PA.11
Curato C, Timm M, Brinckmann MP, Grzesiak A, Altarche-Xifro W, Kaschina E, Unger T,
                                                          ´                                      DIRECT CARDIAC EFFECTS OF DIFFERENT IKR BLOCKERS ON
Li J                                                                                             LANGENDORFF-PERFUSED RABBIT HEARTS AND ISOLATED PAPILLARY
Center for Cardiovascular Research, Berlin, Germany                                              MUSCLES

Following acute myocardial infarction (MI), the heart suffers, beside ischemia-induced               ´
                                                                                                 Mako E., Orosz Sz., Farkas S.
direct myocardial damage, from a subsequent indirect cardiac injury through an                   Gedeon Richter Lpc., Budapest, Hungary
inflammatory reaction. Several recent studies have directly or indirectly shown that
angiotensin receptors, referred to as AT1 and AT2 receptor subtypes, are constitutively          We studied the cardiac effects of dofetilide (selective IKr blocker, antiarrhythmic), quinidine
expressed in the heart and significantly upregulated during inflammatory reaction following      (non-selective IKr blocker, Na-channel blocker, antiarrhythmic) and sertindole (high affinity
cardiovascular injury. Our preliminary data demonstrated that cardiac AT2 receptors              antagonist of the human cardiac potassium channel, antipsychotic). On the Langendorff-
mediate IL-10 production in CD8 T-cells, a mechanism that may contribute to                      heart the following parameters were measured: QT-intervals of the surface electrocardio-
anti-inflammatory effects of AT2 receptors in cardiac ischemia and reveal a novel function       gram, left ventricular pressure and the coronary flow. The heart rate and the corrected
of AT2 receptors in modulating adaptive immune responses to injury. To further                   QT-interval (QTc) were calculated. As expected each drug elicited QTc-prolongation, which
characterize the role of angiotensin AT2 receptors in the pathogenesis of MI-induced             was 12.5, 28.7 and 57.6% for 10, 30 and 100nM dofetilide, 23.9, 31.2 and 52.4% for 1,
cardiac injury, we established a method for direct isolation of ex vivo T-cell populations       3 and 10 M quinidine and 10.6, 28.7 and 37.1% for 0.3, 1 and 3 M sertindole. In our
from rat spleens and infarcted hearts, in particular CD8 /AT2 and CD8 /AT2- T-cell               experimental conditions 1 M dofetilide and10 M sertindole caused TdP in a few cases.
populations. CD8 cells freshly isolated from rat spleens and infarcted hearts were able          On the isolated, electrically stimulated papillary muscle the following parameters were
to suppress the proliferation of embryonic rat cardiomyocytes (H9C2). The expression of          measured: extracellularly recorded action potential amplitude (APA), conduction velocity
the AT2 receptor may reduce CD8 T cell induced cardiac injury by the production of               (APC), refractory period (RP) and contraction amplitude (CA). Dofetilide did not exert any
anti-inflammatory cytokines in the context of the MI outcome.                                    effect on APA, APC and CA, but significantly increased the RP by 62.9, 86.8 and 96.6%
                                                                                                                       2007 ECCR Poster Presentations                                   809

at 1, 3 and 10 M respectively. Quinidine had a slight increasing effect of APC at the            luciferase expression vector containing the Il-6 promoter including the NF- B binding site.
highest administered concentration (10 M) and moderately increased the RP by 6.5, 15.9           In order to be able to detect a putative inhibitory effect of AT2R stimulation on enhanced
and 24.6% at the same concentration range. Sertindole slightly increased APC, decreased          IL-6 expression or NF- B activity, respectively, cells were pre-treated with TNF (10
CA at 10 M and markedly increased the RP by 7.2, 25.6 and 34.4% at 1, 3 and 10 M.                ng/ml). Incubation with either Ang II (10 –7 M) under concomitant AT1R blockade or with
Our results confirm a good correlation as far as the QT-prolonging effect of the tested          C21 (10 – 6M) led to a significant and dose-dependent reduction of IL-6 levels. This effect
drugs on isolated heart and RP increasing effect on papillary muscle are concerned.              could be inhibited by the specific AT2R antagonist PD123319. C21 related IL-6 inhibition
                                                                                                 did not occur in fibroblasts derived from AT2R-KO mice. Furthermore, C21 had an
                                                                                                 inhibitory effect on TNF -induced NF- B activity; an effect that could also be blocked by
                                                                                      PB.01      PD123319. We conclude that direct AT2-R stimulation causes an inhibition of NF- B
ANGIOTENSIN AT2 RECEPTOR MEDIATES CARDIAC C-KIT CELL                                             activity leading to reduced IL-6 levels suggesting that AT2R stimulation acts anti-
PROLIFERATION AND SUPPORTS SURVIVAL OF CARDIOMYOCYTE                                             inflammatory.

             ´
Altarche-Xifro W, Birka C, Kaschina E, Curato C, Brinckmann MP, Timm M, Grzesiak A,
Unger Th, Li J                                                                                                                                                                        PB.04
Center for Cardiovascular Research (CCR), Charite – University Medicine Berlin, Berlin,
                                                 ´                                               FERMENTED MILK DOES NOT INHIBIT ANGIOTENSIN CONVERTING
Germany                                                                                          ENZYME IN HUMANS

The expression pattern of angiotensin AT2 receptors with predominance during fetal life          Wuerzner G, Peyrard S, Blanchard A, Lalanne F, Marelle L, Azizi M
and upregulation under pathological conditions during tissue injury/repair suggests that         CIC-HEGP, Paris, France
AT2 receptors may exert an important action in injury/repair adaptive mechanisms. Less
is known about AT2 receptors in acute ischemia-induced heart injury. We aimed here to            Ingestion of fermented milk (FM) has been shown to decrease BP in hypertensive patients.
elucidate the role of AT2 receptors after acute myocardial infarction. Double immunoflu-         The putative mechanism could be inhibition of angiotensin converting enzyme (ACE) by
orescence staining showed that increased AT2 receptors were mainly detected in clusters          dairy peptides, since these peptides are weak ACE inhibitors in vitro. Objective: To
of small c-kit cells accumulating in the peri-infarct zone. Further, we isolated the c-kit       investigate the effects of FM on ACE activity in humans using sensitive methods. Design
cell population from rat infarcted hearts by modified MACS technology and FACS sorting.          and methods: 12 healthy male volunteers were given 330 ml of FM once daily (9 am)
Ex vivo cardiac c-kit cells, which are characterized by both sca-1 and AT2 receptor              during one week (D1 to D7) and a single dose of captopril 50 mg on D8 (active control).
expression, proliferate slowly and maintain a stable phenotype under in vitro conditions for     ACE inhibition was assessed in vivo by urine (U) and plasma (P) AcSDKP, by plasma active
more than 18 months after their isolation. In vitro, exogenous angiotensin II (Ang II)           renin and in vitro by plasma ACE activity (Cushman assay). Subjects completed a 12H urine
stimulated proliferation of cardiac c-kit cells but the AT2 receptor antagonist, PD123319,       collection (U0) before FM (D1, D7) and captopril (D8) intakes followed by two 2H collections
abolished the effect. When cardiomyocytes isolated from adult rat hearts were co-cultured        (U1, U2) and a 4H collection (U3). Blood was sampled before FM (D1, D7) and captopril (D8)
with cardiac c-kit cells, treatment with Ang II or the AT2 receptor agonist, compound 21,        intakes and 0.5, 1 and 6 hours post-dose. Results: Compared to captopril (0.42 0.12 at
supported survival of cardiomyocytes and reduced apoptosis of cardiomyocytes. Together,          U0 vs 14 5 at U1, p 0.01), FM increased UAcSDKP (nM/h) (0.49 0.12 at U0 vs
these data demonstrate that AT2 receptors may mediate cardiac c-kit cell proliferation           0.73 0.34 at U1, p 0.05) only slighty and transiently. No effect on plasma AcASDKP,
to support survival of cardiomyocytes in response to ischemic injury.                            plasma ACE activity and plasma active renin was noted after repeated doses of FM,
                                                                                                 whereas captopril increased plasma AcASDKP, active renin and decreased plasma ACE
                                                                                                 activity from H0.5 to H6. Conclusion: The minimal ACE inhibition achieved after repeated
                                                          PB.02                                  doses of FM cannot explain their hypotensive effect observed in hypertensive patients,
IDENTIFICATION OF THE CALCIUM BINDING SITE OF AMINOPEPTIDASE A                                   indicating that other mechanisms should be investigated.

Claperon C1, Rozenfeld R1, Okada M1, Iturrioz X1, Maigret B2, Inguimbert N3, Roques
BP3, Llorens-Cortes C1                                                                                                                                   PB.05
1
  Inserm U691 College de France, Paris, France, 2CNRS UMR 7503, Nancy, France,                   THE EFFECT OF THE RENIN-ANGIOTENSIN SYSTEM AND BRADYKININ ON
3
  Inserm U640, Paris, France                                                                     AUTOREGULATION OF CEREBRAL BLOOD FLOW
Aminopeptidase A (EC 3.4.11.7, APA) is implicated in the conversion of angiotensin (Ang)         Sigurdsson ST2, Strandgaard S1
                                                                                                 1
II to Ang III, which is one of the main effector peptides of the brain renin-angiotensin           Department of Nephrology, Copenhagen University Hospital, Herlev, Denmark,
                                                                                                 2
system. Ang III exerts a tonic stimulatory effect on the central control of blood pressure.        Neurobiology Research Unit, Rigshospitalet, Copenhagen University Hospital,
APA activity is enhanced by the presence of calcium which confers to APA a substrate             Copenhagen, Denmark
specificity for N-terminal acidic amino-acid residues. In this study, we identified in the APA
3D model the calcium binding site in the S1 subsite. This calcium atom is located in an          Background: The renin-angiotensin system maintains a tone in the cerebral resistance
hydrophilic pocket where it interacts directly with aspartate (Asp) 213 and Asp 218 and the      vessels. Blockade of the system with an ACE-inhibitor or an angiotensin II receptor
side chain of the N-terminal acidic residue of the substrate or inhibitor via a water            antagonist shifts the lower and upper blood pressure limits of autoregulation of cerebral
molecule. We validated this model by site-directed mutagenesis studies by substituting           blood flow (CBF) towards lower blood pressure. This has been demonstrated by our group
Asp 213 and Asp 218 by an alanine, a glutamate, an asparagine and a glutamine. We                with the ACE-inhibitors captopril, ceranopril and fosinopril and the angiotensin II subtype
observed that the negative charge in position 213 and 218 is essential for maintaining the       1 receptor antagonist candesartan with the use of the intracarotid xenon technique. By
ability of recombinant mutated and wild-type APAs to be activated by calcium.                    contrast, the angiotensin II subtype 2 antagonist PD 123319 did not influence the limits of
Determination of kinetic parameters for recombinant APAs and inhibitory potencies for            autoregulation. Methods: The study was carried out in Sprague-Dawley rats in general
several inhibitors showed that Asp 213 and Asp 218 are involved not only in calcium              anesthesia with isofluran. Temperature and pCO2 were kept stable. Catheters were placed
binding but also in APA enzymatic activity playing a major role in substrate or inhibitor        in both femoral arteries for recording blood pressure, for blood sampling and controlled
specificity. This led also to the design of new APA inhibitors, which allow to confirm the       bleeding. A third catheter was placed in a femoral vein for drug administration. CBF was
localization of Asp 213 and Asp 218 in the S1 subsite.                                           measured on the surface of the brain with laser Doppler technique. The lower limit of the
                                                                                                 autoregulation was then calculated via a computer program. Results and conclusion:
                                                                                                 Acute treatment with candesartan (30 mmHg) lowered the lower limit of the CBF
                                                      PB.03                                      autoregulation compared to controls (40 mmHg), also demonstrating the feasibility of the
DIRECT ANGIOTENSIN AT2R STIMULATION BY THE SPECIFIC                                              laser-Doppler method for this type of study.
NON-PEPTIDE AGONIST COMPOUND 21 ACTS ANTI-INFLAMMATORY IN
VITRO BY REDUCING NF- B ACTIVITY
                                                                                                                                                                                      PB.06
Rompe F1, Hallberg A2, Artuc M3, Bader M4, Unger Th1, Steckelings UM1                            TYPE II DIABETES REDUCES ANGIOTENSIN II TYPE 2
1
  Center for Cardiovascular Research, Charite-Universitatsmedizin Berlin, Berlin,
                                            ´          ¨                                         RECEPTOR-DEPENDENT DILATION THROUGH COX-2 GENERATED
Germany, 2Dept. of Medicinal Chemistry, Organic Pharmaceutical Chemistry, Upsalla                THROMBOXANE A2
                              3
                                                           ´          ¨
University, Upsalla, Sweden, Dept. of Dermatology, Charite-Universitatsmedizin Berlin,
Berlin, Germany, 4Max Delbruck Center for Molecular Medicine, Berlin, Germany
                             ¨                                                                   Retaileau K, Guillot A-L, Vessieres E, Chanoine S, Jardel Al, Belin de Chantemele E,
                                                                                                 Henrion D, Loufrani L
There is substantial evidence that the AT1-receptor (AT1R) mediates pro-inflammatory             INSERM771 CNRS6214, Angers, France
actions of angiotensin II (Ang II), while the role of the AT2-receptor (AT2R) in inflammation
is uncertain. In order to get more insight into AT2R actions in inflammation, we determined      The role of angiotensin II type 2 receptors (AT2R) is not fully understood in vascular
the effect of direct AT2R stimulation by the novel non-peptide AT2R agonist Compound 21          diseases. Abnormalities in the renin-angiotensin system have a key role in cardiovascular
(C21) on interleukin-6 (IL-6) expression in human and murine primary dermal fibroblasts          disorders associated with the evolution of type II diabetes. Thus, we hypothesized that
in vitro. Furthermore, NF- B activity was determined by immunohistochemical detection            AT2R function might be altered in diabetic resistance arteries. Mesenteric resistance
of NF- B-p50-subunit nuclear translocation and by luciferase reporter assay using a              arteries were isolated from type II diabetic Zucker (ZDF) and control (LZ) rats receiving the
810          Hypertension Vol 50, No 4 October 2007


antioxidant tempol or water for 3 weeks. Dilation to angiotensin II in the presence of                                                                PB.09
candesartan was studied in arteries. AT2R induced endothelium and NO-dependent                LOG-LINEAR DOMINANT DIETARY CONTROL OF RENIN SECRETION AND
dilation in LZ (inhibited by L-NAME). It was not affected by cyclooxygenase or reactive       RESPONSES TO PULSE CHANGE IN TOTAL BODY SODIUM
oxygen species inhibition. In ZDF rats AT2R-induced dilation was lower than in LZ rats and
independent of NO. It was restored to control level after ROS and cycloxygenase inhibition.   Kjolby M, Bie P
Cycloxygenase-2 and Thromboxane A2 receptor inhibition restored AT2R-dilation in ZDF          Dept. of Physiology and Pharmacology, Institute of Medical Biology, University of
rats to control (LZ) level. Cycloxygenase-2 and ROS were overexpressed in ZDF rats            Southern Denmark, Odense, Denmark
arteries. In ZDF rats chronically treated with Tempol AT2R-induced dilation was equivalent
to that in LZ rats and it involved NO and cycloxygenase-2-derivatives in equivalent           Reactions to sodium loading depend on the rate and magnitude of the change in body
proportion. In tempol-treated rats SQ29548 had no effect. We conclude that in type II         sodium and the sodium status. We hypothesized that a step increase in body sodium elicits
diabetic rats AT2R induced dilation is reduced due to ROS and thromboxane A2-dependent        a natriuretic response depending on sodium status, and measured the effects of a bolus
vasoconstriction due to COX-2 activation. Chronic treatment of diabetic rats with tempol      of hypertonic NaCl during different low-salt diets representing a 25-fold change in sodium
restored AT2R-dependent dilation through the suppression of ROS and TxA2 production.          intake. To custom-made, low-sodium chow (0.003 % Na ), NaCl was added to provide
These findings might be important to consider in the choice of vasoactive drugs in diabetic   four levels of intake, 0.03 to 0.75 mmol kg-1 d-1 for 7 days. In these steady states,
patients.                                                                                     hypertonic NaCl (1.2 mmol kg-1) was given i.v. to conscious dogs over 2 min. Arterial blood
                                                                                              pressure (MABP), glomerular filtration rate (GFR), plasma volume (PV), plasma renin activity
                                                                                              (PRA) and hormone concentrations were determined conventionally. Acute NaCl adminis-
                                                                                              tration increased PV ( 7.4%) and sodium ( 2 %) and decreased plasma protein (-7.3 %).
                                                                                              Plasma angiotensin II and aldosterone decreased transiently. Sodium excretion remained
                                                        PB.07                                 unchanged, but potassium excretion increased substantially. MABP, GFR, urine flow,
METABOLIC EFFECTS AND END ORGAN PROTECTION BY THE                                             plasma potassium, PRA did not change measurably. The results indicate that renal sodium
                                                                                              excretion is controlled by neurohumoral mechanisms remarkably resistant to acute
COMBINATION OF TELMISARTAN AND NIFEDIPINE IN SHR/NDmcr-cp
                                                                                              changes in blood volume and colloid osmotic pressure, and driven by mechanisms, which
RATS, AN ANIMAL MODEL OF METABOLIC SYNDROME                                                   are not down-regulated within a few hours. Together with previous data, the results
                                                                                              demonstrate that plasma renin is log-linearly related to sodium intake over a 250-fold
                                                  ¨
Tinel H, Bischoff H, Stelte-Ludwig B, Sandner P, Hutter J                                     change, thereby defining a renin function line by slope and intercept.
Bayer Healthcare AG, Product Related Research, Wuppertal, Germany

The organ protective effects of the combination of the angiotensin II type 1 receptor
blocker telmisartan and the calcium channel blocker nifedipine were examined in                                                                       PB.10
genetically hypertensive and obese SHR/NDmcr-cp rats. The animals were treated with           TELMISARTAN REDUCES ABDOMINAL ANEURYSM FORMATION IN THE
telmisartan (1 mg/kg/day), nifedipine (4.8 mg/kg/day) or with a combination of both drugs     RAT BY DECREASING PROTEOLYSIS, APOPTOSIS AND INFLAMMATION
for 22 weeks. After 22 weeks of treatment SBP of SHR/NDmcr-cp rats was 205.6 6.6
mmHg and was reduced by all three treatment regimen significantly. Telmisartan and the        Schrader F, Grzesiak A, Kemnitz UR, Krikov M, Sommerfeld M, Unger T, Kaschina E
combination therapy reduced the left ventricular hypertrophy as well as the mRNA                                                                         ¨
                                                                                              Center for Cardiovascular Research (CCR), Charite Universitatsmedizin, Berlin, Germany
expression of ANP. The BNP expression was reduced significantly only by the combination
therapy. Renal function and expression of extracellular matrix proteins were improved by      We investigated the effects of treatment with the AT1 receptor antagonist, telmisartan, on
nifedipine and the combination therapy. Telmisartan had no effects on these parameters.       abdominal aortic aneurysm (AAA) formation in the rat. AAA was induced by perfusion of
The liver weight was increased in SHR/NDmcr-cp rats compared with control. Plasma             isolated aortic segment with elastase (Anidjar/Dobrin model). Treatment with telmisatran
concentration of hepatic enzymes was reduced by all antihypertensive treatment. The level     (0.5 mg/kg i.p.) or hydralazine (15 mg/kg) was started after the operation and continued
of GLDH was reduced by telmisartan, AST and GLDH by nifedipine, and GLDH, AST and ALT         for 14 days. Sham operated animals and placebo treated animals after aneurysm induction
by the combination therapy. The impaired glucose tolerance of SHR/NDmcr-cp rats was           (AI) served as controls. Aortic diameter was measured using ultrasound biomicroscopy
not changed. Nifedipine and the combination treatment increased the plasma level of LDL       before AI and on days 7 and 14 after AI. Aortic tissues were studied using immunohis-
and HDL, and significantly reduced the level of plasma triglycerides and free fatty acids.    tology, immunoblotting and real-time PCR. Blood pressure, hemodynamic parameters and
These results indicate that in SHR/NDmcr-cp rats telmisartan and nifedipine were effective    serum cytokines were also investigated. On day 14 after AI, aortic diameter was increased
in lowering blood pressure and showed positive effects concerning organ protection and        2-fold in the placebo group compared to sham operated animals (2.02 0.12mm vs
metabolic parameters. Thus, optimal profile might be expected from the combination            0.87 0.02mm, p 0.005, n 8). Telmisartan treatment significantly reduced aneurysmal
therapy of telmisartan and nifedipine.                                                        size (1.65 0.06mm vs 2.02 0.12mm in placebo, p 0.05, n 8). Treatment with
                                                                                              hydralazine had no effect on aneurysmal size. MMP-3 and cathepsin D, TGF-1 beta, as
                                                                                              well as caspase 3 and FasL were significantly down-regulated in aortic tissue under
                                                                                              telmisartan treatment compared to placebo. MCP-1 levels were significantly decreased.
                                                                                              Telmisartan and hydralazine slightly reduced the blood pressure to a similar extent. The
                                                                                              AT1 receptor antagonist telmisartan protects from AAA formation independently of blood
                                                         PB.08
                                                                                              pressure reduction by inhibiting proteolysis, apoptosis and inflammation in the aorta.
EFFECT OF CHRONIC ALISKIREN ON BODY WEIGHT, LIPID PARAMETERS
AND ADIPOSE TISSUE RENIN-ANGIOTENSIN SYSTEM IN A MOUSE MODEL
OF DIET-INDUCED OBESITY
                                                                                                                                                                                  PC.01
Stucchi P1, Cano V2, de las Heras AI1, Ruiz-Gayo M2, Fernandez-Alfonso MS1                    SOLUBLE ENDOGLIN CAUSES IMPAIRED ENDOTHELIUM DEPENDENT
1
  Instituto Pluridisciplinar, UCM, Madrid, Spain, 2Departamento de Farmacologıa,
                                                                             ´                RELAXATION IN RAT TAIL ARTERY: A POSSIBLE PATHOGENIC
Tecnologıa y Desarrollo Farmaceutico. Facultad de Farmacia. USP-CEU., Madrid, Spain
           ´                       ´                                                          MECHANISM IN PRE-ECLAMPSIA

Overfeeding of rodents leads to an increase in adipose tissue mass related to an elevation    Sujenthiran A, Wijetunge S, Hughes AD
of adipose angiotensinogen and angiotensin II (Ang II) formation. The aim of this study was   Imperial College London, London, United Kingdom
to determine the effect of a chronic treatment with the renin inhibitor, aliskiren, on
diet-induced obesity. We used mice (C57BL/6) treated with either saline or aliskiren (A; 50   Background: Endoglin (CD105) is highly expressed in endothelial cells and modulates the
mg/kg/day), and exposed either to a low fat (LF, n 10; LFA, n 10) or a high fat (HF,          actions of TGF- 1 by interacting with TGF- 1 receptors types I and II. The role of endoglin
n 10; HFA, n 10) diet during 6 weeks. Aliskiren induced a significant reduction               in vascular homeostasis is unclear, but increased circulating levels of soluble endoglin are
(*p 0.05) of body weight independently of the type of diet (LF 26 0.5g;                       found in pre-eclampsia and are implicated in its pathogenesis [1]. However, the acute
LFA 25 0.3g*; HF 28 0.4g; HFA 27 0.4g*), without modifying food intake or                     effects of soluble endoglin on vascular reactivity have not been studied in vitro. Methods:
caloric efficiency. Moreover, aliskiren significantly reduced the amount of lumbar            Isometric tension was measured in rat tail arteries mounted in a myograph containing a
(LF 82 8g; LFA 71 5g; HF 208 17g; HFA 130 12g*), epididimal                                   physiological saline solution (PSS). Arteries were bubbled with 95% O2, 5% CO2 and
(LF 317 19g; LFA 241 14g*; HF 529 44g; HFA 415 23g*), and mesenteric                          maintained at 37°C. Results: In endothelium-intact arteries, pre-incubation with endoglin
adipose tissue (LF 280 15g; LFA 237 20g*; HF 374 34g; HFA 322 18g*).                          (20ng/ml) for 1h, inhibited the acetylcholine (Ach) concentration-response curve (pEC50
These data correlated with a decrease in plasma leptin levels (LF 10 1ng/ml; LFA 6.5          control -7.5 0.2, endoglin -6.7 0.2, p 0.005, n 6) in arteries pre-contracted
   0.3ng/ml*; HF 14 2ng/ml; HFA 7.5 0.5ng/ml*). No changes were observed in                   with phenylephrine (1 M). Endoglin did not affect phenylephrine- induced contractions. In
either lipidic or glycemic parameters between groups. In addition, Ang I (LF 0.5 0.2pg/       contrast TGF- 1 (20ng/ml) did not have significant effects on phenylephrine -induced
mg; LFA 0.3 0.04pg/mg; HF 0.7 0.2pg/mg; HFA 0.3 0.1pg/mg) and Ang II levels                   contraction or Ach-induced relaxations. Conclusion: Soluble endoglin inhibits
(LF 1.8 0.6pg/mg; LFA 0.7 0.1pg/mg; HF 1.7 0.6pg/mg; HFA 1 0.3pg/mg),                         endothelium-dependent relaxation. This effect of endoglin in inducing impaired vasodila-
determined by EIA after HPLC were reduced by aliskiren, although not significantly. These     tion and endothelial dysfunction could have implications in vivo in the pathogenesis of
results suggest that aliskiren reduces body weight by acting on adipose tissue likely         pre-eclampsia, in particular the associated hypertensive state. [1]. Venkatesha et al., Nat
through a reduction of adipogenic effects of Ang II (supported by Novartis).                  Med. 2006;12:642–9.
                                                                                                                      2007 ECCR Poster Presentations                                   811

                                                       PC.02                                    dependent reduction in EPC number (3 mol/l Hcy 7.7 2.1, 10 mol/l 6.3 1.9, 30 mol/l
PARACRINE STIMULATION OF VASCULAR SMOOTH MUSCLE                                                 5.4 0.8 and 100 mol/l 1.9 0.9 cells per view, P value             0.01 for each). Cys also
PROLIFERATION BY DIADENOSINE POLYPHOSPHATES RELEASED FROM                                       caused a similar concentration-dependent reduction in EPC number (3 mol/l Cys
                                                                                                8.3 1.5, 10 mol/l 7.2 1.2, 30 mol/l 4.5 2.3 and 100 mol/l 1.2 0.6, P value 0.01
PROXIMAL TUBULE EPITHELIAL CELLS
                                                                                                for each). There was no significant difference in the concentration response curve for Cys
                                                                                                and Hcy (P value        0.05). The effect of Cys on EPC number has not previously been
Jankowski V, Zidek W, Jankowski J
                                                                                                described. In conclusion, Cys may be as damaging as Hcy with regard to EPCs, and
Charite Med Klinik IV, Berlin, Germany
                                                                                                extracellular thiols other than Hcy may also contribute to the risk of atherosclerosis.
Although it is well known that the purinergic receptor system plays an important role in the
regulation of tubular functions, it is less known whether other purinoceptor agonists than
ATP are released by renal tissue. Therefore, in the present study we examined whether
                                                                                                                                                         PC.05
renal tissue is the source of a specific group of endogenous purinergic agonists, the           ENDOTHELIUM-DEPENDENT ATTENUATION OF NORADRENERGIC
diadenosine polyphosphates. Because of close vicinity of tubules and peritubular vessels,       VASOCONSTRICTION IS PREVENTED BY NO SYNTHASE INHIBITION OR
making a paracrine function of mediators secreted by renal tubules not unlikely, and            CALCIUM-DEPENDENT POTASSIUM CHANNELS BLOCKADE
because of the high-energy consumption by renal tubular cells, where a feedback
regulation of vascular supply by renal tubules is necessary, the effect of the identified       Liskova S1, Kunes J2, Zicha J2
                                                                                                1
diadenosine polyphosphates on VSMC proliferation was analysed subsequently. Renal                 Institute of Pharmacology, School of Medicine, Comenius University, Bratislava,
tissue was homogenated and fractionated by different chromatographic methods. In                Slovakia, 2CRC, Institute of Physiology AS CR, Prague, Czech Republic
fractions purified to homogeneity, Ap5A as well as Ap6A were identified by MALDI-MS,
retention time comparison and enzymatic cleavage analysis. Next, we examined whether            The influence of endothelium on tonic noradrenergic contraction as well as the impact of
cultured renal proximal tubule cells secreted Ap5A and Ap6A. Ap5A and Ap6A amounts were         NO synthase inhibition and Ca2 -dependent K channel blockade were studied. Femoral
increased in the supernatants after stimulation suggesting that Ap5A and Ap6A may have          arteries isolated from Wistar-Kyoto rats were mounted to Mulvany-Halpern myograph. NO
a functional impact on structural elements of the kidneys. Both Ap5A and Ap6A stimulated        synthase inhibitor (L-NNA, 10-4M), Ca2 -dependent K channels (KCa) blocker (TEA,
growth of VSMC in nanomolar concentrations ranging from 10-9 to 10-5 mol/l. The                 10-3M) and voltage-dependent calcium channels (VDCC) blocker nifedipine (NIF, 10-7M)
purinergic receptor antagonist suramin did not inhibit growth stimulatory effects of Ap5A       were added either prior (L-NNA) or after (TEA, NIF) norepinephrine (NE)-induced
or Ap6A significantly, indicating that suramin-insensitive receptors may be involved in the     contraction. Presence of endothelium largely inhibited NE-induced tonic contraction. The
growth promoting effect. Besides their direct growth stimulatory action, both Ap5A and          ability of endothelium to attenuate NE-induced contraction was diminished either by NO
Ap6A markedly potentiated the growth stimulation by platelet-derived growth factor              synthase inhibition or by the blockade of KCa. KCa blockade by TEA augmented
(PDGF). In summary, renal tubular cells release the purinergic agonists Ap5A and Ap6A after     NE-induced tonic contraction of vessels with intact endothelium to the values seen either
stimulation, suggesting a functional role for these substances in the kidney. Moreover, the     after NO synthase inhibition or in the absence of endothelium. NIF inhibited tonic
diadenosine polyphosphates induced VSMC proliferation may be one element of tubular             contraction proportionally to its magnitude. Thus greater NIF-induced reduction of wall
vasoregulatory properties, providing a link between tubular function and energy supply.         tension was seen in vessels with removed endothelium or in vessels with intact
                                                                                                endothelium subjected either to NO synthase inhibition or KCa blockade. We can conclude
                                                                                                that intact endothelium attenuates tonic noradrenergic contraction by inhibiting nifedipine-
                                                         PC.03                                  sensitive calcium influx (through VDCC). Blockade of Ca2 -dependent K channels by TEA,
A NEW ZAPRINAST DERIVATIVE SELECTIVELY AND POTENTLY INHIBITS                                    which prevents membrane hyperpolarization, was able to diminish the endothelium-
VASCULAR PDE5 IN COMPARISON WITH PLATELET PDE5                                                  dependent inhibition of NE-induced tonic contraction. (grant 1M0510)

Kameni-Tcheudji JF1, Keravis T1, Le Bec A1, Lebeau L1, Strub JM2, van Dorsselaer A2,
Lugnier C1                                                                                                                                                                            PC.06
1
  Institut Gilbert Laustriat CNRS UMR 7175, Illkirch, France, 2Institut Pluridisciplinaire      5-HT AND HISTAMINE RECEPTOR SUBTYPES INVOLVED IN THE
Curien, Strasbourg, France                                                                      REGULATION OF HUMAN UMBILICAL ARTERY TONE

Cyclic nucleotide phosphodiesterases (PDEs) play a key role in signal transduction by                                         ´
                                                                                                Santos-Silva AJ1, Cairrao E1, Alvarez E1, Marques B1, Lavrador I1, Verde I1
                                                                                                                        ˜
                                                                                                1
hydrolyzing specifically cyclic nucleotides. We have shown that specific inhibition of                               ¸˜         ˆ           ´
                                                                                                  Centro de Investigacao em Ciencias da Saude (CICS). Universidade da Beira Interior,
vascular PDE5 by zaprinast induces relaxation of rat aorta along with an increase in cGMP       Covilha, Portugal, 2Centro Hospitalar da Cova da Beira E.P.E, Covilha, Portugal
                                                                                                       ˜                                                            ˜
level and also that PDE5 in platelets participates in controlling aggregation. The aim of the
present study was to compare peptidic sequences, biochemical properties and pharma-             The human umbilical artery (HUA) is involved in fetoplacental circulation. Endocrine and
cological sensitivities of vascular PDE5 and platelet PDE5. Cytosolic fractions of smooth       paracrine mechanisms that regulate the tonus of this artery are very important for optimum
muscle from bovine aorta and from human platelet were submitted to DEAE-sepharose               gas and nutrients exchange between the foetus and the placenta. To investigate the
chromatography and subsequently to affinity chromatography on ECH-Sepharose 4B resin            involvement of serotonin (5-HT) and histamine (His) receptors in the regulation of
coupled with NH2-zaprinast. Fractions containing PDE5 were submitted to Western blot            endothelium denuded HUA contractility, the effect of several agonists of these receptors
and to gel electrophoresis. PDE5 bands were excised from gel and submitted to                   were analysed. Concerning the 5-HT receptors, only the 5-HT1B/D and 5-HT2A agonists
gel-digestion in order to perform MALDI-TOF analysis; Km and Vmax determinations as well        induced significant contractile effects. 5-HT1B/D- agonist had dual effects: in some arteries
as Ki were determined on purified PDE5s. Western blot shows that platelet PDE5 is a             it induced a small contractile effect (150mg of tension approximately) and contraction
85-kDa protein whereas vascular smooth muscle PDE5 is a 90-kDa protein. MALDI-TOF               around 1g was obtained in other arteries. The 5-HT-induced contractions were fully
analysis reveals that vascular smooth muscle PDE5 and human platelet PDE5 are different         inhibited by ketanserin (5-HT2A-selective antagonist), and partially inhibited (29%) by
proteins. N-alkylated-zaprinast inhibits vascular PDE5 (Ki 0.032 nM) platelet PDE5              nifedipine, a L-type calcium channel (LTCC) blocker. Concerning the His receptors, only
(Ki 1.8 nM). Our results show that vascular PDE5 and platelet PDE5 are different proteins       H1-receptor activation by betahistine induced significant contractile effect. His induced
which can be discriminate by N-alkylated-zaprinast and suggest that they might represent        contraction was blocked by the H1-selective antagonist pyrilamine (25%) and partially
different PDE5 variants which can be selectively inhibited. These data open new                 inhibited by nifedipine (54%). Also, Betahistine induced contraction was partially blocked
therapeutic opportunities for using PDE5 inhibitor treatment in patients treated with           by dimaprit (H2-selective agonist). These findings show that activated 5-HT2A and 5-HT1B/1D
antiplatelets.                                                                                  receptors are responsible for the vasoconstriction induced by 5-HT. Activation of H1
                                                                                                receptors induces contraction of HUA, meanwhile H2 receptor activation induces vasore-
                                                                                                laxation. Finally, the activation of LTCC is partially responsible of the contractions induced
                                                       PC.04                                    by the activation of 5-HT2A, 5-HT1B/D, and H1 receptors. We thank the Gynaecology-
COMPARISON OF THE EFFECTS OF CYSTEINE AND HOMOCYSTEINE ON                                       Obstetrics Department staff of “Centro Hospitalar da Cova da Beira” and “Fundacao para¸˜
ENDOTHELIAL PROGENITOR CELLS IN VITRO                                                                ˆ
                                                                                                a Ciencia e a Tecnologia” which supported the fellowship SFRH/BDE/15532/2004.

Oakland KA, Schachter M, Hughes AD
Imperial College, London, United Kingdom                                                                                                               PC.07
                                                                                                ROLE OF THE SOLUBLE GUANYLYL CYCLASE 1 1 (sGC 1 1) ISOFORM
Homocysteine (Hcy) has been implicated in atherosclerosis partly because of its effects on      IN MICE CORPUS CAVERNOSUM SMOOTH MUSCLE RELAXATION
the endothelial progenitor cells (EPCs) which repair the endothelium. In vitro, Hcy has been
shown to reduce EPC number. The aim of this study was to investigate whether a                  Nimmegeers S1, Sips P2, Buys E2, Brouckaert P2, Van de Voorde J1
                                                                                                1
structurally related thiol, cysteine (Cys) has a similar effect. EPCs were isolated from          Department of Physiology and Physiopathology, Ghent University, Ghent, Belgium,
                                                                                                2
peripheral blood of healthy volunteers (n 11) by density centrifugation. After culture on         Department of Molecular Biomedical Research, Flanders Interuniversity Institute for
fibronectin coated cover slips, EPCs were treated with 3, 10, 30 or 100 mol/l Cys or Hcy.       Biotechnology and Ghent University, Ghent, Belgium
EPCs were characterised by staining for lectin and AcLDL-Dil and cell counts were
performed using 5 randomly selected fields per slide. Data are presented as means SD.           As the major effector molecule for NO, soluble guanylyl cyclase (sGC) plays a key role
In comparison to vehicle control (12.8 1.8 cells) Hcy caused a significant concentration-       within the NO/cGMP signalling cascade which participates in penile erection. The enzyme
812          Hypertension Vol 50, No 4 October 2007


exist as an      -heterodimer, but only two isoforms have been reported to be active                                                                                                 PC.10
(sGC 1 1 and sGC 2 1). The functional importance of the 1-subunit in corpus caverno-           THE ROLE OF ENDOTHELIUM IN APELIN INDUCED VASCULAR
sum (CC) smooth muscle relaxation was assessed by mounting CC from male sGC 1-/-               RELAXATION
mice and wild type littermates in organ baths for isometric tension recording. The
endothelium-dependent relaxation to acetylcholine (ACh) or bradykinin (BK) and the             Gurzu B, Dumitriu IL, Slatineanu SM, Petrescu G
neurogenic response to electrical field stimulation (EFS) were nearly abolished in the         Departament of Physiology, Faculty of Medicine, University of Medicine and Pharmacy,
sGC 1-/- CC. The relaxing influence of exogenous NO (from sodium nitroprusside (SNP) and       Gr. T. Popa, Iasi, Romania
NO-gas) was also significantly decreased in the sGC 1-/- mice. The remaining relaxation
seen in the sGC 1-/- mice with exogenous NO, was strongly but not completely inhibited         Apelin (AP) is the endogenous ligand of the recently de-orphanized APJ receptor. Despite
by the sGC-inhibitor ODQ. In the preparations of the sGC 1-/- mice, the response to BAY        the highly expression of both AP and APJ in the vascular wall, the vasomotor effects of AP
41–2272 (NO-independent sGC-stimulator) and to T-1032 (phosphodiesterase type 5                are still under investigation. We have recently shown that AP plays a counter-regulatory
inhibitor) were also significantly reduced. The specificity of the impairment of the           role against the angiotensin II -induced vasoconstriction. The aim of this study was to
sGC-related responses was demonstrated by the similar forskolin (adenylyl cyclase              investigate the action of AP (cumulative dose, 1nM - 10 M) on aorta (Ao), pulmonary
activator) and 8 pCPT-cGMP (cGMP-analogue)-induced responses. In conclusion, our               artery (PA), renal vein (RV) and portal vein (PV) precontracted with phenylephrine (Phe,
findings indicate the involvement of an sGC isoform with the 1-subunit in NO-induced CC        10 M) and to what extent, if any, NO mediates the AP effects. Experiments were
smooth muscle relaxation. However, the remaining relaxing influence of exogenous NO in         performed on rat vessels rings with (EI) or without endothelium (ED), in absence or
the sGC 1-/- mice, suggests the contribution of (an) additional pathway(s).                    presence of 10 M N(G)-nitro L-arginine methyl ester (L-NAME, NO synthase inhibitor). The
                                                                                               NO production was continuously monitored by a NO meter. The AP-induced relaxation
                                                                                               were the highest on PV (Emax        56.41 3.18%) and the lowest on the Ao intact rings
                                                       PC.08                                   (Emax       16.93 6.24%). The absence of intact endothelium or L-NAME pretreatment
HYDRALAZINE RESTORES FLOW (SHEAR STRESS)-MEDIATED                                              prevented AP effects on Ao and RV rings. On PA ED rings or on PA EI rings but after
                                                                                               L-NAME pretreatment AP effects were still significantly but 2.5 and 1.7 times lower.
STRUCTURAL AND FUNCTIONAL REMODELING IN RESISTANCE ARTERIES                                    Blocking NO synthesis decreased AP-induced relaxation of Phe precontracted PV rings,
FROM AGED RATS                                                                                 L-NAME being significantly powerful on endothelium denuded (up to 90%) than endothe-
                                                                                               lium intact rings (up to 60%). These results, together with NO measuring, suggested that,
Dumont O, Guihot AL, Pinaud F, Baufreton C, Loufrani L, Henrion D                              depending on vessel type, AP could relax pre-contracted vessels by NO dependent and
Inserm771-cnrs6214, Angers, France                                                             partially endothelium – independent mechanisms.
Aging, a major risk factor for cardiovascular diseases is associated with dramatic changes
in large arteries structure and function. The link between aging and vascular diseases                                                                  PC.11
remains unclear. Nevertheless, a decreased vasodilator capacity of the endothelium is          NIFEDIPINE-INDUCED SHIFT OF NOREPINEPHRINE DOSE-RESPONSE
usually described in aging. Thus, we hypothesized that arteriolar remodeling in response
to a chronic increase in blood flow might be altered, thus preventing tissues to adapt to
                                                                                               CURVE IS SIMILAR IN RESISTANCE VASCULATURE OF CONSCIOUS SHR
chronic changes in either exercise or in ischemic diseases. We also tested a vasodilator       AND WKY RATS
treatment with hydralazine, hypothesizing that it might restore remodeling. Methods and
results: Mesenteric resistance arteries from 3-month- and 24-month-old rats were               Pinterova M, Kunes J, Dobesova Z, Zicha J
submitted to high blood flow (HF) and normal blood flow (NF) by ligating sequentially          CRC, Institute of Physiology AS CR, Prague, Czech Republic
second order arteries in vivo. After 2 weeks, diameter increased in HF arteries (by 21%,
at 100 mmHg) in association with a rise in eNOS expression and superoxide production in        Higher sympathetic tone and augmented vascular responsiveness to noradrenergic
young rats. In old rats, diameter and eNOS failed to increase in HF arteries. Hydralazine      stimulation were reported in spontaneously hypertensive rats (SHR). Our studies with acute
restored HF-remodeling in old rats without increasing eNOS expression. Furthermore             i.v. nifedipine administration suggested that an important part of hypertensive action of
hydralazine also restored HF-remodeling in eNOS knockout mice suggesting that its effect       sympathetic nervous system was exerted by calcium influx through voltage-dependent
is eNOS-independent. Hydralazine increased superoxide production and decreased SOD             calcium channels. The present work was focused on 1) the comparison of dose-dependent
expression in old rats HF arteries. The SOD-mimetic TEMPOL prevented the effect of             responses to norepinephrine (NE) in SHR and Wistar Kyoto rats (WKY), and 2) the influence
hydralazine. Conclusion: Flow-remodelling in resistance arteries failed to occur but could     of nifedipine (NIF) on these NE-induced dose-response curves in both rat strains. We used
be restored by hydralazine via a ROS-dependent mechanism. These finding may have               conscious WKY and SHR male rats (aged 16 weeks) subjected to prior inhibition of
serious pathophysiological consequences in situations requiring flow-dependent remod-          angiotensin converting enzyme (captopril, 10 mg/kg) and ganglionic blockade (pento-
eling such as ischemic and metabolic diseases, more frequent in elderly. Nevertheless,         linium, 5 mg/kg). The increasing noncumulative NE doses were administered i.v. before
vasodilator treatments remain efficient.                                                       and after nifedipine injection (0.4 mg/kg). The maximal blood pressure rises after each of
                                                                                               NE doses were used to construct dose-response curves from which the ED50, maximal
                                                                                               response and slope were calculated. NE elicited similar pressor responses in both strains,
                                                                                               although there was a nonsignificant tendency to greater slope of the curve in SHR
                                                        PC.09                                  compared to WKY. Nifedipine shifted these dose-response curves to the right. This was
IN LONG-TERM HYPERGLYCAEMIA, OXIDATIVE STRESS RATHER THAN                                      associated with the steepening of the curve in SHR but not in WKY. Thus the augmented
HYPERGLYCAEMIA PER SE IMPAIRS ENDOTHELIAL FUNCTION                                             hypotensive effect of nifedipine in SHR contrasts with the absence of enhanced reactivity
                                                                                               of vascular smooth muscle to NE. It remains to determine whether increased sympathetic
Guterbaum TG1, Braunstein TB2, Raunsø JR2, Torp-Pedersen CTP1, Domınguez HD3
                                                                          ´                    tone or greater NE release from sympathetic nerve endings is responsible for these major
1
  Cardiology Clinic Y, Research Unit, Bispebjerg University Hospital, Copenhagen,              abnormalities of SHR. (grant 1M0510)
           2
Denmark, Department of Medical Physiology. Renal and Cardiovascular Division,
Faculty of Health Sciences, Panum, Copenhagen, Denmark, 3Cardiology Department and
Lab. P, Gentofte University Hospital, Hellerup, Denmark                                                                                               PC.12
                                                                                               RETINAL TISSUE PRODUCES COMPARABLE RELAXATIONS ON RETINAL,
In diabetes, a major risk factor for cardiovascular disease, endothelial dysfunction is        CAROTID AND MESENTERIC ARTERIES
present before atherosclerosis becomes apparent. Hyperglycaemia confers a poor
prognosis during acute coronary syndrome and percutaneous revascularization. Nitric            Agus M, Takir S, Uydes-Dogan BS, Ozdemir O
oxide production by the endothelium is elicited by phosphorylation of the serin-1177 site      Istanbul University, Faculty of Pharmacy, Dept. of Pharmacology, Istanbul, Turkey
of the enzyme endothelial NO synthase (eNOS). This study aims to elucidate the ratio of
thr495 and ser1177 in settings of high glucose levels during ischemia and reperfusion in       Retinal tissue was recently shown to release a novel transferable relaxing factor, namely
human endothelial cells. Methods: Human Umbilical Vein Endothelial Cells (HUVECs) have         retinal relaxing factor (RRF) , which is probably involved in the regulation of retinal vessel
been incubated in 5 mM, 10 mM and 20 mM glucose medium respectively for 20h, under             tone. The characteristics of this factor were suggested to be different from those of other
normal oxygen conditions. Western blot of cell lysates (n 8) were probed for phospho-          well known vasoactive substances derived from the retina. Herein, we aimed to investigate
thr495-eNOS, phospho-ser1177-eNOS and total eNOS. Cells in parallel batches were               whether RRF also induced relaxations in other vascular smooth muscle preparations such
additionally co-incubated with hydrogen peroxide (H2O2, 400 mM for the last 30 minutes)        as carotid and mesenteric arteries in comparison with retinal artery. Bovine retinal, rat
as a model for the oxidative stress that occurs during reperfusion. Results: Cells incubated   carotid and rat mesenteric arteries were mounted in a multichamber wire myograph
in high glucose media did not show differences in eNOS phosphorylation sites while cells       system. After equilibration, preparations were precontracted with prostaglandin F2 (30
co-incubated with H2O2 displayed a four-fold increase in phospho-thr495-eNOS (p                   M) and bovine retina was placed in close proximity to the arteries. Retinal tissue
0.001 ) and the ratio phospho-thr495-eNOS to phospho-ser1177-eNOS increased by 50%             produced relaxations in bovine retinal arteries (98. 61 2.95%, n 12) as well as in rat
in the presence of H2O2, p 0,04. Total eNOS expression did not change under                    carotid (90.67 5.69%, n 7) and mesenteric (92.12 2.35%, n 11) arteries. Retinal
hyperglycaemic conditions. Conclusion: Endothelial dysfunction during reperfusion is           tissue-induced relaxations in carotid and mesenteric arteries were comparable to that of
probably related to oxidative stress rather than to the presence of hyperglycaemia per se.     obtained in retinal arteries. Retinal relaxation was immediate and generally displayed a
In this model, increase of the thr495-to-ser1177 ratio of eNOS phosphorylation accounts        biphasic character in retinal as well as in carotid and mesenteric arteries. Unlike retina,
for endothelial dysfunction.                                                                   choroidal tissue did not produce a relaxation in either of these arteries which excludes the
                                                                                                                    2007 ECCR Poster Presentations                                   813

possibility of a mechanical influence. Our results suggest that RRF may also play a role in    may act as a glucose-sensor in the wall of retinal microvessels, translating changes in
the regulation of peripheral vascular tone in addition to retinal arteries.                    Ca2 signals into changes in gene expression. Here we show that NFATc3 is expressed in
                                                                                               the endothelium of murine retinal microvessels and that it translocates to the nucleus upon
                                                                                               30 min stimulation with 20 mM glucose. Activation of NFAT is inhibited by a specific
                                                        PC.13                                  blocker of NFAT as well as by the ecto-nucleotidase apyrase, suggesting the involvement
                 ˙
EFFECT OF ALAGEBRIUM AND SILDENAFIL CITRATE ON ENDOTHELIAL                                     of extracellular nucleotides also in the endothelial response to high glucose.
SMOOTH MUSCLE RELAXATION IN CORPUS CAVERNOSUM

Sagdic Gamze1, Gurbuz Nilgun1, Sanli Ahmet2, Baykal Asli1, Usta M.F2                                                                                 PD.02
1
  Akdeniz University, School of Medicine, Department of Biochemistry, Antalya, Turkey,         THE A3 RECEPTOR MEDIATES ADENOSINE ENHANCEMENT OF MATRIX
2
  Akdeniz University, School of Medicine, Depatrment of Urology, Antalya, Turkey               METALLOPROTEINASE-9 SECRETION BY MACROPHAGES

Objectives: Erectile dysfunction (ED) associated with diabetes is caused by endothelial        Velot E1, Ernens I1, Haas B1, Jeanty C1, Longrois D2, Devaux Y1, Wagner DR1
                                                                                               1
dysfunction, neuropathy, and a decrease in Nitric Oxide (NO) production. Furthermore, it         CRP-Sante, Luxembourg, Luxembourg, 2Brabois Hospital and INSERM U684,
                                                                                                          ´
has been reported that the increased levels of advanced glycation end products (AGEs) in       Vandoeuvre-les Nancy, France
diabetes lead to the alteration of erectile function. AGE can quench nitric oxide (NO) both
in vivo and in vitro resulting in decreased endothelial smooth muscle relaxation. The aim      Purpose: Matrix Metalloproteinase-9 (MMP-9) plays an important role in ventricular
of this study was to investigate whether a combination of sildenafil and alagebrium could      remodeling after myocardial infarction. We have shown that adenosine inhibits the
enhance the erectile capacity in STZ-diabetic rats. Methods: Four groups of animals were       secretion of MMP-9 by human neutrophils. Whether adenosine modulates MMP-9
utilized: (1) control rats, (2) diabetic rats(40 mg/kg i.p.), (3) diabetes treated with        secretion by monocytes/macrophages is unknown. Methods: Cells from the monocyte-like
sildenafil(5 mg/kg/day p.o) and (4) sildenafil (5mg/kg/day p.o) Alagebrium(10 mg/kg/day        cell line THP-1 and monocytes from healthy volunteers were used. Monocytes were
p.o) for the final one month of two months of period of diabetes. At 2 months after i.p.       treated with adenosine (0.01 to 100 M) for 15 min and differentiated to macrophages
injection of STZ, all animals underwent cavernosal nerve stimulation (CNS) to assess           with 150 nM phorbol myristyl acetate for 48 hours (THP-1) or 50 ng/mL macrophage-
erectile function. 5-Hydroxymethylfurfural (5-HMF), Malondialdehyde(MDA) and cGMP              colony stimulating factor for 7 days (primary cells). Nucleofection was used to transfect
levels, and the protein expressions of nNOS and eNOS in cavernosal tissue were                 cells with either expression vectors or siRNA specific for each adenosine receptor (A1, A2a,
investigated. Results: The decreased levels of ICP (intra cavernosal pressure) in the          A2b, A3). MMP-9 secretion was assessed in culture supernatants using zymography.
STZ-diabetic rats was significantly improved after treatment with sildenafil Alagebrium,       Adenosine receptor expression was measured by quantitative PCR. Results: Adenosine
when compared to sidenafil treatment group. Additionally, combination therapy had the          time- and dose- dependently increased MMP-9 secretion by macrophages, reaching
same effect on 5HMF, MDA and cGMP levels, and eNOS, nNOS protein expression.                   three-fold above control (p 0.01). Dipyridamole and EHNA, inhibitors of adenosine
Conclusions: The combination of sildenafil alagebrium is more effective on the                 transferase and adenosine deaminase which increase levels of endogenous adenosine,
decreased erectile capacity through the cGMP/NO pathway than sildenafil treatment alone.       also increased levels of MMP-9. Adenosine induced a two-fold increase in A3 receptor
                                                                                               mRNA expression (p 0.004). The effect of adenosine on MMP-9 was replicated by the A3
                                                                                               agonist Cl-IB-MECA and over-expression of the A3 receptor, and abrogated by silencing of
                                                                                    PC.14      the A3 receptor. Conclusion: In sharp contrast to our previous results obtained in
ELEVATED L-NAME-SENSITIVE COMPONENT OF ENDOTHELIUM-                                            neutrophils, we report here that adenosine increases the secretion of MMP-9 by
DEPENDENT VASORELAXATION IN EXPERIMENTAL HYPERTENSION                                          macrophages. This increase appears to be mediated by the adenosine A3 receptor.
                                                                                               Therefore, antagonists of the A3 receptor may represent a new approach to prevent
Bernatova I, Puzserova A                                                                       ventricular remodeling.
Institute of Normal and Pathological Physiology, Slovak Academy of Sciences,
Bratislava, Slovakia
                                                                                                                                                        PD.03
The aim of this study was to investigate the magnitude of NG-nitro-L-arginine methyl ester     ENHANCED CONTRIBUTION OF NIFEDIPINE-SENSITIVE CALCIUM INFLUX
(L-NAME)-sensitive component of endothelium-dependent vasorelaxation and nitric oxide          TO SYMPATHETIC VASOCONSTRICTION IN SPONTANEOUSLY
(NO) production in adult normotensive Wistar-Kyoto rats (WKY), borderline hypertensive         HYPERTENSIVE RATS
rats (BHR, offspring of SHR dams and WKY sires) and spontaneously hypertensive rats
(SHR). Blood pressure (BP) of WKY, BHR and SHR (determined by tail-cuff) was 111 3,            Zicha J1, Pinterova M1, Liskova S2, Dobesova Z1, Kunes J1
140 4 (p 0.01 vs. WKY) and 184 6 mm Hg (p 0.01 vs. BHR), respectively. NO                      1
                                                                                                 CRC, Institute of Physiology AS CR, Prague, Czech Republic, 2Institute of
synthase activity (determined by conversion of [3H]-L-arginine to [3H]-L-citrulline) was       Pharmacology, School of Medicine, Comenius University, Bratislava, Slovakia
significantly higher in the aorta of BHR and SHR vs. WKY as well as in the left ventricle of
SHR vs. both BHR and WKY. L-NAME-sensitive component of endothelium-dependent                  Greater blood pressure (BP) decrease after acute blockade of voltage-dependent calcium
vasorelaxation was investigated in the pre-constricted femoral arteries using the wire         channels (VDCC) by nifedipine is an important abnormality in spontaneously hypertensive
myograph during isometric conditions as a difference between acetylcholine-induced             rats (SHR) characterized by high sympathetic tone. Norepinephrine (NE)-induced vasocon-
vasorelaxation before and after acute L-NAME pre-treatment (10-5 mol/l). Acetylcholine-        striction is based upon initial phasic contraction (mobilization of calcium stores) which is
induced vasorelaxation of SHR was significantly greater than that in WKY. Average value        followed by sustained tonic contraction (calcium influx through various calcium channels),
of L-NAME-sensitive component of vasorelaxation in WKY, BHR and SHR was 20 3%,                 which are both augmented in SHR. Our present study aimed to evaluate the contribution
29 4% (p 0.05 vs. WKY) and 37 3% (p 0.05 vs. BHR), respectively. There was a                   of calcium influx through VDCC to sympathetic vasoconstriction in SHR and WKY rats. The
significant positive correlation between BP and L-NAME-sensitive component of vasore-          influence of nifedipine (10-7 M) on phasic and tonic NE-induced contractions of femoral
laxation of the femoral artery (r 0.614, p 0.007, n 18). In conclusion, results suggest        arteries isolated from 14-week-old SHR and WKY were studied using Mulvany myograph.
the absence of endothelial dysfunction in the femoral artery of adult borderline and           BP effects of nifedipine injection (0.75 mg/kg) prior and after acute ganglionic blockade
spontaneously hypertensive rats and elevation of the L-NAME-sensitive component of             (pentolinium, 5 mg/kg) as well as those of pentolinium injection prior and after acute VDCC
endothelium-dependent vasorelaxation with increasing BP. Supported by the grants Nos.          blockade were determined in conscious SHR and WKY rats. Nifedipine did not influence
APVT-51– 018004 and VEGA 2/7064/27.                                                            phasic NE-induced contractions, but it caused a substantial reduction of tonic contractions,
                                                                                               which was greater in SHR than WKY vessels. Nifedipine-induced BP fall was almost
                                                                                               completely abolished by pentolinium pretreatment in both rat strains. On the contrary,
                                                        PD.01                                  nifedipine pretreatment prevented only 50% and 75% of BP decrease elicited by ganglionic
ACTIVATION OF CALCIUM/CALCINEURIN/NFAT SIGNALING PATHWAY IN                                    blockade in WKY and SHR, respectively. Augmented calcium influx through VDCC during
RETINAL MICROVESSELS                                                                           NE-induced tonic contractions of SHR arteries explains a more important contribution of
                                                                                               nifedipine-sensitive component to sympathetic vasoconstriction in SHR than WKY. (grant
        ¨
Nilsson-Ohman J, Gomez MF                                                                      1M0510)
Department of Clinical Science, Lund University, Lund, Sweden

Diabetic retinopathy is a major cause of blindness worldwide. Even though the                                                                                                      PD.04
mechanisms leading to diabetic retinopathy are not clear, hyperglycemia has been               ER IS A NEGATIVE REGULATOR OF THE PPAR IN VIVO
identified as an important risk factor. Retinal vasoconstriction and changes in Ca2
signaling have been shown to precede the onset of diabetic retinopathy. We have recently       Foryst-Ludwig A1, Clemenz M1, Hohmann S1, Hartge M1, Sprang Ch1, Frost N1, Krikov
shown that high glucose activates the Ca2 /calcineurin-dependent transcription factor          M1, Gustafsson J-A2, Unger T1, Kintscher U1
                                                                                               1
NFAT in vascular smooth muscle (VSM). This effect involves the local release of                  CCR, Charite-Berlin, Berlin, Germany, 2NOVUM, Karolinska Institute, Huddinge, Sweden
                                                                                                            ´
extracellular nucleotides, such as UTP, acting on P2Y receptors, leading to increased
intracellular Ca2 and subsequent activation of calcineurin. Once activated, NFAT has been      The estrogen receptors beta (ERbeta) and the PPARgamma are members of the nuclear
shown to promote VSM cell proliferation and to regulate the expression of inflammatory         hormone receptor family. The molecular mechanisms underlying PPARgamma function are
markers, such as IL-6, COX-2 and osteopontin. We therefore hypothesize whether NFAT            similar to those of ER-signaling. Importantly, PPARgamma is sharing an identical pool of
814          Hypertension Vol 50, No 4 October 2007


nuclear cofactors with ERbeta which provides a platform for mutual interactions between                                                                 PD.07
these two receptors. To determine the functional significance of ERbeta in cardiovascular      PERINDOPRILAT OVERCOMES THE INCREASES OF RAT KIDNEY CORTEX
and metabolic diseases in vivo we undertook studies in ERKO mice. When set on high             cAMP-PDE AND PDE3 ACTIVITIES CONSECUTIVE TO CONGESTIVE HEART
fat diet (59% kcal from fat for 12 weeks) female ERKO mice show higher endogenous
                                                                                               FAILURE
PPARgamma activity in gonadal fat tissue in comparison to wild-type (WT) mice in EMSA
assay. Moreover ERKO mice have increased PPARgamma-target gene expression in                   Charloux A2, Clauss F1, Piquard J2, Geny B2, Lugnier C1
gonadal fat (wt vs. ERKO) including lpl (1, 48 0,1 vs. 4,3 0,35) and adiponectin               1
                                                                                                 Institut Gilbert Laustriat CNRS UMR 7175, Illkirch, France, 2Service de Physiologie et
(0,92 0,14 vs. 1,42 0,28). Consistently, adiponectin serum level was significantly                                               ˆ
                                                                                               d’Explorations Fonctionnelles, Hopitaux Universitaires de Strasbourg, Strasbourg, France
elevated in ERKO mice. To investigate the mechanism of increased adiponectin
expression in vivo, we performed ChIP experiments in gonadal fat from HFD-fed ERKO             Congestive heart failure (HF) is characterized by left ventricular dysfunction, altered renal
and wt mice. PPARgamma binding to the adiponectin promoter was present in both                 metabolism and neuro-humoral activations of vasoconstrictor and antidiuretic systems
adipose tissues. However, binding of the coactivators SRC1 and TIF2 was enhanced in            such as the renin-angiotensin-aldosterone system and it is associated with cardiac
gonadal fat from ERKO mice, indicating that the absence of ERbeta in adipose tissue            natriuretic peptides hyporesponsiveness. We hypothesised that the inactivation of cAMP,
results in exaggerated coactivator binding to a PPARgamma target promoter. Together            a vasodilatatory intracellular second messenger, could result from an upregulation of renal
these data suggest that the coactivators SRC1 and TIF2 are involved in the negative            cAMP phosphodiesterase (PDE) activity during HF leading thus to renal retention and that
regulation of PPARgamma by ERbeta in-vivo. Molecular interactions between ERs and              inhibition of angiotensin converting enzyme could have protective effects related to PDE.
PPARs may help to understand the function of sex nuclear hormone receptors in                  Rat with congestive HF were obtained by coronary artery ligation and treated with
cardiovascular and metabolic diseases.                                                         Perindoprilat (AECI at 2 mg/kg/day) or placebo. After 1 month, plasmatic levels of ANP
                                                                                               were assessed. Cortical specific cAMP-PDE activities from kidneys were determined. ACEI
                                                                                               reduced rat mortality and improved their echocardiographic ejection fractions. Rats with
                                                                                               HF showed an increase in plasmatic ANP levels at 1 month of heart failure. Cortical renal
                                                                                               specific activities of both total cAMP-PDE and PDE3 were increased 1.6 fold, at 1 month
                                                                                               of HF as compared to sham (P 0.05). Interestingly, ACEI significantly reduced and
                                                                                               normalized specific activities of total cAMP-PDE and PDE3 in treated rats (P 0.05). This
                                                        PD.05                                  suggests that an alteration of renal vasodilatatory effect of cAMP, linked to cAMP-PDE
GPR30: NEW INSIGHTS IN CARDIOVASCULAR AND METABOLIC DISEASES                                   upregulation during HF, leads to alterations in filtration rate and diuretic response. AECI
                                                                                               may have a protective effect through a reduction of intra-renal angiotensin II which can
Meoli L, Isensee J, Nabzdyk C, Zazzu V, Ruiz Noppinger P                                       increase cAMP-PDE activities via intracellular calcium increase and MAP kinase cascade.
Center Cardiovascular Research, Charite University Medicine, Berlin, Germany

Estrogen (E2) regulates gene expression via nuclear estrogen receptors (ER and ) acting                                                               PD.08
as a ligand-activated transcription factors. However, accumulating evidence suggests that      THE IMPACT OF THE COMBINATION OF ALAGEBRIUM AND SILDENAFIL
the G-protein coupled receptor 30 (GPR30) binds E2 with high affinity and mediates rapid       ON iNOS EXPRESSION THROUGH MAP KINASE AND NFKB IN THE
estrogen signals in tissue culture. Although it is known that ER and ER are involved in        CORPUS CAVERNOSUM OF RATS WITH DIABETES MELLITUS
glucose, fatty acid and cholesterol metabolism, the role of GPR30 in these systems is
unknown. Currently we are analyzing the phenotype of GPR30 mutant mice generated by            Gurbuz N1, Sagdic G1, Sanli A2, Baykal A1, Usta MF2
                                                                                               1
targeted gene disruption (T181, Deltagen, Inc.) to decipher the physiological function of        Department of Biochemistry, School of Medicine, Akdeniz University, Antalya, Turkey,
                                                                                               2
GPR30. High levels of reporter gene expression were detected in small arterial vessels,          Department of Urology, School of Medicine, Akdeniz University, Antalya, Turkey
predominantly in the brain and the kidney vasculature, suggesting a regulatory role of this
G-protein coupled receptor in the cardiovascular as well as in the renal system. Preliminary   The dysfunction of the corpus cavernosum associated with diabetes mellitus (DM) results
phenotypic analysis showed elevated levels of plasma LDL as well as an increased heart         from the disordered endothelial smooth muscle relaxation and Nitric Oxide (NO) bioavail-
to the body weight ratio in homozygous mutant females, after 300 days of standard chow         ability. Advanced glycosylation end products (AGEs), which are accumulated in diabetic
diet. Remarkably, these alterations could not be detected in homozygous male mutants.          tissue, have been reported to induce iNOS expression through the NF-kappaB and p38
Based on the results obtained we will evaluate the role of this new estrogen receptor in       MAPK-dependent pathway. AIM: To investigate whether a combination of sildenafil and
the development and progression as well as the gender-specific aspects of cardiovascular       alagebrium, which is an AGE cross-link breaker, could have a synergistic effect on iNOS
and metabolic diseases.                                                                        expression via this pathway in the corpus cavernosum of STZ-induced DM rats. METHODS:
                                                                                               Wistar rats (18 –22 weeks old) were divided into four groups as (1) control; (2) STZ-diabetic
                                                                                               rats; STZ-diabetic rats treated with (3) sildenafil (5mg/kg/day p.o.) and (4) sildenafil
                                                                                               (5mg/kg/day p.o.) alagebrium(10mg/kg/day p.o.). Treatments started 1 month after STZ
                                                                                               injection. Blood glucose and AGE levels were determined by colorimetric methods. The
                                                                                               measurements of iNOS, MAP kinase and NFkB, and apoptosis were performed respectively
                                                                                               by western blot, immunohistochemistry and TUNEL. RESULTS: STZ-diabetic rats had an
                                                                                               increase in protein expressions of iNOS, MAP kinase and NFkB, and cavernosal cell
                                                        PD.06                                  apoptosis; when compared to control rats. In contrast to STZ-diabetic rats treated with
BIOCHEMICAL AND FUNCTIONAL ANALYSIS OF THE G PROTEIN-COUPLED                                   sildenafil alone, the combination therapy decreased these values similar to control rats.
RECEPTOR, GPR30 IN ENDOTHELIAL CELLS                                                           CONCLUSIONS: Our findings demonstrate that alagebrium may provide a novel therapeutic
                                                                                               approach in endothelial dysfunction, which is commonly seen with DM.
Zazzu V, Isensee J, Meoli L, Effertz K, Ruiz Noppinger P
           ´
CCR-Charite, Berlin, Germany
                                                                                                                                                                                    PE.01
Estrogen is a member of the steroid hormone family that acts through intracellular             DIET-INDUCED OBESITY (DIO) INDUCES ALTERATIONS IN THE
receptors and regulates many functionally unrelated processes in numerous tissues. The         MESENTERIC BED FUNCTION IN MICE
mode of action of two such estrogen-binding receptors, ER and ER , is reasonably well
understood. Novel receptors and novel forms of ERs have been postulated to mediate             Gil-Ortega M2, de las Heras AI1, Guzman R2, Aranguez I1, Ruiz-Gayo M2,
many of these signal transduction events. It has been suggested that rapid non-genomic         Fernandez-Alfonso MS1, Somoza B2
estrogen-mediated signaling may involve the G protein-coupled receptor, GPR30. This            1
                                                                                                 Instituto Pluridisciplinar, UCM, Madrid, Spain, 2Departamento de Farmacologıa,
                                                                                                                                                                            ´
protein has been reported to respond to estrogen at the plasma membrane or in the                         ´                       ´
                                                                                               Tecnologıa y Desarrollo Farmaceutico. Facultad de Farmacia. USP-CEU, Madrid, Spain
endoplasmic reticulum. This interaction occurs through unknown mechanisms leading to
the activation of downstream kinases and to the generation of second messengers via            The aim of this work was to determine the effect of a high fat diet on vascular reactivity
EGFR transactivation. In order to decipher the in vivo function of GPR30, we are currently     of mice resistance vessels. Five-week old male CB57BL/6J mice were assigned either to
analyzing GPR30 mutant mice generated by targeted gene disruption. Reporter gene               a low-fat (LF, 10 kcal% from fat, n 20) or to a high-fat diet (HF, 45 kcal% from fat, n 20)
expression shows predominant GPR30 expression in blood vessels of several organs of the        during 8 or 14 weeks. HF animals weighed significantly more than LF mice, either after
mouse including the heart and the kidney. To analyse the signaling pathway of GPR30 we         8 weeks (LF 25.47 0.57 g; HF              30.12 0.82g; p 0.05) or after 14 weeks of
screened a human heart cDNA library using the yeast-2-hybrid assay. Several interaction        treatment (LF 27.4 0.82 g; HF 33.6 1.8 g; p 0.05). Vascular reactivity was studied
partners were identified, among them PALS1-associated tight junction (PATJ). We are            in the isolated perfused mesenteric bed. There were no differences between groups in
currently characterising this interaction in mammalian cells by immunocytochemistry and        basal perfusion pressure, concentration-response curves to KCl (6 to 75 mM) or to
co-immunoprecipitation. Since GPR30 was originally cloned from human umbilical vein            noradrenaline (10-7 to 10-5 M). Endothelium-dependent relaxation to acetylcholine (Ach,
endothelial cells (HUVECs), we hypothesize that estrogen-mediated signalling in HUVECs         10-9-10-4M) was significantly higher in 8-week HF animals compared to the LF group (LF
might be mediated by GPR30. Therefore, the expression as well as localisation of GPR30            70.53 7.07%; HF 88.15 4.18%; p 0.05). This effect was lost after 14 weeks of
and PATJ are being evaluated at transcript and protein level in these cells upon E2            diet. Preincubation with L-NAME (10-4 M) or indomethacin (5x10-6M) reduced relaxation to
stimulation.                                                                                   Ach only in the LF group after 8 weeks of diet (L-NAME: LF 46.24 6.58%; HF
                                                                                                                     2007 ECCR Poster Presentations                                   815

72.87 12.76%; Indomethacin: LF 51 15.14%; HF               73.32 9.1%). These results          caused by unclear mechanisms. The effects of Ang II and ANP on adipocyte growth and
suggest a reduced contribution of nitric oxide and prostacyclin to Ach-induced vasodila-       the expression of AT1, AT2, NPRA, and NPRC were evaluated in VAT, cultured and freshly
tation together with a compensatory increase of endothelium-derived hyperpolarizing            isolated adipocytes. Methods: Primary human cultures and freshly isolated adipocytes
factor already at early states of DIO. Moreover, this compensatory effect is lost after 14     were incubated for 24 hours with physiological concentration of ANP (10-11M), Ang II
weeks of diet probably due to the initiation of vascular damage. (Supported by SAF             (10-12M), 8Br-cGMP (10-9M). Adipocyte proliferation was evaluated by manual count and
2006 – 02456, SAF 2005– 05180, FUSP-CEU. MG-O, RG and AH are recipients of a grant             BrdU incorporation. Gene expression of AT1, AT2, NPRA and NPRC in cells, adipose and
FUSP-CEU, and MEC, respectively).                                                              renal tissue (as control), was analyzed by Real-Time PCR. Results: Ang II has a
                                                                                               proliferative effect on cultured adipocytes whereas ANP had antiproliferative effect
                                                                                               (p 0.01). Gene expression in adipocytes showed that: AT1 was less expressed than in
                                                                                     PE.02     VAT (p 0.037); AT2 expression was absent. NPRA expression in adipocyte was similar but
CHARACTERIZATION OF AORTIC FUNCTION IN MICE DURING                                             lower than in VAT (p 0.05); VAT and cells showed equivalent NPRC gene expression level.
DEVELOPMENT OF DIET-INDUCED OBESITY (DIO)                                                      AT1 was the mediator of Ang II effect whereas ANP acted through NPRA via 8Br-cGMP that
                                                                                               produced similar effects (p 0.012). Discussion: Proliferation of human cultured adipo-
de las Heras AI1, Gil-Ortega M2, Guzman R2, Aranguez I1, Ruiz-Gayo M2, Somoza B2,              cytes is stimulated by Ang II, through AT1 and inhibited by ANP through NPRA. This growth
Fernandez-Alfonso MS1                                                                          control system is likely to be functional also in vivo because of the physiological Ang II and
1
  Instituto Pluridisciplinar, UCM, Madrid, Spain, 2Departamento de Farmacologıa,
                                                                             ´                 ANP concentrations used and the equivalent receptors gene expression.
           ´                       ´
Tecnologıa y Desarrollo Farmaceutico. Facultad de Farmacia. USP-CEU, Madrid, Spain

This work aims to assess the effect of a high fat diet on body and adipose tissue (AT)
weight and on aortic function. Five-week old male mice were assigned either to a low-fat
(LF, 10 kcal% from fat, n 20) or to a high-fat diet, (HF, 45 kcal%, n 20) for 8 or 14                                                                  PE.05
weeks. Contractile response of aortic segments to 75 mM KCl was not different between          HETEROLOGOUS REGULATION OF THE CB1 RECEPTOR (CB1) EXPRESSION
groups. The vasodilatory response elicited by acetylcholine (Ach, 10-9 to 10-4 M in rings
                                                                                               BY ATRIAL NATRIURETIC PEPTIDE (ANP) IN HUMAN ADIPOCYTES
precontracted with 10-7M phenylephrine) or to sodium nitroprusside (10-12 to 10-5 M) were
similar between groups. Exposure of rings to leptin (10-9 M) induced a slight and transitory
                                                                                               Sarzani R, Bordicchia M, Marcucci P, Cafarelli F, Giannini E, Pimpini L, Franchi E, Arma
relaxation which was similar between the types of diet. However, relaxation to Ach was
                                                                                                      `
                                                                                               P, Dessı Fulgheri P, Rappelli A
significantly reduced by leptin preincubation only in LF animals after 8 weeks of diet.
                                                                                               Department of Internal Medicine - University Politecnica delle Marche, Ancona, Italy
These data suggest that leptin induces an impairment of endothelium-dependent
relaxation in normal vessels which is lost after HF diet, probably due to the development
                                                                                               Introduction: The hyperactivity of the endocannabinoid system is associated with
of leptin resistance during early states of DIO. (Supported by SAF 2006 – 02456,
                                                                                               metabolic and cardiovascular risk. Rimonabant, a CB1 blocker, induced a significant
                           ´
SAF2005– 05180, Fundacion Universitaria San Pablo CEU. MG-O and RG are recipients of
                                                                                               reduction of visceral obesity and an improvement of cardiovascular risk in overweight
a grant FUSP-CEU. AH is recipient of a fellowship from MEC).
                                                                                               patients. We investigate the relationship between CB1 receptor and the cardiovascular
                                                   Lumbar AT               Periaortic AT       system through the analysis of CB1 regulation by ANP and Angiotensin II (AngII) that not
Weeks          Diet        Body Weight            (mg/mm tibia)           (mg/mm tibia)        only have effects on cardiovascular system but also have metabolic effects on adipocytes.
                                                                                               Methods: Primary cultures of subcutaneous and visceral adipocytes were treated with
8              LF          25.5   0.6g             6.9   0.5                 1.2   0.1
               HF          30.1   0.8g***         20.4   1.7***              1.6   0.2
                                                                                               physiological concentration of AngII (10-12 M), ANP (10-11 M), 8Br-cGMP (10-9 M). The
14             LF          27.4   0.8g             6.6   0.7                 1.4   0.2         expression levels of CB1, AngII receptors (AT1, AT2), and ANP receptors (NPRA, NPRC)
               HF          33.6   1.8g**          21.9   2.1***              2.3   0.4*        were measured with Taq-Man assay. Results: In paired patients ANP induced an increase
                                                                                               of CB1 expression in visceral adipocytes (P 0.007) and a down-regulation in subcuta-
                                                                                               neous primary cultures (P 0.005). AngII did not cause any variation in CB1 mRNA levels
                                                                                     PE.03     despite high expression level of AT1. Both NPRA and NPRC were expressed but ANP effect
ALTERATIONS IN KIDNEY ANTIOXIDANT STATUS AND LIPID                                             was mediated by NPRA because 8Br-cGMP reproduced the same effect of ANP (P 0.01).
                                                                                               ANP heterologous regulation was evident even with NPRA expression levels significantly
PEROXIDATION IN RATS WITH DIFFERENT HYPERTENSION MODELS
                                                                                               lower than in kidney and adipose control tissues (P 0.037;P 0.050). NPRC mRNA levels
Kipmen-Korgun D1, Hacioglu G2, Agar A2, Gumuslu S1                                             shown similar expression in cells and tissues. Discussion: ANP (cGMP mediated) opposite
1
  Akdeniz University, School of Medicine, Department of Biochemistry, Antalya, Turkey,         effects on CB1 gene expression regulation underlined the opposite behaviour of visceral
2
  Akdeniz University, School of Medicine, Department of Physiology, Antalya, Turkey            and subcutaneous adipocytes. ANP heterologous upregulation of CB1 could be involved in
                                                                                               a mechanism to maintain the balance between lipogenesis (endocannabinoid-dependent)
Oxidative stress plays a major role in the initiation and progression of cardiovascular        and the lipolysis induced by ANP.
dysfunction associated with diseases such as hypertension, diabetes mellitus, and chronic
heart failure. The aim of the present study was to investigate the effects of the different
hypertension models on antioxidant status and lipid peroxidation of rat kidneys. We also
compared the effects of different hypertension models and found the most effective
hypertension model, which alters these parameters in the kidney. Male Swiss-Albino rats                                                                                              PE.06
were divided into the following six groups namely; control, SHAM operated (SHAM), 1            THE IMPACT OF MACROPHAGES IN THE INITIATION OF
kidney-1 clamp (1K-1C), 2 kidney-1 clamp (2K-1C), deoxycorticosterone acetate (DOCA)           OBESITY-ASSOCIATED INSULIN RESISTANCE
(15 mg/kg twice per week for 10 weeks), N-omega-nitro-L arginine-methyl ester (L-NAME)
(50mg/kg/day for 75 days). The activities of catalase (CAT), Cu,Zn-superoxide dismutase        Hartge M1, Hess K2, Foryst-Ludwig A1, Clemenz M1, Sprang C1, Unger T1, Marx N2,
(Cu,Zn-SOD) and selenium-dependent glutathione peroxidase(Se-GSH-Px) and the levels of         Kintscher U1
glutathione (GSH), conjugated diene (CD) and thiobarbituric-acid-reactive substances           1
                                                                                                 Institute for Pharmacology, CCR, Carite Berlin, Berlin, Germany, 2Department of
                                                                                                                                        ´
(TBARS) were determined. The activities of Cu,Zn-SOD and CAT were found to be                  Internal Medicine II, University of Ulm, Ulm, Germany
decreased in all groups. Se-GSH-Px activities were significantly higher in 1K-1C, 2K-1C
and L-NAME groups compared with controls. The levels of GSH were significantly                 Obesity as a risk factor for type 2 diabetes mellitus is associated with insulin resistance
increased in 2K-1C and L-NAME groups, whereas decreased in 1K-1C and DOCA groups.              (IR). The infiltration of adipose tissue by macrophages was established as an important
CD and TBARS levels were increased in all hypertensive groups. Our results suggest that        pathophysiological mechanism of IR during obesity, however, it’s still unknown whether
different hypertension models have different degrees of influences on antioxidant defence      macrophages initiate or maintain IR. To analyze the temporal pattern of IR development
systems, and lipid peroxidation in the kidney.                                                 and macrophage infiltration into adipose tissue during obesity C57BL/6J mice were fed for
                                                                                               15 weeks with high fed diet (HFD) and compared to low fat diet-fed controls. Animals were
                                                                                               metabolically characterized (GTT, ITT) after 5, 10, and 15 weeks, and sacrificed
                                                             PE.04                             accordingly for gene expression analysis (qPCR) in adipose tissue. Insulin sensitivity of HFD
HUMAN VISCERAL ADIPOSE TISSUE EXPANSION: EFFECTS OF                                            mice was reduced by 11% 2,6 (AUC-ITT) at week 5 (p 0.01 vs. LFD), and was further
ANGIOTENSIN II (ANG II) AND ATRIAL NATRIURETIC PEPTIDE (ANP) ON                                decreased by 35% 6 at week 15 (p 0.01). Consistently, glucose tolerance was impaired
PERIRENAL ADIPOCYTES IN PRIMARY CULTURES                                                       by 59% 6 (AUC-GTT) at week 5 (vs. LFD, p 0.01), and remained at this level until week
                                                                                               15. Macrophage infiltration in adipose tissue was quantified qPCR of the macrophage
Sarzani R1, Marcucci P1, Bordicchia M1, Salvi F1, Espinosa E1, Mucci L1, Muzzonigro G2,        marker F4/80 in gonadal fat. F4/80 expression increased markedly at week 10: 5,2-fold
Minardi D2, Dessı Fulgheri P1, Rappelli A1
                 `                                                                                0,1 vs. LFD, (p 0.05) but was absent at week 5. Interestingly, an increase of the
1
  Department of Internal Medicine - University Politecnica delle Marche, Ancona, Italy,        T-lymphocyte marker CD3 was observed early after week 5 with further increase to
2
  Department of Urology - University Politecnica delle Marche, Ancona, Italy                   5,2-fold at week 15. The present study demonstrates that during the initiation of
                                                                                               obesity-mediated IR adipose tissue macrophages are absent making it unlikely that they
Introduction: Visceral adipose tissue (VAT) has a key role in the metabolism and               act as early pathogenetic mediators. Presence of T-lymphocytes at early stages of IR
cardiovascular system. VAT expansion by increase of size and number of adipocytes is           development may indicate their disease-initiating function.
816           Hypertension Vol 50, No 4 October 2007


                                                         PE.07                                   hybrydized to Affymetrix Rat 230 2.0 Genechips. Only results with at least a two-fold were
ACTIVATION OF THE CELLULAR CD40/CD40L SYSTEM IN PATIENTS WITH                                    further considered. Selected data were validated by Real Time RT-PCR, Western Blot and
METABOLIC SYNDROME                                                                               2D-gel plus LC/MS/MS. About 1000 mRNAs were affected by arteriotomy (p 0.0001) and
                                                                                                 were classified into 19 functional groups. Interestingly, changes of mRNAs and proteins
Natal C1, Restituto P2, Colina I2, Diez J1, Varo N1                                              (e.g. Arginase I and ACE) occurred also in contralateral uninjured carotids and in sham
1
  Centre for Applied Medical Research, Pamplona (Navarra), Spain, 2University Clinic of          control carotids, indicating their involvement in vascular physiology and in early
Navarra, Pamplona (Navarra), Spain                                                               inflammatory systemic reaction to surgery. Gene expression analysis at early stages after
                                                                                                 arteriotomy gives insights into molecular mechanisms of negative remodelling, providing
The proinflammatory dyad CD40/CD40L induces atherosclerosis but it has not been                  a basis for the identification of new potential therapeutic targets. Finally, changes in
                                                                                                 contralateral uninjured carotids and in sham control rats indicate systemic inflammatory
characterised in circulating cells of patients with metabolic syndrome (MS). We hypoth-
                                                                                                 reactions and vasocompensative mechanisms, suggesting caution in the choice of
esised that patients with MS have increased monocytes and platelets expression of
                                                                                                 reference control during experimental design.
CD40/CD40L system. 13 healthy subjects and 11 patients with MS (ATPIII criteria) provided
a sample of blood at the University Clinic of Navarra. Plasma levels of sCD40L were
measured by ELISA. Monocytes and platelets were obtained from peripheral blood. Gene                                                                     PF.02
(6h) and protein (24) expression of CD40/CD40L were determined by real time RT-PCR and
western-blot in monocytes. sCD40L content and release after stimulation with thrombin
                                                                                                 MOLECULAR CONSEQUENCES OF THE WNK1 FIRST INTRON DELETION,
were measured on platelets by ELISA. The relative expression of CD40L was significantly          RESPONSIBLE FOR FAMILIAL HYPERKALIEMIC HYPERTENSION (FHHT)
increased (CD40L/18S: 0.036 0.044) in monocytes from patients with MS compared with
controls (0.026 0.018). However, there were no differences in mRNA expression of the             Elvira E, Delaloy C, Houot AM, Hadchouel J, Jeunemaitre X
receptor CD40 between two groups. The protein expression of CD40 was significantly               INSERM U772 College de France, Paris, France
increased in patients with MS compared with healthy donors (arbitrary units: 1.31 0.92
vs. 0.79 0.49, respectively). In the studied group, patients had higher circulating sCD40L       The genetic study of Familial Hyperkalaemic Hypertension (FHHt), a rare form of
levels than controls (1.11 0.94 vs. 0.59 0.86 ng/ml, respectively p 0.05). In platelets,         hypertension, revealed a new mechanism implicated in blood pressure regulation, through
the thrombin-induced release of the soluble CD40L was higher in patients compared with           the identification of WNK1 and WNK4. These kinases, expressed in the renal distal tubule,
platelets obtained from controls (16.8-fold vs. 4.5-fold). These results suggest that            play a complex role in renal ionic transport regulation. Two proximal promoters, P1 and P2,
monocytes from patients circulate in a pro-inflammatory state which could be due to the          give rise to ubiquitous long isoforms (L-WNK1), while a third promoter, rP, gives rise to a
increased sCD40L release from platelets in these patients.                                       kidney-specific isoform (KS-WNK1). Mutations identified in FHHt are large deletions of
                                                                                                 intron 1. Using a transgenic model, we showed that these deletions induce an
                                                                                                 overexpression of both L- and KS-WNK1 and an ectopic generalised expression of
                                                                                                 KS-WNK1. These data suggest the presence in intron 1 of one ore several silencers and/or
                                                        PE.08
                                                                                                 insulators. Cross-species sequence comparison identified five candidate sequences (C1 to
NOVEL FLUORESCENT PROBE FOR LOW DENSITY LIPOPROTEIN, BASED                                       C5), all localized in the minimal deletion (22kb) observed in FHHt patients. The regulatory
ON THE ENHANCEMENT OF EUROPIUM EMISSION BAND                                                     effect of these candidate sequences was tested on WNK1 promoters by transient
                                                                                                 transfection and luciferase assays. Our results suggest that sequence C1 repress rP
Courrol LC1, Monteiro AM2, Silva FRO3, Gomes L4, Vieira Jr ND4, Gidlund MA2,                     activity. Its deletion would explain KS-WNK1 overexpression in the transgenic model.
Figueiredo Neto AM5                                                                              Sequence C5 can block renal enhancer activity and could therefore play the role of an
1
  Universidade Federal de Sao Paulo, Sao Paulo, Brazil, 2Universidade de Sao Paulo,
                              ˜           ˜                                 ˜                    insulator. Its deletion could explain the ectopic expression of KS-WNK1 in transgenic mice.
Instituto de Ciencias Biologicas, Sao Paulo, Brazil, 3Universidade de Sao Paulo, Escola
                                     ˜                                 ˜                         These results suggest that FHHt is the consequence of a transcriptional misregulation,
Politecnica, LSI, Sao Paulo, Brazil, 4Centro de Lasers e Aplicacoes, IPEN/CNEN-SP, Sao
                    ˜                                                                 ˜          linked to the suppression of at least one silencer and one insulator.
Paulo, Brazil, 5Instituto de Fisica, Universidade de Sao Paulo, Sao Paulo, Brazil
                                                      ˜          ˜

In aqueous solution, Europium trivalent ions exhibit weak luminescence due to their small                                                               PF.03
absorption cross section and strong energy transfer to surrounding water molecules.              A TWIN STUDY SHOWS STRONG HERITABILITY OF PLASMA ADIPONECTIN
However, when the Europium forms a complex with tetracycline, the ion luminescence can           LEVELS AND BODY MASS INDEX
be increased. Due to the different structural characteristics of LDL and oxidized LDL
(OxLDL) in the plasma (or, in vitro, in aqueous solutions), it is expected that their            Cesari M3, Narkiewicz K1, De Toni R3, Aldighieri E3, Williams CJ2, Rossi GP3
environments with respect to the neighboring water molecules (or, more generally, polar          1
                                                                                                   Depts. of Hypertension & Diabetology, University of Gdansk, Gdansk, Poland, 2Dept. of
molecules) are different. In this scheme, the EuTc luminescence in the presence of               Statistics, University of Idaho, Moscow, Idaho, USA, 3Dept. of Clinical & Experimental
aqueous solutions of LDL and OxLDL may be different and a fingerprint of the state of the        Medicine, Padova, Italy
lipoprotein. We report here the observation of the enhancement of Europium-tetracycline
complex emission in Low Density Lipoprotein (LDL) solutions. Europium emission band of           Background and aim: Adiponectin is suspected to exert anti-atherogenic, anti-
tetracycline solution containing Europium (III) chloride hexahydrate was tested to obtain        inflammatory and insulin-sensitizing effects. However, the relative importance of the
effective enhancement in the presence of native LDL and OxLDL. Europium emission                 genetic and the environmental factors in influencing plasma adiponectin levels (ADPN)
lifetime in the presence of lipoproteins was measured, resulting in a simple method to           remains unclear. Thus, we investigate whether ADPN and body mass index are genetically
measure the lipoproteins quantity in an aqueous solution at physiological pH. This method        determined. Methods: In a series of 60 pairs of healthy twins, we estimated genetic
shows that the complex can be used as a sensor to determine the different states of native       variance and heritability of ADPN and body mass index using both analyses of variance and
and oxidized LDL in biological fluids. Our proposal can be used as a complementary               path analysis methods. Twins were genotyped at two biallelic single nucleotide polymor-
technique to measure the amount of native and OxLDL from a previously total LDL sample           phisms (SNPs) at the gene encoding adiponectin: the 45 T/G (on exon 2) and the -11377
                                                                                    ˆ
isolated from a patient by using the conventional techniques. CNPq (Instituto do Milenio de      G/C (on the promoter). Results: 30 pairs of twins were monozygotic (MZ) and 30 were
Fluidos Complexos) and FAPESP (Optics Express, 15, 7066 (2007))                                  dizygotic (DZ). The mean ADPN ( SD) were 10.6 5.7 in MZ and 11.1 4.5 in DZ twins
                                                                                                 (NS). Three tests of heritability (within pair 1.13, p 0.0001; among components 1.62,
                                                                                                 p 0.005; intraclass correlation: 1.34, p 0.0001) consistently showed ADPN heritability.
                                                        PF.01                                    The preferred model of a likelihood-based analysis included an additive genetic influence
PROXIMAL AND DISTAL EFFECTS OF ARTERIOTOMY IN RAT CAROTIDS:                                      (A) and an individually unique environmental (E) influence for ADPN, accounting for 88%
AN EXTENSIVE MICROARRAY ANALYSIS OF GENE EXPRESSION                                              and 12% of ADPN variance, respectively. We found a significantly higher within pair
                                                                                                 difference of ADPN in DZ than in MZ pairs and in 45 T/G SNP discordant compared to
Forte A1, Finicelli M1, De Luca P4, Nordstrom I5, Quarto C2, Sante P2, De Feo M2,
                                                                    `                            concordant DZ twins indicating a significant effect of this adiponectin gene SNP on ADPN.
Onorati F3, Renzulli A3, Galderisi U1, Cipollaro M1, Hellstrand P5, Berrino L1, Rossi F1,        Conclusions: ADPN shows significant genetic variance and heritability, which is indepen-
           2            1                                                                        dent on BMI and partly accounted for by the 45 T/G but not the -11377 G/C adiponectin
Cotrufo M , Cascino A
1
  Dept of Exp. Medicine, Second University of Naples, Naples, Italy, 2Dept of                    gene SNP.
Cardiothoracic Sciences, Second University of Naples, Naples, Italy, 3Unit of Cardiac
Surgery, University of Catanzaro, Magna Graecia, Catanzaro, Italy, 4Biogem S.c.a.r.l.,
Ariano Irpino (AV), Italy, 5Dept of Exp. Medical Science, Lund University, Lund, Sweden                                                                                              PF.04
                                                                                                 GENE EXPRESSION PROFILING OF CELL CYCLE REGULATORS IN
The biology of injury-induced arterial restenosis is complex and not fully understood. We        ALDOSTERONE PRODUCING ADENOMA
analysed the transcriptome variations by microarray analysis at different times after
arteriotomy in inbred WKY rat carotids and in contralateral uninjured carotids, using as         Lenzini L, Giuliani L, Seccia TM, Rossi GP
reference the carotids from uninjured rats. The temporal profile of gene expression has          Department of Clinical and Experimental Medicine, Internal Medicine 4, University of
been analysed in carotids harvested 4h, 48h and 7 days after arteriotomy (n 15 carotids          Padova, Padova, Italy
for each time, divided in subgroups for biological and technical replicates), in contralateral
uninjured carotids and in sham control rats. Carotids from uninjured rats served as              Background: Several molecular mediators, such as cyclins, cyclin-dependent kinases
reference (n 10). Total RNA was amplified according to Eberwine’s method and then                (CDKs) and oncogenes affect cell cycle regulation and cell survival. Dysregulation of these
                                                                                                                    2007 ECCR Poster Presentations                                    817

proteins may determine an abnormal cell growth leading to tumour development. Hence,                                                                PF.07
we used an oligomicroarray-based technique to analyze the whole transcriptome of              MOLECULAR AND FUNCTIONAL INTERACTION BETWEEN ALDOSTERONE
Aldosterone Producing Adenoma (APA) and gather information on cell cycle regulators.          AND ANGII IN THE HEART
Methods: We compared APAs (n 16) with a pool of normal human adrenocortical tissues.
An unsupervised analysis was performed to outline the molecular signatures of the             Zhang A, Soukaseum C, Nguyen Dinh Cat A, Charbili V, Escoubet B, Jaisser F
tumours, using cluster and trend analysis as bioinformatics tools. The microarrays data       INSERM U772, College de France, Paris, France
were validated using quantitative real time RT-PCR. Results: An over-expression of
cyclin-dependent kinase 4 (CDK4) was found in APAs by oligomicroarray and confirmed by        Increasing evidence suggests that aldosterone (aldo), acting via the mineralocorticoid
real time RT-PCR, together with an under-expression of an inhibitor of this kinase,           receptor (MR), may mediate, even exacerbate the damaging effects of Angiotensin II (Ang
cyclin-dependent kinase inhibitor 2B (CDKN2B). At variance, Cyclin D was heterogeneously      II) but little is known about the local mechanisms underlying these interactions. Using a
expressed across APAs. Cullin 7, an ubiquitin cascade mediator was found over-                previously generated transgenic mouse model that allows conditional cardiac-specific
expressed. Among the genes involved in transcription, the nuclear receptor subfamily          overexpression of the human MR, we analyzed the effects of AngII signalling in the
(NR0B1) and sirtuin 4 (SIRT4) were over-expressed, while the inhibitor of DNA binding 1       presence of locally increased MR activation. Experiments were conducted in 4 mo old
(ID1) was under-expressed. Finally, two genes regulating DNA damage repair resulted           transgenic (Tg) and controls (Ctl) male mice treated or not with 200ng/min/Kg Ang II for
under-expressed in APAs: the thymine-DNA glycosylase (TDG) and the growth arrest and          2 months. Gene expression analyses showed a synergic effect between local MR and AT1
DNA-damage-inducible (GADD45B). Conclusions: While demonstrating the feasibility of a         activation with increased cardiac mRNA expression of collagen (col1a, relative value, Ctl
whole transcriptome approach for understanding the biology of APA development, these          : 1.06 0.2, Tg : 18.6 1.7, Ctl AngII : 32.6 17 Tg AngII : 87.4 15.3, n 8) and
results suggest a role of CDK4 and ubiquitin regulation in the growth of these tumours.       increased cardiac fibrosis (Ctl : 1 0.24, Tg : 1.8 0.2, Ctl AngII : 2.4 0.1, Tg AngII
Moreover, the under-expression of genes involved in DNA repair outlines the occurrence        : 3.7 0.7, n 4) and MMP2 activity (Ctl : 1 0.2, Tg : 1.45 0.23, Ctl AngII :
of genomic instability in the cell.                                                           1.63 0.26, Tg         AngII : 2.6 0.3, n 4). This may explain the diastolic dysfunction
                                                                                              evidenced in transgenic mice treated by the AngII. Increased expression of the gp91
                                                                                              subunit of NADHP oxydase,, (relative value, Ctl : 1.2 0.4, Tg : 8.4 1.9, Ctl AngII :
                                                                                              16.0 7, Tg AngII : 105.7 30, n 8) is observed as well as increased maximal activity
                                                        PF.05                                 of NADPH oxidase (Ctl : 49.5 4.4, Tg : 93.5 4.0, Ctl AngII : 104.4 6.5, Tg AngII
COMPARATIVE EXPRESSION OF EIGHT MAIN GENES OF THE                                             : 150.4 9.8, n 8). These results provide evidence that local interactions between AT1
                                                                                              and MR signalling pathways lead to increased fibrotic processes and cardiac remodeling,
RENIN-ANGIOTENSIN-ALDOSTERONE SYSTEM (RAAS) IN HUMAN KIDNEY
                                                                                              possibly as a consequence of their combined action on oxidative stress.
AND ADIPOSE TISSUES

Sarzani R1, Pietrucci F1, Salvi F1, Vannarelli A1, Bordicchia M1, Marcucci P1, Minardi D2,                                                            PF.08
Muzzonigro G2, Dessı Fulgheri P1, Rappelli A1
                     `
1
  Department of Internal Medicine - University Politecnica delle Marche, Ancona, Italy,       IMPORTANCE OF PROREGION IN RENIN BIOSYNTHESIS MATURATION
2
  Department of Urology - University Politecnica delle Marche, Ancona, Italy                  AND SECRETION

Published data on RAAS expression in human adipose tissue are fragmentary and a               Brakch N
correlation with tissues classically associated with the RAAS, such as the kidney, is         Division of Angiology & Vascular Medicine, Centre Hospitalier Universitaire Vaudois et
lacking. The aim of the study was to compare RAAS expression in renal and adipose                      ´
                                                                                              Universite de Lausanne, Lausanne, Switzerland
tissues. Samples of kidney cortex, renal medulla, perirenal visceral (VAT) and subcuta-
neous adipose tissue (SAT) were obtained from 11 patients who underwent nephrectomy.          Renin, an aspartyl protease, is cleaved from a larger precursor (called prorenin), into
Renin, (pro)renin receptor (PRR), AGT, ACE1, ACE2, AT1 and AT2 receptors, aldosterone         mature renin. Although the precise role of the renin proregion (1– 43) remains unknown,
synthase gene expression (all normalized for GAPDH) was evaluated by Real Time-PCR.           it has been reported that it could act as an auto-inhibitor. Works on other proteolytic
Kidney cortex is the main expression site for renin (15 times more than medulla; 60 times     enzymes showed that their proregion could act as chaperones. In the present study we
more than VAT, and over 300 times more than SAT) and ACE2 (9 times less expressed in          have investigated the possible role of the proregion of renin in the generation of
medulla and 6 times less in VAT and SAT). VAT is the highest expression site for AGT (4       enzymatically active renin. We have generated several pro-renin mutants: prorenin (1– 43),
times more than SAT; 2.5 times more than cortex; 8.8 times more than medulla), whereas        prorenin [1– 62], prorenin [ 43] and prorenin [ 62]. These constructs were expressed in
SAT expresses the highest levels of ACE1 and AT1, that were also more expressed in VAT        CV1 cells (constitutive pathway model) and AtT-20 cells (regulated pathway model) and we
than kidney cortex (4.6 times and 9 times, respectively). AT2 and PRR have low expression     measured the concentration of active renin molecules and renin activity in cell extracts and
levels both in adipose than renal tissues, whereas, aldosterone synthase did not resulted     culture media. Our data showed that the renin proregion influences renin generation in
expressed in either kidney or adipose tissue. VAT is likely to be an important source of      both an intra- and intermolecular way. The impact increases with the molecular size of the
angiotensin II (AngII) also because of the high co-expression of ACE1 with renin and PRR      proregion: prorenin (383 amino acids)        pro62 (62 amino acids)         pro43 (43 amino
expression. Adipose tissues also highly express AT1, implying important local AngII effects   acids). The addition of Br-CAMP to the cell culture hardly increased renin secretion.
and an increased number of binding sites for angiotensin receptor blockers in obesity.        Therefore, only a small amount of the cellular renin pool appears to be targeted to the
                                                                                              regulated pathway, and the remaining intracellular renin may be targeted to the
                                                                                              lysosomes. In conclusion, the proregion of renin plays a role in the generation of active
                                                                                              renin and this depends on the length of prorenin sequence. Since the mannose-6-
                                                                                              phosphate receptor mediates the transport of intracellular renin to the lysosomes, our data
                                                      PF.06                                   also support the possible transport by this receptor of extracellular circulating (pro)renin to
RELATIONSHIPS AMONG ENDOGENOUS OUABAIN, ALPHA-ADDUCIN                                         the lysosomes.
POLYMORPHISMS AND RENAL SODIUM HANDLING IN RAT AND HUMAN
PRIMARY HYPERTENSION
                                                                                                                                                                                     PF.09
Casamassima N, Manunta P, Delli Carpini S, Ferrandi M, Rastaldi MP, Bianchi G                 INTERLEUKIN-10 GENE POLYMORPHISM IS ASSOCIATED WITH
Nephrology, Dialysis, and Hypertension, Milano, Italy                                         ESSENTIAL HYPERTENSION IN TATARS FROM RUSSIA
Previous studies in cultured renal cells indicated that human and rat mutant forms of the     Timasheva Y.R.1, Nasibullin T.R.1, Zakirova A.N.2, Mustafina O.E.1
                                                                                              1
cytoskeletal protein adducin as well as nanomolar ouabain concentrations stimulate Na-K         Institute of Biochemistry and Genetics Ufa Science Centre RAS, Ufa, Russian
pump activity. To determine whether the common cellular effect of the two mechanisms          Federation, 2Bashkir State Medical University, Ufa, Russian Federation
translates into similar changes of blood pressure and renal Na handling in vivo, we studied
rats made hypertensive by chronic ouabain infusion, congenic normotensive rats harboring      Interleukin-10 is an immunoregulatory cytokine with multiple actions. Experiments on
the DNA sequence containing the “mutant” adducin and never treated patients with mild         knockout and transgenic mice showed that it had possible atheroprotective effect.
hypertension subdivided according to levels of endogenous ouabain or adducin genotype.        Essential hypertension is thought to be a multifactorial disorder involved in endothelial
Results: In ouabain hypertensive and congenic rats, renal immunohistochemistry and            dysfunction and atherosclerosis. Previously it was reported that -627 C/A promoter
western blotting of renal caveolae showed over-expression of the Na-K pump, and Na-H          polymorphism was associated with decreased interleukin-10 plasma level. The aim of the
exchange. Among 155 hypertensive patients, those with higher endogenous ouabain levels        present study was to evaluate the association between the interleukin-10 gene – 627 C/A
or that carried the mutated Trp ADD1 allele had higher blood pressure (p 0.002), plasma       promoter polymorphism and genetic susceptibility to essential hypertension. We geno-
Na (p 0.05), and increased proximal tubular reabsorption (p 0.001) as estimated by            typed 274 men with essential hypertension and 99 healthy subjects from the Tatar ethnic
endogenous lithium clearance. Following an acute Na load, a positive interaction between      group from Bashkortostan, Russia, by the polymerase chain reaction restriction fragment
circulating EO levels and the ADD1 genotype was detected (p 0.028). The hypertensive          length polymorphism method. Statistical analysis was performed using Fisher’s exact test.
effect of EO was evident in carriers of the Trp ADD1 allele and was not associated with       Odds ratios with 95% confidence intervals were calculated. The frequencies of IL-10
variation of tubular reabsorption. We conclude that Trp alleles and EO enhance renal          – 627C/A genotypes were the following: CC 71.72%, CA 27.27% and AA 1.01% in control
tubular sodium reabsorption and raise BP in humans and rats. These factors appear to be       group, CC 51.81 %, CA 39.8% and AA 8.39% in hypertensive patients. Thus, in Tatar
important in the genesis of essential hypertension.                                           ethnic group AA genotype was associated with increased risk of essential hypertension
818          Hypertension Vol 50, No 4 October 2007


(OR 8.98, P 0,006). Our results suggest a significant role for interleukin-10 in the            “no touch” technique preserves an intact function of vasa vasorum with communication
pathogenesis of cardiovascular disease.                                                         to the vein graft lumen. Evidence of restored transmural flow at graft implantation during
                                                                                                CABG operations, utilizing the “no touch” technique, will be presented. 1. Souza DS,
                                                                                                Johansson B, Bojo L, Karlsson R, Geijer H, Filbey D, et al. Harvesting the saphenous vein
                                                       PG.01                                    with surrounding tissue for CABG provides long-term graft patency comparable to the left
DOSE-RELATIONSHIPS OF BLOOD PRESSURE AND METABOLIC EFFECTS                                      internal thoracic artery: results of a randomized longitudinal trial. J Thorac Cardiovasc Surg
OF HYDROCHLOROTHIAZIDE, CHLORTHALIDONE AND                                                      2006;132(2):373– 8.
BENDROFLUMETHIAZIDE MONOTHERAPY: A META-ANALYSIS

Peterzan MA1, Chaturvedi N2, Hughes AD2                                                                                                                                               PG.04
1
  Imperial College School of Medicine, London, United Kingdom, 2Imperial College School         PROGNOSTIC EFFECT OF PLASMA ADIPONECTIN (ADPN) IN NON
of Medicine St Mary’s Campus, Cardiovascular Science, National Heart and Lung                   DIABETIC HIGH-RISK PATIENTS
Institute, International Centre for Circulatory Health, London, United Kingdom
                                                                                                Maiolino G2, Cesari M2, Antezza K2, Zanchetta M1, Pedon L1, Pessina AC2, Rossi GP1
                                                                                                1
Context: It is widely believed that doses of thiazide diuretics that have BP-lowering             Division of Cardiology, Cittadella Hospital, Italy, 2Dept. of Clinical & Experimental
efficacy are lower than those that provoke unwanted metabolic side-effects and                  Medicine, University of Padova, Padova, Italy
neuroendocrine activation. However, rigorous evidence for this is lacking. Objective: To
estimate the dose-related effect-size of hydrochlorothiazide (H), chlorthalidone (C) and        Background and aim: Cross sectional studies have suggested an inverse association of
bendroflumethiazide (B), each used as monotherapy versus placebo control, on BP and             ADPN, a circulating adipocytokine with anti-atherogenic, anti-inflammatory and insulin-
serum parameters. Data sources: Ten electronic databases; relevant references. Trial            sensitizing effects, with atherosclerosis. However, its prognostic value is under debate as
inclusion criteria: Randomized, double-blinded, parallel placebo arm, 2 different               prospective cohort studies have suggested an adverse effect of high plasma (p)ADPN.
monotherapy thiazide dose arms, duration of follow-up 4 weeks, baseline washout of              Thus, we investigated if pADPN levels predict cardiovascular (CV) events and mortality in
medication 2 weeks. Titration intervals in step-up protocols 4 weeks for all                    non-diabetic high-risk coronary artery disease (CAD) patients. Methods: We selected for
participants, regardless of BP response. Results: Of 28 included trials, BP data were           this study 674 non-diabetic patients of the GENICA study who underwent coronary
available in 25 (5196 subjects). The dose-hypotensive-relation was quantifiable over the        angiography for suspected CAD. We measured pADPN and, after splitting of our patients
entire range for H, but not for the lower parts of the relations for C and B. The lowest H      in high and low pADPN subgroups, we examined the incidence of CV deaths and events
dose with hypotensive effect was 6.25mg/day (effect-size (systolic/diastolic) -3.15/-           at follow-up. Results: The median pADPN in this population was 6.6 g/mL (IQR
2.44mmHg); this was also the highest dose free of hypokalaemic and hyperuricaemic               4.4 –10.8). Complete follow-up data were obtained in 100% of patients. After a median
effect. Conclusions: Low dose H reduces BP and is largely free of side-effects, suggesting      follow-up of 1341 days (range 9 –2018 days) 43 (6.4%) CV deaths were recorded. While
that the mechanism of action at this dose is less reliant on induced natriuresis per se and     Kaplan-Meier analysis suggested an adverse impact of high pADPN on survival,
less hindered by secondary activation of the renin-angiotensin-aldosterone system.              multivariate Cox regression analysis, where potential confounders including ongoing drug
Additional studies of low dose C and B are required to establish the dose response              treatment were taken into due consideration, showed an association of low pADPN with
relations for BP, metabolic side-effects and neuroendocrine activation.                         CV death (HRR 0.50, CI 0.2– 0.9, p 0.029) and events (HRR 0.50, CI 0.3– 0.9,
                                                                                                p 0.028). Conclusions: In non-diabetic high-risk CAD patients pADPN above the median
                                                                                                was associated with roughly a 50% reduction of the risk of CV death and events
                                                      PG.02                                     independently of major CV risk factors.
ENDOTHELIAL DYSFUNCTION AND CARDIOVASCULAR REMODELING IN
RHEUMATOID ARTHRITIS
                                                                                                                                                         PG.05
Myasoedova EE                                                                                   VISCERAL ADIPOSITY AS MAIN DETERMINANT OF ARTERIAL STIFFNESS
Ivanovo State Medical Academy, Ivanovo, Russian Federation                                      IN A NORMOTENSIVE POPULATION AT LOW CARDIOVASCULAR RISK:
                                                                                                EVIDENCE FROM A NEW ECHO-TRACKING APPROACH
To study the endothelial and myocardial alterations and the pathogenesis of cardiovascular
remodeling 163 patients with rheumatoid arthritis (RA) and 94 matched controls were             Malshi E, Morizzo C, Muscelli E, Kozakova M, Camastra S, Guerra E, Nannipieri M,
investigated. Endothelium-dependent (EDVD), endothelium-independent brachial vasodi-            Ferrannini E, Palombo C
latation (EIVD), carotid intima-media thickness (IMT), left-ventricular myocardium mass         Department of Interna Medicine, University of Pisa, Pisa, Italy
index (MMI) and diastolic function, desquamated endothelial cells (DE), L-arginine, NOx
plasma levels were assessed. Results: High MMI was seen in 73.5% of RA patients and             Introduction: Increasing evidence highlights the role of obesity in anticipating atheroscle-
2.1% of controls (p 0,001), diastolic dysfunction (DD) was seen in 57.4% and 1.1%,              rotic vascular disease. Aim of the study: to evaluate determinants of carotid stiffness in
respectively (p 0,001). DD in RA was associated with EDVD (r – 0.45, p 0.05), and RA            a population with a wide BMI and age range. Materials and methods: 145 subjects [(age
activity (r 0.40, p 0.05). IMT 0.9 mm was present in 73.3% of RA patients and 1.07%             range 18 – 64 years), Framingham Risk Score (FRS range 1–17%)] were selected.
of controls (p 0.001). IMT was associated with RA duration (r 0.35, p 0.05), RA activity        According to the WHO classification, study population included 44 normal weight, 30
(r 0.46, p 0.05), rheumatoid factor (RF) (r 0.43, p 0.05), -globulin level (r 0.94,             overweight and 71 obese subjects [(NL, OW and OB: mean age 40 11.4, 44.4 10 and
p 0.05), MMI (r 0.31, p 0.05), left-atrium length (r 0.39, p 0.05). 34% of RA                   39.2      12 yrs yrs, mean BMI 22.6         1.9, 28.2     1.6 and 39.5     6.2 kg/m2). 106
patients had subnormal EDVD and 54% had increased EIVD compared with 1.1% and                   subjects underwent (OGTT), and blood samples for glucose, insulin, C-peptide, HDL, total
4.3%, in controls (p 0.001). EDVD in RA was associated with RF (r – 0.30, p 0.05). DE           cholesterol and triglyceride levels collected. Arterial mechanics was evaluated in common
and NOx levels in RA were higher than in control (p 0,05). L-arginine level didn’t differ       carotid artery (CCA) by a cardiovascular ultrasound system (Aloka SSD-5500) implemented
between groups. L-arginine in RA correlated to RF (r – 0.32, p 0.05), while NOx                 with E-tracking providing calibrated diameter-derived pressure curves. Indices carotid
correlated with the number of cardiovascular risk factors (r 0.53, p 0.05), DE – to MMI         stiffness (pressure-strain elastic modulus (EP), index, arterial compliance (AC), pulse
(r 0.40, p 0.05). RA is associated with cardiovascular disease. Vasospasm, L-                   wave velocity (PVW)) and augmentation index (AIx), were obtained. Results: in the overall
arginine–NO disbalance, endothelial cell desquamation predispose to myocardial and              population WS correlated directly with age (r 0,407, p 0,0001); MBP (r 0,330,
vascular remodeling.                                                                            p 0,001); CCA IMT (r 0,219, p 0.05), waist-to-hip ratio (WHR, r 0,511, p 0,001);
                                                                                                AUC for C-peptide (in 60 subjects, r 0,359, p 0,005), FRS (r 0,319, p 0.05), and
                                                                                                inversely with HDL cholesterol (r -0,231, p 0.05). In stepwise regression analysis, WHR
                                                                                     PG.03      remained the main independent determinant of carotid PWV (r2 0,396, p 0.01).
PRESERVATION OF VEIN GRAFT MICROCIRCULATION WITH THE “NO                                        Conclusions: in an otherwise healthy obese population, visceral adiposity is independently
TOUCH”-HARVESTING TECHNIQUE                                                                     associated with subclinical involvement of large artery function.

Dreifaldt MO1, Dashwood MR2, Loesch A3, Norgren L4, Souza DS1
1                                                       ¨
  Dept. of Cardiothoracic Surgery, University Hospital, Orebro, Sweden, 2Dept. of Clinical                                                               PG.06
Biochemistry, Royal Free Hospital, London, United Kingdom, 3Dept. Anatomy, Royal Free           LOCAL ARTERIAL WAVE SPEED AT CAROTID ARTERY LEVEL IS
                                    4                                      ¨
Hospital, London, United Kingdom, Dept. of Surgery, University Hospital, Orebro,                REPRESENTATIVE OF CAROTID-FEMORAL PULSE WAVE VELOCITY AND
Sweden                                                                                          AORTIC STIFFNESS: EVIDENCE BY A NEW ECHO-TRACKING TECHNIQUE

Vein grafts are, in spite of their limited patency, still the most used conduits for coronary   Guerra E1, Malshi M1, Morizzo C1, Palombo C1, Kozakova M1, Florescu M1, Vinereanu D1
                                                                                                1
artery bypass grafting (CABG). Recently a prospective randomised study was published              Department of Internal Medicine, University of Pisa, Pisa, Italy, 2University Hospital of
where vein grafts harvested with the new “no touch” technique showed the same patency           Bucharest, Bucharest, Romania
as the Internal Mammary Artery, known to be the most superior conduit for CABG.1) During
conventional harvesting of vein grafts the surrounding tissue, and partly the adventitia, is    Background: Carotid-femoral (CF) pulse wave velocity (PWV) is a marker of aortic stiffness
stripped off and the vasa vasorum is damaged. This might cause ischemia to the vein             and predictor of cardiovascular risk. A new ultrasound technique of real-time arterial
tunica media, which causes neointimal hyperplasia, a major cause of vein graft failure. The     waveform analysis (“E-track”, Aloka, Tokyo, Japan) has been developed. By this method
                                                                                                                     2007 ECCR Poster Presentations                                   819

arterial stiffness parameters can be obtained: pressure-strain elastic modulus (EP),                                                                 PG.09
stiffness index ( ) and a single point local wave speed (WS). Aim:: evaluate whether           VASOMOTOR DYSFUNCTION AND INCREASED ENDOTHELIN-1 LEVELS IN
common carotid artery WS may be representative of CF-PWV. Methods: 150 patients, [110          PATIENTS WITH INSULIN RESISTANCE
free of cardiovascular disease (NL, mean age 52 10 yrs), 40 hypertensive (HBP, mean
age 55 11 yrs)] underwent common carotid artery scanning by high resolution linear US          Jurka A1, Tretjakovs P1, Reihmane D1, Bormane I1, Mikelsone I1, Stifts A2, Aivars JI1,
probe (7.5 to 10 MHz, Aloka SSD-5500) for E-track evaluation and EP, stiffness index,          Pirags V2
and WS were obtained. CF-PWV and carotid-radial PWV (CR-PWV) were assessed by                  1
                                                                                                 University of Latvia, Riga, Latvia, 2Pauls Stradins Clinical University Hospital, Riga,
Complior device (Artech, Paris, France). Results: HBP showed significantly (p 0.001)           Latvia
higher WS (6,5 1,2 vs 7.6 1.6), CF-PWV (9,4 1,6 vs 11,2 2,3), CR-PWV (10,4 1,3 vs
11 1,5), EP (120,4 49 vs 180 71) and           stiffness index (9,2 3,7 vs 11,4 3,9,           Aim: to evaluate the relationship between endothelin-1 (ET-1), insulin resistance and
p 0.01). WS and CF-PWV significantly correlated with age and mean blood pressure               cutaneous vasomotor responses (endothelial-dependent vasodilatation and peripheral
[(r 0,454 and r 0,461, p 0.001, for CF-PWV); (r 0,535 and 0,552, p 0.01, for WS)].             sympathetic failure: noradrenergic control of smooth muscle cells – vasoconstriction and
WS, EP and stiffness index correlated directly with CF-PWV (r 0.484, p 0.001; r                neuropeptides induced vasodilatation) in metabolic syndrome (MS) patients with insulin
0.467 and 0.364, p 0.005, respectively) but not with CR-PWV, in the entire population.         resistance. MS patients with insulin resistance, but without hypertension were divided into
Conclusions: Common carotid artery stiffness indices and local wave speed appear               two groups: 18 patients with type-2 diabetes mellitus (without insulin therapy and
representative of aortic stiffness.                                                            pronounced diabetic complications) (DM) and 18 patients without DM. 18 healthy subjects
                                                                                               were selected as controls (C). The study groups were matched for age and sex. Insulin
                                                                                               resistance was measured by HOMA-IR method and ET-1 was measured by ELISA. We
                                                                                               recorded changes in laser Doppler flux (LDF; PeriFlux 4001, Perimed) on the foot. Basal
                                                                                    PG.07      LDF (b-LDF), postocclusive hyperemia (m1-LDF), vasoconstrictor response (v-LDF) to deep
AORTIC STIFFNESS AND KIDNEY FUNCTION                                                           inspiration on the pulp of the toe; and heat ( 44°C; PeriTemp 4005) induced hyperemia
                                                                                               (m2-LDF) on the dorsum of the foot. Results: b-LDF and local skin temperature did not
Baumann M1, Liu D1, Kribben A2, Heemann U1, Witzke O2                                          differ between the study groups (p 0.05). Only the patient group with diabetes
1
  Dept. of Nephrology, Klinikum rechts der Isar, TU Munich, Munich, Germany, 2Dept. of         demonstrated a significant diminution in v-LDF compared to that of the group of healthy
Nephrology, University Hopsital Essen, Essen, Germany                                          subjects (p 0.05). m1-LDF was decreased in both patient groups in comparison with the
                                                                                               controls (p 0.05), but only in diabetics the decrease of m2-LDF was significant (p 0.05).
Background: Increased stiffness of central arteries is independently associated with           ET-1 levels were elevated (p 0.05) in both patient groups. Our findings show that MS
reduced creatinine clearance in subjects with mild to severe renal insufficiency, indicating   patients with insulin resistance have significant cutaneous vasomotor dysfunction and
that kidney diseases may interact with large conduit arteries. We aimed to investigate         elevated ET-1 levels.
whether glomerular function or renal blood flow are associated with aortic stiffness in
individuals with normal renal function. Methods: We investigated the 24-hour ambulatory
                                                                                                                                                        PH.01
blood pressure in subjects (n 78) who were prepared for living kidney donation and
calculated the ambulatory aortic stiffness index (AASI). Glomerular function was deter-        CURRENT STANDARD TREATMENT DOES NOT PREVENT RECURRENT
mined by 2x2-hour creatinine-clearance and renal blood flow (RBF) by para-aminohippur-         CARDIOVASCULAR EVENTS IN SUBJECTS WITH A POSITIVE FAMILY
clearance (PAH), both corrected for body surface area (BSA). AASI quartiles were compared      HISTORY
to renal function and blood pressure. Data is presented as mean SD. Results:
Distribution of subjects to the quartiles was as follow: 1st: 17, 2nd: 39, 3rd: 17 , 4th:5.    Meijer Z1, Mulders T1, van der Donk C1, Kroon AA1, Ferreira I2, Stehouwer CDA1,
AASI ranged from 0.14 to 0.99. Quartiles did not significantly differ regarding age, sex,      Pinto-Sietsma SJ1
                                                                                               1
body mass index (BMI), hypertension and smoking status. Creatinine-clearance did not             Academic Hospital Maastricht, Maastricht, Netherlands, 2University Maastricht,
differ between the quartiles (1st:106.9 19.7, 2nd:97.6 18.6, 3rd:96.4 14.1,                    Maastricht, Netherlands
4th:103.0 31.2 ml/min). PAH-clearance was negatively correlated with AASI (-0.18). The
1st quartile demonstrated significantly higher PAH clearance than the 3rd quartile             Background: Premature cardiovascular disease (PCVD) is treated in the same manner as
(448.3 55.8 and 401.3 66.2 ml/min/BSA; P 0.05). Blood pressure did not differ                  CVD of advanced age. However, different (possibly genetic) mechanisms may underlie
between the quartiles (1st:125.7 10.1, 2nd:122.9 12.5, 3rd:123.0 9.8,                          PCVD, to which current medical treatment is not targeted. This suggests that particularly
4th:126.9 13.2 mmHg). Discussion: Aortic stiffness determined by AASI is not associ-           in families with PCVD, standard treatment may be insufficient to prevent recurrent events.
ated with glomerular function, but is negatively correlated with RBF in a cohort with normal   Hypothesis: Subjects with PCVD and a positive family history for PCVD are more likely to
renal function. We conclude that a relationship exists between kidney function and aortic      have recurrent cardiovascular events despite standard medical treatment. Methods: We
stiffness predominantly on the level of extraglomerular renal microcirculation.                retrospectively investigated 151 subjects with PCVD. All subjects had to have a first
                                                                                               cardiovascular event at least 5 or more years ago. PCVD was defined as an event before
                                                                                               the age of 50 for men or 55 for women. A positive family history was defined as a
                                                                                               first-degree family member with PCVD before the age of 55 for men or 60 for women. We
                                                                                               used multinomial logistic regression to analyze the relationship between a positive family
                                                       PG.08                                   history and the reoccurrence of 1 or 2 or more cardiovascular events. Results: Subjects
ACUTE AND SHORT TERM EFFECTS OF ATORVASTATIN ON RENAL                                          with a positive family history were more likely to have a single recurrent event [age and
HEMODYNAMICS, TUBULAR FUNCTION, VASOACTIVE HORMONES, BLOOD                                     sex adjusted odds ratio (OR): 1.8, 95% confidence intervals (CI): 0.9 –3.7]. This relationship
PRESSURE AND PULSE RATE IN HEALTHY, NORMOCHOLESTEROLEMIC                                       was even stronger for subjects who had 2 or more recurrent events. (OR: 2.8; 0.9 – 8.7).
HUMANS                                                                                         Besides, there was no significant difference between subjects with or without a positive
                                                                                               family history in treatment duration (9.3 4.8 years versus 9.2 5.2 years, respectively).
Paulsen L1, Holm C1, Starklint J1, Bech JN2, Pedersen EB1                                      Conclusion: Subjects with PCVD and a positive family history are not protected for the
1
  Department of Medical Research, Holstebro, Denmark, 2Department of Medicine,                 recurrence of cardiovascular events. This might imply that different pathophysiological
Holstebro, Denmark                                                                             mechanisms are active within families with PCVD, which are not treated to day.

Background: Statins have a beneficial effect on cardiovascular morbidity and mortality
due to reduction in plasma cholesterol. However, statins seem to have effects beyond                                                                   PH.02
cholesterol-lowering and we hypothesize that these effects might be attributed to an effect    ASSOCIATIONS BETWEEN LOWER LIMB ARTERIAL CALCIFICATION AND
on renal function. Methods: In two randomised, placebo-controlled, double blinded and          MARKERS OF CORONARY AND PERIPHERAL ARTERIAL DISEASE IN
cross-over studies we measured the effects of atorvastatin (AS) on renal function. In Study    SOUTH ASIAN AND EUROPEAN MEN
1 19 subjects received either 80 mg AS as a single dose or placebo. In Study 2 20 subjects
received either 80 mg AS or placebo daily for four weeks. In both studies glomerular           Joshi M1, Tillin T1, Malik I1, Coady E1, Raynor S1, Mayet J1, Wright A1, Kooner J1, Shore
filtration rate (GFR), renal plasma flow (RPF), plasma concentrations of atrial natriuretic    A2, Thom S1, Chaturvedi N1, Hughes A1
                                                                                               1
peptide, brain natriuretic peptide, aldosterone, vasopressin, angiotensin II and renin were      International Centre for Circulatory Health, St Mary’s Hospital, Hammersmith Hospital &
determined. Results: In Study 1 the fractional excretion of sodium (FENa) (P 0.035) and        Imperial College London, London, United Kingdom, 2Peninsula Medical School, Exeter,
the diastolic BP (DBP) (P 0.024) were significantly decreased in the AS treated group.         United Kingdom
In Study 2 we found that the 24-h urinary sodium excretion (24h U-Na) was significantly
lower (P       0.005) in the AS treated group. In both studies we found no significant         Background: South Asians have high rates of heart disease and stroke, but low rates of
differences in GFR, RPF or vasoactive hormones. CONCLUSIONS: A single dose of AS               peripheral arterial disease (PAD). The reasons for this are unclear. Objectives: To
significantly decreases FENa and DBP in healthy humans. The decrease in FENa by AS might       investigate ethnic differences in arterial calcification in the legs and associations between
be due to the antihypertensive effect of AS. The effect of AS on FENa was very modest and      leg and coronary calcification and other measures of PAD. Methods: Cross sectional study
disappeared after 4 weeks treatment. Treatment with AS for four weeks decreases 24h            of 83 European and 84 South Asian men, with and without coronary heart disease,
U-Na in healthy humans. The beneficial effect of statin could not be attributed to changes     recruited from West London cardiology clinics and general practices. Participants
in renal haemodynamics in healthy subjects.                                                    underwent blood tests and routine measurements, including mean ankle brachial pressure
820          Hypertension Vol 50, No 4 October 2007


index (ABI). Multislice computed tomography measured coronary and leg calcification.            (43.08 3.37) compared to those without atrial fibrillation (52.80 2.36, P 0.03). VEGF
Femoral intima-media thickness (IMT) was measured using ultrasound. Results: South              expression on the vasculature did not differ but was increased on cardiomyocytes in
Asian men without diabetes (n 128) were significantly less likely than Europeans to have        patients with atrial fibrillation (2.82 0.18) compared to those without atrial fibrillation
leg calcification above the median score of 70 Hounsfield Units (odds ratio (OR): 0.29 (95%     (2.42 0.15), P 0.08. VEGFR2 expression on the vasculature and cardiomyocytes did not
CI: 0.14,0.62) p 0.001). The ethnic difference was partially attenuated on adjustment for       differ between groups. Atrial fibrillation is not associated with an alteration in atrial
age and systolic blood pressure (adjusted OR: 0.43 (0.18,1.03) p 0.058). There was no           appendage vascular morphology. HIF-1 expression is reduced in patients with atrial
ethnic difference in leg calcification in men with diabetes (n 35) (age adjusted OR: 1.10       fibrillation with a trend for an increase in VEGF but no change in VEGFR2 expression.
(0.18, 6.88). In all men, leg calcification was positively correlated with coronary
calcification (rho:0.67, p 0.001) and femoral IMT (rho:0.49, p 0.001) and negatively
correlated with ABI (rho:-0.25, p 0.001). Conclusions: For the first time we show that in                                                                PH.06
men without diabetes, South Asians have less leg arterial calcification than Europeans and      URINARY ALBUMIN EXCRETION IN CHRONIC HEART FAILURE PATIENTS
that leg calcification is strongly associated with coronary calcification and with markers of
PAD in all men. Medial calcification may mask ethnic differences in people with diabetes.       Reznik EV, Gendlin GE, Volinkina VM, Storozhakov G.I.
                                                                                                Russian State Medical University, Moscow, Russian Federation

                                                       PH.03                                    The relation between urinary albumin excretion (UAE) and chronic heart failure (CHF) has
INCREASED URIDINE ADENOSINE TETRAPHOSPHATE CONCENTRATIONS                                       not been well studied. We investigated 70 CHF patients with I-IV NYHA functional class
IN PLASMA OF JUVENILE HYPERTENSIVES                                                             without primary renal, endocrine, autoimmune and oncological diseases. Medians of age,
                                                                                                LVEF (Simpson), UAE (immunoturbidimetric assay) and glomerular filtration rate (GFR;
Jankowski V, Zidek W, Jankowski J                                                               MDRD1 equation) were 61.5 years, 28.9%, 45 mg/24-h and 72.5 ml/min/1.73 m2
Charite (CBF) Med Klinik IV, Berlin, Germany                                                    respectively. Microalbuminuria (MAU) was revealed in 69.4% (95% confidence interval
                                                                                                57.1– 81.2), high-normal albuminuria (HNA; 15–30 mg/24-h) in 22.6% and macroalbu-
Uridine adenosine tetraphosphate (Up4A) was been recently characterized as a potent             minuria in 4.8% of patients. GFR was 60 – 89 ml/min/1.73 m2 in 46.8% of MAU patients
vasoconstrictor. Up4A occurs in plasma from healthy subjects at concentrations sufficient       and 56.3% of HNA patients and 60 ml/min/1.73 m2 in 31.9% of MAU patients. GFR
to cause strong vasoconstrictive effects. In this study, Up4A concentrations in plasma from     (median 68.1 vs. 78.7 ml/min/1.73m2, p 0.046), basal duplex ultrasonography renal
juvenile hypertensives and normotensives were determined. Up4A was purified to                  blood flow (423.9 vs. 598.4 ml/min/1.73m2, p 0.043), Kansas City Cardiomyopathy
homogeneity by preparative reverse phase high performance liquid-chromatography                 Questionnaire (KCCQ) severity of shortness of breath (4 vs. 5, p 0.047), swelling in feet,
(HPLC), affinity HPLC and analytic reverse phase HPLC from deproteinized plasma of              ankles or legs (4 vs. 5, p 0.039), fatigue (3 vs. 4, p 0.049) and KCCQ summary score
juvenile hypertensives and normotensives. Mean total plasma Up4A concentration was              (65 vs. 83, p 0.018) were lower in patients with UAE 45 mg/24-h in comparison with
significantly increased in juvenile hypertensives compared to juvenile normotensives.           others. However these patients didn’t differ in age, arterial pressure, LVEF and cardiac
Accordingly, Up4A showed a significant association with juvenile hypertension. Plasma           output. Subjects with MAU and macroalbuminuria had increased al-cause mortality in
Up4A concentrations correlated with left ventricular mass and intima media wall thickness       comparison with others (p 0.042). UAE did not correlate with any echocardiographic
in the hypertensives. Since the increased intima media thickness may be related to              parameters. Thus, elevated UAE is widespread and not early sing of renal dysfunction in
proliferative effects of Up4A, we studied the effects of Up4A on human VSMC proliferation.      CHF patients. UAE is not associated with cardiac function, but with renal function, renal
Circulating levels of Up4A are strongly associated with juvenile hypertension. The              hemodynamics, severity and prognosis of CHF.
endothelium-derived vasoconstrictor Up4A may contribute to the early development of
primary hypertension and is moreover an important risk factor of juvenile hypertension.
                                                                                                                                                          PH.07
                                                                                                CALCIUM AND PHOSPHORUS METABOLISM DISTURBANCES IN PATIENTS
                                                                                     PH.04      WITH CHRONIC HEART FAILURE (CHF): ASSOCIATION WITH CLINICAL
GLUCOSE METABOLISM IN PATIENTS WITH ACUTE MYOCARDIAL                                            SYMPTOMATIC, RENAL FUNCTION AND PROGNOSIS
INFARCTION
                                                                                                Reznik EV, Gendlin GE, Volinkina VM, Storozhakov GI
Ahmad S, Ashawesh K, Abdulqawi R, Warner DP, Barton DM                                          Russian State Medical University, Moscow, Russian Federation
Princess Royal Hospital, Telford, United Kingdom
                                                                                                Abnormalities of calcium and phosphorus metabolism are common and important in CKD
BACKGROUND: Diabetes and impaired glucose tolerance are associated with increased               pts, but they have not been described well in CHF. In 70 I-IV NYHA class pts with chronic
mortality in patients with acute myocardial infarction. Objective: to investigate the           left ventricular systolic dysfunction (EF 40%) without renal, endocrine, autoimmune,
prevalence of newly diagnosed abnormal glucose tolerance (AGT) in patients with acute ST        oncological, inflammatory bowel and bone diseases, we measured serum calcium
elevation myocardial infarction (STEMI), without known diabetes mellitus (DM), in a district    corrected for albumin, phosphorus and 24-h urinary excretion of these substances.
general hospital. Methods: all patients admitted to the coronary care unit with acute           Hypocalcaemia ( 8.6 mg/dl) was revealed in 28.5%, hypercalcaemia ( 10.3 mg/dl) in
STEMI and without known DM, over a 6 month period, were prospectively enrolled. A               10.8%, hyperphosphatemia in 5.7%, phosphaturia in 52.7% of pts. Urinary excretion of
standardised oral glucose tolerance test (GTT) with 75 g of glucose, 5 days post admission,     calcium was increased in 4.3% and was decreased in 30.1% of pts. The severity of main
was performed. Results: 54 patients (14 women) were included; 6 patients were                   CHF symptoms assessed by Minnesota Living with Heart Failure and Kansas City
previously diagnosed with type 2 DM; 5 patients died and 7 transferred to tertiary centre       Cardiomyopathy Questionnaires was higher (p 0.05), creatinine clearance was lower
for further management, within 4 days post admission. 32 patients underwent GTT five            (p 0.039) in pts with calcium-phosphorus product (Ca*P) 33.1 mg2/dl2 (median) in
days post admission; 8 of 32 (27%) patients had impaired glucose tolerance with mean            comparison with others. Maximal physical capacity (treadmill test) was lower, elevated
2-h post-load glucose concentration of 8.9 mmol/L and 2 of 32 (6%) had previously               urinary albumin excretion was more often in pts with phosphaturia in comparison with
undiagnosed DM. Conclusion: AGT is common in patients with acute STEMI; the lower               others (p 0.039 and 0.005). Ca*P 33.1 mg2/dl2 and phosphaturia were associated with
prevalence rate of AGT observed in our study, compared to the previous, can be explained        increased all-cause mortality (p 0.021 and 0.033). There were not differences in age,
by the fact that the most serious patients admitted with STEMI (22% of the total number),       body mass, echocardiographic parameters between pts with and without Ca*P 33.1 or
who were high likely to have AGT, were excluded (because they either died or were               phosphaturia. Thus, disturbances of calcium and phosphorus metabolism are common and
transferred to the tertiary centre). AGT early post-MI is a strong risk factor for further      associated with clinical symptoms, mortality and renal dysfunction in CHF patients.
cardiovascular events and should be considered a novel target for secondary prevention.         Screening and correction of these abnormalities may improve quality and length of life in
                                                                                                CHF patients, especially those with renal dysfunction.

                                                         PH.05
ALTERATIONS IN MYOCARDIAL VASCULAR MORPHOLOGY AND                                                                                                                                   PH.08
EXPRESSION OF KEY ANGIOGENIC FACTORS IN PATIENTS WITH ATRIAL                                    THE PREVALENCE OF HYPERGLYCEMIA AND LIPID DISORDERS IN
FIBRILLATION                                                                                    PATIENTS WITH PRIMARY HYPERALDOSTERONISM AND ESSENTIAL
                                                                                                HYPERTENSION: A CASE-CONTROL STUDY
Ben Darif A, Jeziorska M, Prendagast B, Malik R
University of Manchester, Manchester, United Kingdom                                            Matrozova J2, Steichen O1, Jeunemaitre X1, Plouin PF1
                                                                                                1
                                                                                                  Hopital Europeen Georges Pompidou, Paris, France, 2Clinical Center of Endocrinology,
                                                                                                   ˆ           ´
The left atrial appendage was obtained prior to coronary bypass surgery in 53 patients with     Akad. Ivan Penchev, Sofia, Bulgaria
Ischaemic heart disease, with (n 20) and without (n 33) atrial fibrillation. Microvascular
density (96.63 5.07 v 105.26 4.86, P 0.347), Endothelial cell area (200.96 14.51 v              Background: The association of primary hyperaldosteronism (PH) with hyperglycemia and
175.81 8.42, P 0.201), Luminal area (162.17 14.27 v 187.34 17.39, P 0.387 and                   the metabolic syndrome has been reported. The aim of this retrospective study was to test
percentage of closed vessels (76.44 1.99 v 74.80 1.32, P 0.419) did not differ                  this association in a large cohort of patients with PH and with essential hypertension (EH).
significantly between patients with and without atrial fibrillation. There was a significant    Methods: We retrieved the records of 237 cases with PH (142 with aldosterone-producing
reduction in the percentage expression of HIF-1 in patients with atrial fibrillation            adenomas/APA/ and 95 with idiopathic hyperaldosteronism/IH/) and matched for age and
                                                                                                                   2007 ECCR Poster Presentations                                   821

sex 1176 controls with EH, selected from the ARTEMIS-database. We compared history,          overall in-hospital mortality in older patients with hip fracture (HF). Methods: In 240
BP levels, BMI, fasting blood glucose and lipids between cases with APA and IH and           consecutive patients (age 81.9 7.8 years; 72% females) with low-trauma HF we
between cases and their matched controls. Results: Compared to cases with APA, cases         measured serum concentrations of cardiac troponin I (cTnI), leptin, PTH, 25(OH)vitamin D,
with IH were older (51 vs 47 yrs, p 0.01), more frequently male (73 vs 55%, p 0.006),        estimated glomerular filtration rate (GFR) and prospectively collected clinical data. Results:
had a more frequent history of diabetes (18 vs 5%, p 0.002), higher BMI (27.7 vs 26.3        Myocardial injury (cTnI 0.06 g/L) was present in 29% patients, 25(OH)vitamin D
kg/m2, p 0.008) and triglycerides (1.15 vs 0.84 mmol/l, p 0.001) and lower HDL-
                                                                                             deficiency ( 50nmol/L) in 81%, elevated PTH ( 6.5pmol/L) in 53%, the leptin level was
cholesterol levels (1.16 vs 1.25 mmol/l, p 0.011). Cases with APA had higher BP levels
and lower BMI than matched controls, but did not differ in metabolic parameters. Cases         12 ng/ml in 48.3%, and a history of coronary artery disease (CAD) in 19.7%. Patients
with IH also had higher BP levels than matched controls, but did not differ in BMI and       with myocardial injury compared to the rest of the cohort had decreased leptin
metabolic parameters. Conclusion: Our findings do not confirm the hypothesis that high       concentrations (14.6 16.2 vs 22.5 28.5ng/ml; p 0.030) and increased PTH levels
aldosterone and/or low potassium levels have a diabetogenic effect. It seems unlikely that   (8.6 6.5 vs 6.1 4.2pmol/L; p 0.001). In multivariate analysis, myocardial injury was
PH and EH differ in the prevalence of the metabolic syndrome. Studies on the metabolic       independently associated only with leptin 12ng/ml (OR 2.3; p 0.018), history of CAD
consequences of PH should carefully distinguish between cases with IH and APA, since         (OR 2.3; p 0.035) and older age (for every 5 years after 60 OR 1.5; p 0.002). Patients
they have different metabolic profiles.                                                      who died compared to survivors had significantly higher levels of PTH (10.1 4.0 vs
                                                                                             6.6 5.0pmol/L; p 0.028). After adjusting for age, sex, history of CAD, vitamin D status
                                                        PH.09                                and GFR, only elevated PTH (OR 19.5; p 0.010) and cTnI 1 g/L (OR 8.4; p 0.028)
LEPTIN AND PARATHYROID HORMONE (PTH) AS PREDICTORS OF                                        were associated with all-cause mortality. Conclusions: In older HF patients serum leptin
MYOCARDIAL INJURY AND ALL-CAUSE MORTALITY IN OLDER PATIENTS                                    12ng/ml is an independent predictor of myocardial injury, while elevated PTH and cTnI
WITH OSTEOPOROTIC HIP FRACTURE                                                               ( 1 g/L) are independent predictors of all-cause mortality.

Fisher A, Davis M, Goh S, Southcott E, Srikusalanukul W, Hickman P, Smith P
The Canberra Hospital and Australian National University Medical School, Canberra,
ACT, Australia

Aim: To examine the relationship between leptin and PTH, important regulators of the
cardiovascular system and bone homeostasis, and peri-operative myocardial injury and
Author Index From the 12th Annual Meeting of the ECCR

Aalkjaer C, 5.03                      ˜
                                 Cairrao E, PC.06                        Ferrandi M, PF.06                  Hughes AD, 01.05, 02.06, 04.02,
Abdulqawi R, PH.04               Calippe B, 2.05                         Ferrannini E, PG.05                  PC.01, PC.04, PG.01, PH.02
Achard JM, 5.02                  Camastra S, PG.05                       Ferreira I, 05.05, PH.01            ¨
                                                                                                            Hutter J, PB.07
Agabiti Rosei E, 5.04            Cambien C, 4.01                         Figueiredo Neto AM, PE.08
Agar A, PE.03                    Cambien F, W3.03                        Finicelli M, PF.01                 Inguimbert N, 03.03, PB.02
Agus M, PC.12                    Cano V, PB.08                           Fischer JA, 1.03                    ˜
                                                                                                            Inigo C, PA.01
Ahmad S, PH.04                   Casamassima N, PF.06                    Fisher A, PH.09                    Isensee J, PD.05, PD.06
Aivars JI, PG.09                 Cascino A, PF.01                        Fliegner D, PA.09                  Ittner L, 1.03
Alakakone A, 4.02                Castoldi G, PA.06                       Florescu M, PG.06                  Iturrioz X, PB.02
Aldighieri E, PF.03              Cesari M, PF.03, PG.04                  Forte A, PF.01                     Jaisser F, PF.07
Allemann Y, 3.01                 Chambers J, 4.02                              ˜
                                                                         Fortuno MA, PA.01, PA.02
Allison W, PA.07                 Chanoine S, 02.03, PB.06                Foryst-Ludwig A, 02.07, 03.04,     Jankowski J, 02.01, PC.02,
              ´
Altarche-Xifro W, PA.08, PB.01   Charbili V, PF.07                          PD.04, PE.06                       PH.03
´
Alvarez E, PC.06                 Charloux A, PD.07                       Franchi E, PE.05                   Jankowski V, 02.01, PC.02,
Amouyel P, PA.10                 Chaturvedi N, 01.05, 04.02,             Frost N, PD.04                        PH.03
Ansar S, 1.06                      PG.01, PH.02                          Frugiere A, 3.03                   Jardel AL, PB.06
Antezza K, PG.04                 Cieniewski-Bernard C, PA.10             Funke-Kaiser H, 3.02               Jeanty C, PD.02
Aranguez I, PE.01, PE.02         Cipollaro M, PF.01                                                         Jeunemaitre X, 05.02, PF.02,
Ariel I, PA.05                   Claperon C, PB.02                       Galderisi U, PF.01                    PH.08
Arma P, PE.05                    Clauss F, PD.07                         Gassmann M, 2.04                   Jeziorska M, PH.05
Arnal JF, 2.05                   Clemenz M, 02.07, PD.04, PE.06          Geldyyev A, 1.01                   Jordan J, 3.01
Artuc M, PB.03                   Coady E, 04.02, PH.02                   Gendlin GE, PH.06, PH.07           Joshi M, PH.02
Ashawesh K, PH.04                Cody R, 3.01                            Geny B, PD.07                      Jurka A, PG.09
Azimonti S, W3.04                Colina I, PE.07                         Gianni M, PA.04
Azizi M, PB.04                   Cotrufo M, PF.01                        Giannini E, PE.05                  Kameni-Tcheudji JF, PC.03
                                 Courrol LC, PE.08                       Gidlund MA, PE.08                  Kaschina E, 02.07, 03.04, PA.08,
Bader M, 05.01, PB.03            Curato C, 03.04, PA.08, PB.01           Gilibert S, 4.04                     PB.01, PB.10
Barlassina C, W3.04              Cusi D, W3.04                           Gil-Ortega M, PE.01, PE.02         Kemnitz UR, 02.07, 03.04,
Barton DM, PH.04                                                         Giuliani L, PF.04                    PB.10
Bataillard A, 4.04               Dashwood MR, PG.03                      Godes M, 3.02                      Keravis T, PC.03
Bathula R, 1.05                  Davis M, PH.09                          Godsland I, 1.05                   Kintscher U, 02.07, PD.04,
Baufreton C, PC.08               De Feo M, PF.01                         Goh S, PH.09                         PE.06
Baumann M, PG.07                 de las Heras AI, PB.08, PE.01,          Gomes L, PE.08                     Kipmen-Korgun D, PE.03
Baumgart P, W3.03                   PE.02                                Gomez MF, PD.01                    Kjolby M, PB.09
Bayard F, 2.05                   de Leeuw PW, 03.01, 04.06               Gotthold C, W2.03                  Kleine-Katthoefer P, W3.03
Baykal A, PC.13, PD.08           De Luca P, PF.01                        Gourdy P, 2.05                     Kojsova S, 02.02
Bech JN, 04.03, PG.08            De Mey JGR, 05.04, W2.04                Grandi AM, PA.04                   Kold-Petersen H, 5.03
Belin de Chantemele E, 02.03,    De Toni R, PF.03                        Grimm D, PA.03                     Koleganova N, 01.01, PA.05
  PB.06                          Del Vecchio L, W3.04                    Gross ML, 01.01, PA.05             Kooner J, 04.02, PH.02
Belloni AS, 4.05                 Delaloy C, PF.02                        Gruber C, W2.03                    Kozakova M, PG.05, PG.06
Ben Darif A, PH.05               Delli Carpini S, PF.06                  Grzesiak A, 03.04, PA.08, PB.01,   Kribben A, PG.07
Bergaya S, 5.02                  Dentali F, PA.04                          PB.10                            Krikov M, 03.02, PB.10, PD.04
Berger I, 1.01                        `
                                 Dessı Fulgheri P, PE.04, PE.05,         Guerra E, PG.05, PG.06             Krivokharchenko A, 5.01
Bernatova I, PC.14                  PF.05                                Guihot AL, 02.03, PC.08            Krøjgaard C, 2.06
Berrino L, PF.01                 Devaux Y, 01.02, PD.02                  Guillot AL, PB.06                  Kroon AA, 03.01, 04.06, 05.05,
Bianchi G, PF.06                 di Gioia C, PA.06                       Gumuslu S, PE.03                     PH.01
Bie P, PB.09                       ´
                                 Dıez J, PA.01, PA.02, PE.07             Gurbuz N, PC.13, PD.08             Krucker T, 2.04
Birka C, PB.01                   Dobesova Z, PC.11, PD.03                Gurzu B, PC.10                       ¨
                                                                                                            Kruger F, 2.07
Bischoff H, PB.07                Doerdelmann C, W3.03                    Gustafsson JA, PD.04               Kuhn G, 2.04
Blanchard A, PB.04                    ´
                                 Domınguez HD, PC.09                     Guterbaum TG, PC.09                Kunes J, 02.02, PC.05, PC.11,
Boari GEM, 5.04                    ¨
                                 Dordelmann C, 4.01                      Guzman R, PE.01, PE.02               PD.03
Bodineau L, 3.03                 Douin-Echinard V, 2.05                                                     Kunz TH, 1.03
Bombardi C, PA.06                Dreifaldt MO, PG.03                     Haas B, PD.02                      Kurtz S, W3.03
Bondke A, W2.03                  Duelsner A, W2.03                       Hacioglu G, PE.03                  Kwitek AE, 4.04
Bordicchia M, PE.04, PE.05,      Dumitriu IL, PC.10                      Hadchouel J, 05.02, PF.02
  PF.05                          Dumont O, 02.03, PC.08                  Hagedorn C, W3.03                  Laffargue M, 2.05
Bormane I, PG.09                                                         Hallberg A, 03.04, PB.03           Lahera V, PA.02
Born W, 1.03                     Eckert S, 3.01                          Hansky B, 3.01                     Lalanne F, PB.04
Brakch N, PF.08                  Edvinsson L, 1.06                       Hartge M, PD.04, PE.06             Larsen P, 5.03
Brand E, 04.01, W3.03            Effertz K, PD.06                        Heemann U, PG.07                   Laurell H, 2.05
Brand-Herrmann SM, 04.01,        Elvira E, PF.02                         Heinzer S, 2.04                    Lauridsen TG, 4.03
  W3.03                          Engeli S, 3.01                          Hellstrand P, PF.01                Lavrador I, PC.06
Brands J, W1.03                  Ernens I, 01.02, PD.02                  Henrion D, 02.03, PB.06, PC.08     Le Bec A, PC.03
Braunstein TB, PC.09             Escoubet B, PF.07                       Hess K, PE.06                      Lebeau L, PC.03
Briggs Boedtkjer DM, 5.03        Espinosa E, PE.04                       Hickman P, PH.09                   Lehmann K, W2.03
Brinckmann MP, PA.08, PB.01      Etienne-Selloum N, PA.07                Hilgers RHP, W2.04                 Lenzini L, PF.04
Brouckaert P, PC.07                                                      Hillmeister P, W2.03               Li J, 03.04, PA.08, PB.01
Bursztyn M, PA.05                Faramarzi S, PA.03                      Hohmann S, PD.04                   Liebeskind U, 3.01
Buschmann IR, W2.03              Farkas S, PA.11                         Hojna S, 2.02                      Liskova S, PC.05, PD.03
Buys E, PC.07                    Fassot C, 3.03                          Holm C, PG.08                      Litvin K, 2.06
                                 Faure S, 5.02                            ¨
                                                                         Hortnagl H, 3.02                   Liu D, PG.07
Cachofeiro V, PA.02              Fernandez-Alfonso MS, PB.08,            Houben AJHM, 4.06                  Llorens-Cortes C, 03.03, PB.02
Cafarelli F, PE.05                 PE.01, PE.02                          Houot AM, PF.02                    Loesch A, PG.03

                                                                   822
                                                                                                     Author Index               823

Longrois D, PD.02                 Nordstrom I, PF.01                Ruello A, W3.04                  Thom S, 04.02, PH.02
 ´
Lopez N, PA.01, PA.02             Norgren L, PG.03                  Ruiz Noppinger P, PD.05, PD.06      ¨
                                                                                                     Thone-Reineke C, 3.02
Loufrani L, 02.03, PB.06, PC.08   Nussberger J, 4.04                Ruiz-Gayo M, PB.08, PE.01,       Thuillez C, PA.10
Luft F, 3.01                                                          PE.02                          Tillin T, 01.05, 04.02, PH.02
Lugnier C, PA.07, PC.03, PD.07    Oakland KA, PC.04                 Ruiz-Noppinger P, W2.03          Timasheva YR, PF.09
                                  Okada M, PB.02                     ¨
                                                                    Rußmann C, 4.01                  Timm M, 03.04, PA.08, PB.01
Maigret B, PB.02                  Onorati F, PF.01                                                   Tinel H, PB.07
Maiolino G, PG.04                 Orosz SZ, PA.11                   Sagdic G, PC.13, PD.08           Tiret L, 4.01
    ´
Mako E, PA.11                     Ozdemir O, PC.12                  Salvi F, PE.04, PF.05            Toelle M, 2.01
Malik I, 04.02, PH.02                                               Sandner P, PB.07                 Tooke JE, W1.04
Malik R, PH.05                    Page C, 1.05                      Sanli A, PC.13, PD.08            Tordoir J, 3.01
Malshi E, PG.05                   Palombo C, PG.05, PG.06                `
                                                                    Sante P, PF.01                   Torielli L, W3.04
Malshi M, PG.06                   Panerai R, 1.05                   Santos-Silva AJ, PC.06           Torp-Pedersen CTP, PC.09
Mancini M, PA.06                  Patel S, 1.05                     Sarzani R, PE.04, PE.05, PF.05   Travaglini C, PA.06
Maniero C, 4.05                   Paul M, 01.04, 04.01, PA.03,      Sassard J, 4.04                  Tretjakovs P, PG.09
Manunta P, PF.06                    W3.03                           Satchell SC, W1.04               Tripodi G, W3.04
Marc Y, 3.03                      Paulis L, 2.02                    Schachter M, PC.04
Marcucci P, PE.04, PE.05, PF.05   Paulsen L, PG.08                  Schefe J, 3.02                   Ulbrich C, PA.03
Marelle L, PB.04                  Pechanova O, 02.02                Scheffers I, 3.01                Ulmann-Schuler A, 2.04
Maresca AM, PA.04                 Pedersen EB, 04.03, PG.08         Schiffers PMH, 5.04              Unger T, 02.07, 03.02, 03.04,
Marques B, PC.06                  Pedon L, PG.04                    Schini-Kerth V, PA.07              PA.08, PB.01, PB.03, PB.10,
Marti HH, 2.04                    Penkalla A, PA.09                 Schjørring O, 2.06                 PD.04, PE.06
    ´
Martın B, PA.02                   Perego L, PA.06                   Schmerbach K, 3.02               Usta MF, PC.13, PD.08
    ´
Martınez L, PA.02                 Pessina AC, 04.05, PG.04          Schmidli J, 3.01                 Uydes-Dogan BS, PC.12
Marx N, PE.06                     Peters T, 3.01                    Schoenfelder J, W3.03
Matchkov VV, 5.03                 Peterzan MA, PG.01                    ¨
                                                                    Schone C, PA.09
                                                                                                     Van de Voorde J, PC.07
Mathieson PW, W1.04               Petrescu G, PC.10                 Schrader F, PB.10
                                                                                                     van der Donk C, 05.05, PH.01
Matrozova J, PH.08                Peyrard S, PB.04                  Schubert C, PA.09
                                                                                                     van der Giet M, 2.01
Mayet J, 04.02, PH.02             Piecha G, 01.01, PA.05            Scott M, 1.03
                                                                                                     van Dorsselaer A, PC.03
McKenzie EA, W1.04                Pietrucci F, PF.05                Seccia TM, 04.05, PF.04
                                                                                                     van Engelshoven JMA, 4.06
Meijer Z, 05.05, PH.01            Pimpini L, PE.05                  Shore A, 04.02, PH.02
                                                                                                     Vannarelli A, PF.05
Meoli L, PD.05, PD.06             Pinaud F, PC.08                   Sigurdsson ST, PB.05
                                                                                                     VanTeeffelen JWGE, W1.03
Meyer E, 2.04                     Pinet F, PA.10                    Silva FRO, PE.08
                                                                                                     Varo N, PE.07
Mikelsone I, PG.09                Pinterova M, PC.11, PD.03         Simko F, 2.02
                                                                                                     Vase H, 4.03
Minardi D, PE.04, PF.05           Pinto-Sietsma SJ, 05.05, PH.01    Simonsen U, 2.06
                                                                                                     Vausort M, 1.02
Mohaupt M, 3.01                   Pipy B, 2.05                      Singh A, W1.04
                                                                                                     Velot E, 01.02, PD.02
Mokni W, PA.07                    Piquard J, PD.07                  Sips P, PC.07
                                                                                                     Verde I, PC.06
Mongiardi C, PA.04                Pirags V, PG.09                   Slatineanu SM, PC.10
                                                                                                     Vessieres E, 02.03, PB.06
Monteiro AM, PE.08                Plouin PF, PH.08                  Smith P, PH.09
                                                                                                     Vetter R, 1.04
Monza M, PA.04                    Popova E, 5.01                    Sommerfeld M, 02.07, 03.04,
                                                                                                     Vieira Jr ND, PE.08
Morizzo C, PG.05, PG.06           Potter J, 1.05                       PB.10
                                                                                                     Villringer A, 3.02
Mostard GJM, 4.06                 Prelle K, PA.09                   Somoza B, PE.01, PE.02
                                                                                                     Vinereanu D, PG.06
Mucci L, PE.04                    Prendagast B, PH.05               Soukaseum C, PF.07
                                                                                                     Vink H, W1.03
Mulder P, PA.10                   Puzserova A, PC.14                Southcott E, PH.09
                                                                                                     Vogel J, 01.03, 02.04
Mulders T, 05.05, PH.01                                             Souza DS, PG.03
                                                                                                     Volinkina VM, PH.06, PH.07
Muller R, 2.04                    Quarto C, PF.01                   Sprang C, PE.06
                                                                                                     von Bauer R, PA.03
 ¨
Muller S, 3.02                                                      Sprang CH, PD.04
Muscelli E, PG.05                 Rana-Poussine V, 2.05             Srikusalanukul W, PH.09
Mustafina OE, PF.09               Rappelli A, PE.04, PE.05, PF.05   Stampanoni M, 2.04               Wagner DR, 01.02, PD.02
Muzzonigro G, PE.04, PF.05        Rastaldi MP, PF.06                Stankevicius E, 2.06             Warner DP, PH.04
Myasoedova EE, PG.02              Raunsø JR, PC.09                  Starklint J, PG.08               Weiß W, 1.04
                                  Raynor S, 04.02, PH.02            Steckelings UM, 03.02, 03.04,    Wijetunge S, PC.01
Nabzdyk C, PD.05                  Regitz-Zagrosek V, PA.09             PB.03                         Williams CJ, PF.03
Namsolleck P, 3.02                Reihmane D, PG.09                 Stehouwer CDA, 05.05, PH.01      Witt H, W2.03
Nannipieri M, PG.05               Renzulli A, PF.01                 Steichen O, PH.08                Witzke O, PG.07
Narkiewicz K, PF.03               Restituto P, PE.07                Stella A, PA.06                  Wright A, 04.02, PH.02
Nasibullin TR, PF.09              Retaileau K, PB.06                Stelte-Ludwig B, PB.07           Wuerzner G, PB.04
Natal C, PE.07                    Reznik EV, PH.06, PH.07           Stifts A, PG.09
Neal CR, W1.04                    Richard V, PA.10                  Storozhakov GI, PH.06, PH.07     Zakirova AN, PF.09
Neumann C, 3.02                   Ritz E, 1.01                      Strandgaard S, PB.05             Zanchetta M, PG.04
Nguyen Dinh Cat A, PF.07          Rompe F, PB.03                    Strub JM, PC.03                  Zazzu V, PD.05, PD.06
Nicaud V, 04.01, W3.03            Roques BP, 03.03, PB.02           Stucchi P, PB.08                 Zerbini G, PA.06
Nicolini E, PA.04                 Rossi F, PF.01                    Sujenthiran A, PC.01             Zhang A, PF.07
Nilsson H, 5.03                   Rossi GP, 04.05, PF.03, PF.04,                                     Zicha J, 02.02, PC.05, PC.11,
         ¨
Nilsson-Ohman J, PD.01              PG.04                           Takir S, PC.12                     PD.03
Nimmegeers S, PC.07               Rozenfeld R, PB.02                Telgmann R, 04.01, W3.03         Zidek W, 02.01, PC.02, PH.03

				
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