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IMMUNOHISTOCHEMICAL STUDY OF ESTROGEN AND

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IMMUNOHISTOCHEMICAL STUDY OF ESTROGEN AND Powered By Docstoc
					IMMUNOHISTOCHEMICAL STUDY OF ESTROGEN
AND PROGESTERONE RECEPTOR EXPRESSION IN
              MELASMA


               PROTOCOL OF THESIS

     SUBMITTED FOR THE PARTIAL FULFILLMENT
             OF MASTER DEGREE IN

  DERMATOLOGY, VENEREOLOGY AND ANDROLOGY
                          BY
       EMAN MOHAMMED AHMED ALIAN
                     (M.B, B. CH.)
      FACULTY OF MEDICINE – AIN SHAMS UNIVERSITY

                   SUPERVISED BY
         DR. MAHIRA HAMDY EL-SAYED
PROFESSOR OF DERMATOLOGY, VENEREOLOGY AND ANDROLOGY
      FACULTY OF MEDICINE – AIN SHAMS UNIVERSITY



           DR. AZZA ESMAT MOSTAFA
LECTURER OF DERMATOLOGY, VENEREOLOGY AND ANDROLOGY
      FACULTY OF MEDICINE – AIN SHAMS UNIVERSITY



              FACULTY OF MEDICINE
              AIN SHAMS UNIVERSITY
                      2010
                              INTRODUCTION
       Melasma is a common acquired symmetric hypermelanosis characterized by
irregular light-brown to gray-brown macules and patches involving sun-exposed
areas most commonly the face (1). Although the majority of melasma cases occur
in women, 10% of cases have been reported in males (2).

      Clinically, melasma usually occurs in one of three patterns. The most
common is the centrofacial pattern with involvement of the lateral forehead,
cheeks, nose, upper lip and chin. The less common patterns include the malar and
the mandibular types. Lesions may occasionally occur in other sun-exposed areas
including the forearms and the mid upper chest (3).

     Although histologically, melasma was formerly classified as epidermal,
dermal and mixed type by the locations of pigments. Kang et al., 2002 reported
that there is no true dermal type of melasma (4). In addition, it has been
demonstrated that the melanocytes within affected skin are larger, intensely stained
with prominent dendrites and contain more melanosomes than melanocytes of
unaffected skin, suggesting that these cells may be hyperfunctional in melasma (5).

       The exact etiopathogenesis of melasma is unknown, although multiple
factors have been implicated. These include genetic predisposition; medications,
such as anticonvulsants; ultraviolet light exposure (6); and, most notably, hormonal
influences (7).

       Many cases appear to be related to excess estrogen, either produced
endogenously during pregnancy or delivered exogenously through the use of oral
contraceptive pills and hormone replacement therapy. In vitro studies have shown
that estradiol increases levels of tyrosinase, tyrosinase-related-protein 1 and
tyrosinase-related-protein 2, enzymes involved in human eumelanogenesis, within
normal human melanocytes (8).

      On the other hand, the results of studies on the effects of progesterone on
skin pigmentation have been inconsistent. It has been reported that progesterone
increased cell numbers and tyrosinase activity in melanocytes. The observation that
postmenopausal woman who are given progesterone develop melasma, while those
who are given only estrogen do not, implicates progesterone as playing a critical
role in the development of melasma (9). In contrast, significant inhibitory effect of
progesterone on the proliferation of melanocytes in cell culture was also reported
(10).

     However, estrogen receptor and progesterone receptor expression in
melasma affected skin has not been investigated to date, except for two reports in
the literature (2, 11). Such information is a prerequisite for the understanding the
pathogenesis of melasma in relation to female sex hormones such as estrogen and
progesterone, perhaps then a treatment protocol can be constructed probably
implementing topical anti-estrogen therapy.




                            AIM OF THE WORK
       The aim of this thesis is to investigate the immunohistochemical differences
of the estrogen receptor and progesterone receptor expression between melasma-
affected skin and adjacent-unaffected control skin.
                         SUBJECTS AND METHODS
This thesis is designed to be prospective case-control study. It will be carried on
thirty(30) patients with melasma presenting to the Dermatology outpatient clinic
of Ain Shams University Hospital, Faculty of Medicine, Ain Shams University.

Exclusion criteria:

Patients with other diseases leading to hyperpigmentation will be excluded.

Each patient will be subjected to:

   1-      History taking: including full information about the medical history of
           the patients and history of the disease.
   2-      Full dermatologic examination.
   3-      Skin biopsy:
          i. A 3mm punch biopsy will be taken from an area of melasma affected
               skin.
         ii.   Another 3mm punch biopsy will be taken from unaffected adjacent
               skin as a control.
        iii.   The tissues will be fixed in 10% buffered formalin and embedded in
               paraffin.
        iv.    From the paraffin block, 5 micron thick serial sections will be cut and
               prepared for routine hematoxylin and eosin (H&E) stain for
               histopathological examination by light microscopy.
         v.    Sectioned will be examined for:
                  a. The extend and the distribution of melanin pigment.
                   b. The histologic type of melasma.
                   c. Other relevant histologic findings will be reported.
        vi.    Immunohistochemical staining will be performed using the following
               stains:
                a) Staining for estrogen receptors (ER–β) in lesional and
                   perilesional skin.
                b) Staining for progesterone receptors (PR) in lesional and
                   perilesional skin.
      vii.   All slides were evaluated by two observers in a blinded fashion.



Statistical methodology

Analysis of the data will be done using SPSS (Statistical Program for Social
Science) computer program.



THE THESIS WILL INCLUDE:
     Introduction and aim of the work.
     Review of literature.
     Subjects and methods.
     Results.
     Discussion.
     Conclusion
     Recommendation
     Summary
     References
     Arabic summary
                               REFERENCES

(1) Sanchez NP, Pathak MA, Sato S, Fitzpatrick TB, Sanchez JL and Mihm
    MC Jr (1981): Melasma: a clinical, light microscopic, ultrastructural, and
    immunofluorescence study. J Am Acad Dermatol; 4: 698-710.
(2) Lieberman R and Moy L (2008): Estrogen receptor expression in melasma:
    results from facial skin of affected patients. J Drugs Dermatol; 7(5): 463-465.
(3) Johnston GA, Sviland L and Mc Lelland J (1998): Melasma of the arms
    associated with hormone replacement therapy. Br J Dermatol; 139:932.
(4) Kang WH, Yoon KH, Lee ES, Kim J, Lee KB and Yim H (2002):
    Melasma: histopathological characteristics in 56 Korean patients.         Br J
    Dermatol; 146: 228-237.
(5) Grimes PE, Yamada N and Bhawan J (2005). Light microscopic,
    immunohistochemical, and ultrastructural alterations in patients with
    melasma. Am J Dermatopathol; 27: 96-101.
(6) Ortonne JP, Arellano I, Berneburg M, Cestari T, Chan H, Grimes P,
    Hexsel D, Im S, Lim J, Lui H, Pandya A, Picardo M, Rendon M, Taylor
    S, Van Der Veen JPW and esterhof WW (2009): Global survey of the
    role of ultraviolet radiation and hormonal influences in the development
    of melasma. J Europ Acad Dermatol Venereol; 23, 1254-1262.
(7) Hassan I, Kaur I, Sialy R and Dash RJ (1998): Hormonal milieu in the
    maintenance of melasma in fertile women. J Dermatol; 25: 510-512.
(8) Kippenherger S, Loitsch S, Solano F, Bernd A and Kaufmann R. (1998):
    Quantification of tyrosinase, TRP-1, and TRP-2 transcripts in human
    melanocytes by reverse transcriptase-competitive multiplex PCR-regulation
    by steroid hormones. J Invest Dermatol; 110: 364-367.
(9) Bolanca I, Bolanca Z, Kuna K, Vuković A, Tuckar N, Herman R and
    Grubisić G (2008): Chloasma: the mask of pregnancy. Coll Antropol; 32
    (S2): 139-141.
(10) Wiedemann C, Nägele U, Schramm G and Berking C (2009): Inhibitory
    effects of progestogens on the estrogen stimulation of melanocytes in vitro.
    Contraception; 80: 292-298.
(11) Jang, YH, Lee YJ, Kang HY, Lee E-S and Kim YC (2010): Oestrogen and
    progesterone receptor expression in melasma: an immunohistochemical
    analysis. J Europ Acad Dermatol Venereol; 24: 1312-1316.

				
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