Vet Pathol-1977-Hargis-218-28 by xiaopangnv


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Chronic Dermatosis and Cutaneous Squamous Cell Carcinoma in the Beagle Dog
                        A. M. Hargis, R. W. Thomassen and R. D. Phemister
                                      Vet Pathol 1977 14: 218
                               DOI: 10.1177/030098587701400304

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 Vet. Pathol. 14: 218-228 (1977)

 Chronic Dermatosis and Cutaneous Squamous Cell Carcinoma in the
                           Beagle Dog

                  A . M . HARGIS,R. W . THOMASSEN R. D. PHEMISTER

    Collaborative Radiological Health Laboratory and Department of Pathology, Colorado State
                                  University, Fort Collins, Colo.

   Abstract. Chronic dermatosis developed in sparsely-haired, lightly pigmented, ventral body skin in
397 of 1 680 beagle dogs by the time the dogs were 5 ’ 1 2 years old. Twenty-six of 55 other beagles had
developed such lesions by 12 years of age. Squamous cell carcinomas developed in the sites of
dermatosis in eight of the 397 younger and five of the 26 older beagles. The lesions resembled solar
keratosis (solar dermatosis, actinic dermatosis or senile keratosis) in man. They developed under cir-
cumstances suggesting that solar radiation is involved in the pathogenesis, as it is in man.

  Chronic dermatosis and keratosis occur in human skin inadequately protected
from prolonged exposure to sunlight. T h e changes progress to invasive carcinoma
often enough to be considered precancerous [ 7 ] . Chronic inflammatory and prolif-
erative lesions also occur in the skin of dogs, but with the exception of nasal solar
dermatitis, neither location nor circumstances of development have implicated
chronic solar radiation as a cause, and carcinoma is not a sequel [13]. This report
deals with the development of inflammatory and proliferative lesions in nonpig-
mented and sparsely-haired abdominal, inguinal, preputial, and scrota1 skin of
beagles. T h e circumstances suggest that solar radiation is involved in the pathologic
processes. Further, invasive squamous cell carcinomas have developed frequently
enough in the abdominal, inguinal, and preputial sites of chronic dermatosis to
indicate that the preexisting changes are precancerous a n d that this change may be
considered a canine counterpart of human actinic dermatosis and subsequent

                                      Materials and Methods
   The clinic and pathology records of 858 male and 882 female purebred beagles were reviewed for
data o n inflammatory, proliferative, or neoplastic lesions in nonpigmented skin. The beagles were part
of a large colony in a research laboratory in rural northern Colorado. Most of these dogs will live out
their lives in the colony and at the time of the study were from 3 to 12 years old. One thousand three
hundred and twenty designated “long-term” beagles were exposed to 20 or 100 R (51.6 x 10F or 258 x
10V4Cikg) whole body, “’Co gamma radiation. Three hundred and sixty “long-term” control dogs were
sham irradiated. Fifty-five nonirradiated beagles, the F, breeding group, were the oldest survivors of
original breeding stock in the colony. Five other beagles, designated “short-term”, were included
because cutaneous squamous cell carcinomas developed (table I ) . All beagles were maintained in

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                                                                   Table I. Clinic, pathology and irradiation data from beagles with cutaneous squamous cell carcinomas
                                                                                                                          Age derma-     Age carci-
                                                                                               Age at irra- Irradiation                                Age subsequent     Primary cuta-
                                                                                                                           tosis first   noma first
                                                               Dog         Group         Sex    diation,        level                                 carcinomas noted.   neous location      Condition of dogs'
                                                                                                                             noted,        noted,
                                                                                                  days     (""Co gamma)                                     years         of carcinomas
                                                                                                                             vears         years
                                                                1        F, breeding     F                      0             7.2           7.2       -                    Inguinal        Dead, no metastasis
                                                                2        F, breeding     M                      0             9.4          10.9       11.5                 Prepuce         Alive, with dermatosis
                                                                3        F, breeding     M                      0             4.1          10.2       -                    Abdomen         Alive, with dermatosis       8.
                                                                4        F, breeding     F                      0             6.9          10.9       11.3, 11.5, 11.8     Inguinal        Dead, no metastasis          D
                                                                5        F, breeding     F                      0             2.0           6.6       7.4, 7.8             Abdomen         Dead, equivocal metastasis   3
                                                                6        Long-term       F                   100 R            4.0           7.2       -                    Abdomen         Alive, with dermatosis       rA
                                                                7        Long-term       F                   100 R            3 .O          7.1       -                    Abdomen         Alive, with dermatosis       E
                                                                8        Long-term       M           28'      00 R             .8           4.2       -                    Prepuce         Alive, with dermatosis       3
                                                                9        Long-term       M            8l      20 R              .4          3.2       -                    Prepuce         Alive, with dermatosis       C
                                                               10        Long-term       F            -        0              6.3           7.4       -                    Inguinal        Alive, with dermatosis
                                                               11        Long-term       F          365       00 R            1.6           6.4       6.6, 6 . 9           Inguinal        Dead, no metastasis          I

                                                               12        Long-term       M          365       00 R            3 .O          7.2       -                    Inguinal        Alive, with dermatosis
                                                               13        Long-term       F          365       00 R            2.0           7.1       7.2                  Abdomen         Alive, with dermatosis
                                                               14        Short-term      F                     0              4.6           5.3       5 . 9 , 6.3          Abdomen         Dead, no metastasis

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                                                               15        Short-term      F         2         380 R            4.6           4.6       5.0, 5.2, 5.4        Abdomen         Alive, with dermatosis
                                                               16        Short-term      F      2152         220 R            1.8           6.0       -                    Abdomen         Dead, no metastasis
                                                               17        Short-term      F            -        0              4.1           5.0       -                    Abdomen         Dead, no metastasis
                                                               18        Short-term      F            -        0              2.9           4.1       4.4. 4.8             Inguinal        Dead, metastasis to lung
                                                               1   in utero (days after breeding)
                                                               * as of   April 1976
220                               Hargis, Thomassen and Phemister

outside kennels equipped with a house, shade panel, and self waterers and feeders. Kennel floors were
covered with small, light colored, river rock. In addition to routine veterinary medical, surgical, and
dental care, all beagles were observed daily, were given annual physical examinations and were
immunized against canine distemper, infectious canine hepatitis and leptospirosis. All skin tumors were
excised and submitted for histologic examination. After tumors were removed, the dogs were regularly
examined for recurrence of neoplasia, evidence of metastasis, and occurrence of other tumors. All dogs
that died or were killed were given a thorough postmortem examination. Surgical and postmortem
specimens were fixed by immersion in 10 percent buffered neutral formalin. Tissue sections were em-
bedded in paraffin, cut at 6 micrometers, and stained with hematoxylin and eosin (HE). Selected sec-
tions of skin were stained with periodic acid-Schiff (PAS) with and without diastase, colloidal iron
with and without hyaluronidase, Verhoeff‘s elastic tissue stain, Pinkus’ acid orcein-Giemsa method,
and Snook’s method for reticulum.

   About one fourth of the “long-term” beagles and o n e half of the F, breeding
group had scattered areas of dermatosis of nonpigmented, sparsely-haired skin in
abdominal, inguinal, and scrota1 regions (table 11). Erythema, associated with
edema and scaling and considered to be mild sunburn, was often the earliest noted
change. Exacerbations of erythema occurred in many dogs, especially during sum-
mer when sunlight was intense. T h e erythema subsided during cloudy weather o r
indoor confinement. Scattered areas of cutaneous thickening, induration, and
roughening, often with tan to brown crusts and many comedones (fig. 1, 2 ) , suc-
ceeded and existed with the erythema. The transition from adjacent normal, thin,
pliable, pigmented skin to the lesions in nonpigmented skin was usually abrupt
(fig. 1). After an average of 3.2 years, eight of 397 ‘‘long-term’’ dogs and five of
26 F, breeding dogs developed single or multiple, ovoid, red to tan neoplastic
plaques in sites of preexisting dermatosis (fig. 3 ) . Tumors became nodular o r fungoid
in a few dogs.
   Microscopic examination of moderately indurated skin showed the epidermis
was thickened by hyperkeratosis, parakeratosis, acanthosis, and often by mild
pseudoepitheliomatous hyperplasia (fig. 4). In more extensively involved skin,
epidermal cells in the stratum spinosum often were vacuolated and the superficial
epidermis of hair follicles was acanthotic. T h e dermis also was thickened, initially
by deposition of abundant, finely fibrillar t o granular, lightly eosinophilic collagen
within the papillary layer. With time both the papillary and reticular layers were
greatly thickened with collagen that eventually became dense and poorly vascular-
ized (fig. 5). Few t o many lymphocytes and plasma cells, a n d infrequently neutro-
phils, were most often in the papillary layer where they surrounded telangiectatic
capillaries. In areas of nodules and plaques, the superficial epidermis was usually
ulcerated, and acanthosis resulted in deeply penetrating cords and islands of
squamous epithelium (fig. 6). T h e change was considered neoplastic when the
basement membrane was focally to extensively disrupted and individual epidermal
cells occurred in the dermis (fig. 7). The carcinomas and some areas of acanthosis
and pseudoepitheliomatous hyperplasia contained few to many mitotic figures which
were often atypical, multinucleate and uninucleate giant cells with acidophilic
nucleoli and individual cell keratinization (fig. 8, 9). Deep intradermal fibroplasia

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                           Dermatosis and Squamous Cell Carcinoma                                                    22 I

Table 11. Incidence and mean age of onset of dermatosis and cutaneous squamous cell carcinoma in
                               long-term and F, breeding beagles
                                                   Long-term beagles                         F, breeding beagles

                                                 Male                 Female                  Male        Female
                                              ( n = 840)            ( n = 840)             ( n = 18)     ( n = 37)
Current mean age of dogs, years                5.4                   5.4                   12.2           12.2
Number with dermatosis                       214                   183                     10             16
Mean age of dermatosis onset, years            1.9                   2.2                    5.8            4.2
Number with cutaneous squamous                 3                     5                      2              3
 cell carcinoma
Mean age of carcinoma onset, years              4.9                    7 .O                10.6            8.3
Number with derrnatosis that devel-             3 of 214               5 of 183             2 of 10        3 of 16
 oped carcinomas

 was prominent in association with penetrating cords of epidermal cells and existed
with many lymphocytes, plasma cells, histiocytes, and mast cells. Foci of lympho-
cytes and plasma cells partially surrounded dilated hair follicles and sweat glands
subjacent to the epidermal thickening. Many neutrophils and histiocytes bordered
by lymphocytes, plasma cells, and fibroblasts were seen where adnexal walls had
been disrupted.
   To compare the lesions in man and dog, histochemical procedures used to
characterize solar damaged human skin were done on sections of dermatosis and
squamous cell carcinoma in the beagle. Moderate to large amounts of PAS-
positive, diastase labile material interpreted to be glycogen occurred in the epider-
mis within vacuolated, squamous cells of the spinous layer. Glycogen also was in
the central portions of invading epidermal columns and cords. Colloidal iron
positive, hyaluronidase labile material interpreted to be hyaluronic acid was within
the interstices between many squamous cells in areas of acanthosis. N o colloidal iron
positive, hyaluronidase resistant material was found intracellularly . Elastic tissue
stains in solar-affected beagle skin showed few positive fibers in the papillary layer
and focal tangled masses of positive fibers deep within the reticular layer of the
dermis (fig. 5 ) . These fibers were indistinguishable from collagen when H E
stained. Generally the wide zone of dense collagen within the beagle dermis was
free of positive staining elastic fibers. Snook’s reticulum stain showed proliferation
of reticular fibers in zones of inflammation. Melanin granules were seen in some
squamous cells, in melanocytes, and free and within melanophages in the upper
dermis, although no abnormal pigmentary accumulations were seen clinically.
   No carcinomatous vascular invasion was noted in biopsy specimens from the
beagles; however, postmortem examination showed the lungs of one dog contained
metastatic foci of well-differentiated, squamous cell carcinoma (fig. 10). In another
dog a well-differentiated squamous cell carcinoma was in the external iliac lymph
node; other surgical specimens, however, had squamous cell carcinomas, possibly
originating in the mammary gland duct, in addition to those arising in nonpigmented
  Table I1 shows that within groups the percent of males and females affected is

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222                             Hargis, Thomassen and Phemister

   Fig. 1: Sparsely-haired abdominal skin from female beagle. Abrupt transition between thin and
smooth pigmented skin, and thick and rough nonpigmented skin. Minute black foci in nonpigmented
areas are comedones.
   Fig. 2: Crust formation (hyperkeratosis) in sparsely-haired, nonpigmented areas of abdominal skin
from female beagle.

nearly equal and that the mean age of onset of dermatosis or neoplasia in the F,
breeding group is greater than that of the long-term group. F, dogs were not given
physical examinations as frequently as were long-term beagles, therefore, we may
not have seen dermatosis as early. Also F, animals are older and have had more
time to develop dermatosis and neoplasia. Data indicate no strong tendency for
dermatosis or neoplasia to be associated with previous gamma irradiation.

   Lesions associated with solar damage in beagle skin are morphologically and
histochemically identical to those seen in human skin. There are two exceptions.
First, in man, appropriate stains in solar damaged skin demonstrate abnormal
elastic tissue in the areas of wavy, basophilic, fragmented fibers seen with HE [7,
91. Squamous cell carcinomas, however, have been induced by ultraviolet radiation
with little dermal connective tissue damage in hairless mice, whereas dermis of
haired mice was damaged extensively [4, 211. Solar elastosis is not easily produced
in animals, but it has been induced in haired rat skin without squamous cell

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                           Dermatosis and Squamous Cell Carcinoma                             223

   Fig. 3: Ulcerated surface of squamous cell carcinoma within sparsely-haired, nonpigmented, ab-
dominal skin.
   Fig. 4: Epidermal thickening of nonpigmented abdominal skin. Hyperkeratosis and acanthosis with
downward growth of the epidermis (pseudoepitheliomatous hyperplasia). Parakeratosis and thickening
of the granular cell layer. H E .

carcinoma formation [ 141. Several investigators have concluded that solar elastosis
is often a concomitant finding rather than necessary precursor in the formation of
cutaneous squamous cell carcinoma [ 3 , 211. Secondly, in contrast to m a n , abnor-
mal pigmentary accumulations were not found in solar exposed dog skin, nor have
they been mentioned in rats, cattle, cats o r goats. A few blue-black plaques,
however, have been produced in brown, hairless mouse skin subsequent to ultravi-
olet exposure [4]. Species variation seems great and may account for the different
reaction of solar exposed human a n d beagle dermis.
   It is perhaps too early to draw many conclusions about the biological behavior of
these solar induced carcinomas in beagle skin, but the metastasis to the lung in one
dog indicates malignant potential. Solar induced squamous cell carcinomas on the
noses of dogs and ears of cats rarely metastasize 1131. Similarly, solar induced
neoplasms in man are less malignant than are their de nova counterparts [7]. With
only o n e unequivocal metastasis in 18 squamous cell carcinomas in o u r series, this
may also be the case in ventral body skin of dogs.
   In addition to solar keratosis, several cutaneous and mucocutaneous lesions with
a propensity to develop into invasive carcinoma occur in man. These lesions include

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224                               Hargis, Thornassen and Phemister

   Fig. 5 : Marked thickening of nonpigrnented abdominal skin. Acanthosis and extensive fibrosis of
the dermis. Darkly stained elastic fibers subjacent to the zone of dense collagen. Pinkus’ acid orcein-
   Fig. 6: Deeply penetrating broad cords and islands of epidermal cells within an ulcerated squamous
cell carcinoma. Nonpigmented abdominal skin. HE.

Bowen’s disease, erythroplasia of Queyrat, and extramammary and mammary
Paget’s disease. They resemble solar keratosis and should be differentiated for
treatment and prognosis. Canine counterparts of these lesions have not been
described. Overall, the changes in solar exposed beagle skin most closely resemble
solar keratosis in man which is differentiated from the other precancerous lesions
by anatomic location, morphology, and histochemistry [7].
   The mechanism by which sunlight induces precancerous dermatosis and cuta-
neous neoplasia has been debated. Studies have shown that ultraviolet radiation is
absorbed in exposed epidermis and dermis, more readily by some parts than by
others. Absorption is related to molecular structure, and occurs primarily in conju-
gated systems that contain alternating single and double bonds in a carbon chain
[8]. By far the most susceptible are the pyrimidine bases of the DNA molecule, and
the major change is formation of covalent bonds between those bases yielding
dimers [ 5 , 8 , 181. Other types of damage include DNA-protein cross-links, damage
to bases other than pyrimidines, and single-strand breaks [8]. Much of the damage,
however, is at least partially repairable by enzyme systems in man [20]. Other

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                          Dermatosis and Squamous Cell Carcinoma                           225

  Fig. 7: Narrow epidermal cords and isolated epidermal cells within squamous cell carcinoma.
Nonpigmented abdominal skin. HE.
  Fig. 8: Cellular pleomorphism within an island of epidermal cells in squamous cell carcinoma.
Nonpigmented abdominal skin. HE.

experiments have shown that a carcinogen, cholesterol-a-oxide, has been formed
repeatedly from natural sterols in the skin of albino hairless mice and man as a
result of ultraviolet radiation 111. Enzyme systems for the detoxification of the
carcinogen also have been found [ 11, 21. Because much ultraviolet-induced dam-
age is repairable in normal people, it has been proposed that lack of DNA repair is
not responsible for most neoplastic development but rather it is faulty DNA repair
or replication which leads to mutation, integration with oncogenic viruses, or
activation of oncogens [ 5 ] . It is also thought that cells actively synthesizing DNA,
which occurs in repair, are more susceptible to further ultraviolet exposure [17].
   Regardless of the mechanism, neoplastic development is influenced by many
factors that regulate the quantity of ultraviolet radiation reaching the earth’s
surface. Such factors include the quantity of ozone, which absorbs wavelengths
below 290 nanometers [19]; the quantity of gaseous molecules of the atmosphere
and fine suspended particles that scatter ultraviolet rays; and the quantity of smog,
which may involve absorption and scattering. These factors vary greatly with
altitude, latitude, season of the year, and time of day [ l o , 121 and are partially

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226                              Hargis, Thomassen and Phemister

   Fig. 9: Keratinization within individual epidermal cells (arrows) in an area of pseudoepithelioma-
tous hyperplasia. Nonpigmented abdominal skin. HE.
   Fig. 10: Metastatic focus of squamous cell carcinoma originating in nonpigmented abdominal skin.
Lung. HE.

responsible for regional variation in human and animal cutaneous tumor incidence.
Another important factor that increases exposure to ultraviolet radiation is back-
ground scatter that occurs at beaches mainly by reflection from sand, and not from
water [19]. Increased temperature, humidity, wind velocity, and skin hydration

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                             Dermatosis and Squamous Cell Carcinoma                                  221

also have been shown to enhance squamous cell carcinoma development in ultra-
violet radiated mice [6, 15, 161.
   Environmental and behavioral factors may play a role in chronic dermatosis and
cutaneous squamous cell carcinoma in the beagle, just as it does in man. Our
colony of beagles is at a moderately high altitude in a smog-free area with abundant
sunshine, and the dogs are housed outside on reflective gravel. Although shade
panels and houses are provided, most dogs spend the daylight hours in full
sunshine; they frequently rest on their backs with their ventral surface exposed to
the s u n . All of these factors probably cause maximum exposure to sunlight that
directly or indirectly contacts unprotected, lightly-pigmented, sparsely-haired skin.
Ultraviolet exposure to such areas in the ventral body skin of many beagles
therefore probably is sufficient to produce dermatosis, keratosis, and invasive and
metastatic squamous cell carcinoma.

   Supported in part by United States Public Health Service Contracts F D A 72-302 and 223-76-6002
from the Bureau of Radiological Health, Food and Drug Administration.

  1 Black, H.S.; Lo, W.B.: Formation of a carcinogen in human skin irradiated with ultraviolet light.
    Nature. 234:306-307, 1971
 2 Chan, J.T.; Black, H.S.: Skin carcinogenesis: cholesterol-5 a, 6 a-epoxide hydrase activity in
    mouse skin irradiated with ultraviolet light. Science. 186: I2 16- 12 17, I974
 3 Epstein, J.H.: Ultraviolet light carcinogenesis, in Advances in biology of skin, e d . Montagna; 1st
    ed., pp. 215-234, Pergamon Press, Oxford, 1966
 4 Epstein, J . H . ; Epstein, W.L.: A study of tumor types produced by ultraviolet light in hairless and
     hairy mice. J Invest Dermatol 41:463-473, 1963
 5 Epstein, W.L.; Fukuyama, K . ; Epstein, J.H.: Ultraviolet light, DNA repair and skin carcinogenesis
    in man. Fed Proc 30:1766-1771, 1971
 6 Freeman, R.G.; Knox, J.M.: Influence of temperature on ultraviolet injury. Arch Dermatol
    89:858-864, 1964
 7 Graham, J . H . ; Helwig, E.B.: Premalignant cutaneous and mucocutaneous diseases, in Dermal
    pathology, ed. Graham, Johnson and Helwig; 1st ed., pp. 561-624, Harper and Row, Hagerstown,
 8 Jagger, J.: Ultraviolet effects, in Medical radiation biology. ed. Dalrymple, Gaulden, Kollmorgen
    and Vogel; 1st e d . , pp. 44-51, W. B . Saunders, Philadelphia, 1973
 9 Johnson, W.C.: Histochemistry of the skin, in Dermal pathology, e d . Graham, Johnson and
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10 Koller, L.: The physics of the atmosphere, in The biological effects of ultraviolet radiation, e d .
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11 Lo, W.B.; Black, H.S.; Knox, J.M.: Cholesterol a-oxide hydrase activity in hairless mouse skin.
    CIin Res 22:618A, 1974
12 London, J.: The depletion of ultraviolet radiation by atmospheric ozone, in The biological effects of
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13 Muller, G.H.: Basal cell epithelioma and squamous cell carcinoma in animals. Arch Dermatol
14 Nakamura, K.; Johnson, W.C.: Ultraviolet light induced connective tissue changes in rat skin: A
    histopathologic and histochemical study. J Invest Dermatol 51:253-258, 1968
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    Invest Dermatol 64:250-252, 1975

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228                               Hargis, Thomassen and Phemister

16 Owens, D . W . ; Knox, J.M.; Hudson, H.T.; Troll, D.: Influence of wind on ultraviolet injury. Arch
   Dermatol 109:ZOO-201, 1974
17 Pound, A . W . : Induced cell proliferation and the initiation of skin tumor formation in mice by
   ultraviolet light. Pathology 2269-275, 1970
18 Swanbeck, G.: Aetiological factors in squamous cell skin cancer. Br J Dermatol 85:394-396, 1971
19 Urbach, F.: Geographic distribution of skin cancer. J Surg Oncol 3:219-234, 1971
20 Wilkins, R.J.: DNA repair: a molecular process of medical relevance. N Z Med J 80:210-215, 1974
21 Winkelmann, R.K.; Zollman, P.E.; Baldes, E.J.: Squamous cell carcinoma produced by ultraviolet
   light in hairless mice. J Invest Dermatol 40:217-224, 1963

Request reprints from Ann M . Hargis, DVM, Collaborative Radiological Health Laboratory, Foothills
Campus, Colorado State University, Fort Collins, C O 80523 (USA).

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