Photocarcinogenesis and persistent hyperplasia in UV-irradiated SENCAR mouse skin.

Susceptibility to photocarcinogenesis has been examined in several mouse strains and stocks including SENCAR, CD-1, BALB/c, C3H, C57Bl, and NZB, SENCAR mice are hypersusceptible to tumorigenesis caused by single high dose exposures to ultraviolet (UV) radiation but not by chronic low-dose exposures. SENCAR mice also exhibit an exaggerated and persistent epidermal hyperplasia in response to UV-induced tissue damage. The persistent hyperplasia is apparently due to a sustained proliferation of the epithelial basal cells, rather than to delayed cell differentiation. SENCAR mice did not exhibit persistent hyperplasia following other forms of tissue damage (surgical or thermal). In related studies, the levels of thymine dimers induced in SENCAR epidermis by UV radiation were comparable to those observed in BALB/c epidermis. In addition, no differences were found in the tissue distribution or persistence of thymine dimers in SENCAR and BALB/c skin.


Photocarcinogenesis and Persistent
Hyperplasia in UV-Irradiated SENCAR Mouse Skin by Paul T. Strickland*t Susceptibility to photocarcinogenesis has been examined in several mouse strains and stocks including SENCAR, CD-1, BALB/c, C3H, C57BI, and NZB. SENCAR mice are hypersusceptible to tumorigenesis caused by single high dose exposures to ultraviolet (UV) radiation but not by chronic low-dose exposures. SENCAR mice also exhibit an exaggerated and persistent epidermal hyperplasia in response to UV-induced tissue damage. The persistent hyperplasia is apparently due to a sustained proliferation of the epithelial basal cells, rather than to delayed cell differentiation. SENCAR mice did not exhibit persistent hyperplasia following other forms of tissue damage (surgical or thermal). In related studies, the levels of thymine dimers induced in SENCAR epidermis by UV radiation were comparable to those observed in BALB/c epidermis. In addition, no differences were found in the tissue distribution or persistence of thymine dimers in SENCAR and BALB/c skin.
The development of SENCAR mice, which are highly susceptible to skin carcinogenesis, has provided a useful model for investigating factors that modulate hypersusceptibility as an approach to the study of mechanisms of skin neoplasia (1)(2)(3). SENCAR mice are selectively bred to be hypersusceptible to chemically induced twostage skin carcinogenesis, and this unusual sensitivity has been shown to be an inherent characteristic of the skin of these animals rather than an altered systemic response of the host (4).
We have been investigating certain aspects of photocarcinogenesis regarding susceptibility to tumor induction in mouse skin. A number of experiments have compared strain susceptibility to photocarcinogenesis using single and chronic exposure protocols, and these are summarized in Table 1. These experiments have shown that a single treatment of ultraviolet (UV) radiation (5.8-11.5 x 104 J/m2) from FS 40 sunlamps, without subsequent chemical or physical promotion, results in the dose-dependent induction of epithelial tumors (papillomas and squamous cell carcinomas) in SENCAR mice (5). No tumors of any kind are produced in comparably treated CD-1 or BALB/c mice. This finding demonstrates that the susceptibility determinants selected in SENCAR mice are not specific for chemical carcinogenesis, but can also potentiate photocarcinogenesis. In contrast, chronic exposure to UV radiation (1.5-2.9 x 104 J/m2 three times per week) is not unusually carcinogenic in SENCAR mice compared to BALB/c mice (Table 1). Thus the exhibition of hypersusceptibility depends on how the UV radiation is administered, suggesting that other factors (e.g., tissue damage) may be involved. Three pigmented mouse strains (C3H, C57BV/6, and NZB/N) are somewhat more resistant than the two albino strains to the carcinogenic effects of chronic UV radiation exposure, based on latency period for tumor appearance. It is noteworthy that the tumors induced in SENCAR mice by either exposure protocol (single or chronic) are primarily of epidermal origin and, in this respect, are similar to most human skin tumors.
Much of our effort has focused on investigating the biological basis for the hjpersusceptibility of SENCAR mice to single treatment photocarcinogenesis. One approach we have used to address this question is to examine UV-induced DNA damage in mouse skin to determine whether damage and repair varies among mouse strains. Since several lines of evidence suggest that cyclobutane-pyrimidine dimers produced in DNA by UV radiation may be the initial lesion involved in photomutagenesis and photocarcinogenesis, we have produced monoclonal antibodies specific for thymine dimers for use in immunological assays (8,9). Using these assays, we examined the distribution and repair of thymine dimers in the skin of three different strains of mice (SENCAR, BALB/c, and C3H) exposed to UV radiation. In preliminary studies, an indirect immunofluorescent assay (10) was used to examine the cellular and tissue distributions of thymine dimers in frozen skin 'Percent of tumors in this tissue. eDifference between strains is significant at p = 0.10 (t < 1 hr) and p < 0.01 (t = 24 hr) by t-test. 'Mean ± SD (n = 3); difference between strains not significant by t-test.
"Mean ± SD (n = 3); difference between strains significant (p < 0.025) by t-test. 'Compared to 18-20 wk for BALB/c. sections; no major differences were observed among the three mouse strains (unpublished observation). This finding suggests that the optical properties of the skin that govern penetration and absorption of UV radiation are similar in these mouse strains and probably do not contribute to variations in susceptibility to photocarcinogenesis among the strains. A radioimmunoassay was used to quantitate thymine dimer induction and persistence in DNA extracted from mouse skin at various times after UV irradiation (11). Initial yields of thymine dimers in DNA extracted from ear and dorsal epithelium were similar in the two mouse strains tested, SENCAR and BALB/c ( Table 2). In addition, thymine dimer yields measured 24 hours after UV irradiaiton were similar to initial yields in the epidermis of the two strains. This persistence of thymine dimers in mouse skin is consistent with other studies, indicating that murine skin cells are deficient in excision repair of these photoproducts (12,13). A lower initial yield of dimers was detected in BALB/c dermis than in SENCAR dermis. However, this finding did not correlate with the preponderance of dermal tumors (fibrosarcomas) arising in BALB/c mice compared to SEN-CAR mice, in which the tumors are mainly of epithelial origin. Thus no clear correlation can be drawn between the distribution, induction, or persistence of thymine dimers in the UV-exposed tissues ofthese mouse strains and their varying susceptibilities to photocarcinogenesis. These data suggest that SENCAR mice do not suffer from abnormalities that alter either the levels of DNA photoproducts induced in their epidermis or the rate of removal of thymine dimers. Consistent with this conclusion is the recent report (14) that host cell reactivation of UV-irradiated herpes simplex virus by primary epidermal cells from SENCAR, BALB/c, and CD-1 mice is similar. These data argue against a defective DNA repair mechanism for the observed hypersusceptibility of SENCAR mice to photocarcinogenesis.
It has been postulated (4,15) that untreated SEN-CAR mouse skin contains a population of cells, constitutively initiated for tumorigenesis, that are resistant to terminal differentiation in vitro. In our studies, SEN-CAR mice exhibited an unusual wound healing pattern following single exposure to UV radiation (5,16). These treatments are ulcerogenic, and the gross appearance of the healing wound was unusual in that it was exophytic and very coarse-somewhat reminisicent of hypertrophic scarring in humans. When we examined this abnormality in detail, an exaggerated and persistent epithelial hyperplasia was evident in the exposed skin of SENCAR mice but not BALB/c mice. Epithelial cell proliferation and migration were measured in vivo by autoradiography to determine the cellular basis for the persistence of hyperplasia. Twelve weeks after irradiation (when tissue regeneration is normally completed), the rate of epidermal basal cell proliferation in SEN-CAR mouse skin was approximately four times the rate in BALB/c mouse skin that had been identically treated (Table 3). On the other hand, epidermal cell transit times (time for labeled cells to traverse the epidermis) were similar in the two strains of mice. These results suggest that the abnormal hyperplasia in SENCAR mouse skin is due to a sustained increase in basal cell proliferation rather than to inhibition of cell differentiation.
To determine whether the abnormal hyperplastic response of SENCAR skin is specific for UV-induced tissue damage, two other types of tissue damage, surgical and thermal, were examined. Neither full thickness scalpel incision, nor a thermal treatment, which has been shown (17) to mimic UV-induced ulceration, were effective in producing persistent hyperplasia in SEN-CAR epidermis. Thus persistent hyperplasia is not a general response of SENCAR skin to all forms of tissue damage. In addition, tissue damage caused by surgical incision administered after subcarcinogenic exposures to UV radiation did not enhance the tumorigenic potential of the UV radiation. These results are summarized in Table 4.
The precise relationship between sustained basal cell proliferation and hypersusceptibility to photocarcinogenesis remains to be determined. However, three hypotheses are indicated: (1) that sustained basal cell pro-liferation predisposes SENCAR mice to tumor susceptibility by causing repeated replication of basal cell DNA; (2) that sustained proliferation is an independent manifestation of a genetic defect in SENCAR mice that also mediates susceptibility to skin carcinogenesis; or (3) that sustained proliferation is a premalignant state in which the cells are partially transformed.
Research was sponsored by the National Cancer Institute, DHHS, under contract No. N01-CO-23909 with Litton Bionetics, Inc. The work described in this paper was not funded by EPA and no official endorsement should be inferred. The contents of this publication do not necessarily reflect the views or policies of the Department of Health and Human Services, nor does mention of trade names, commercial products or organizations imply endorsement by the U.S. Government.