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We previously reported that benzo[a]pyrene (BaP) and UVA radiation synergistically induced oxidative DNA damage via 8-hydroxy-2'-deoxyguanosine (8-OHdG) formation in vitro. The present study shows that microsomal BaP metabolites and UVA radiation potently enhance 8-OHdG formation in calf thymus DNA about 3-fold over the parent compound BaP. Utilization of various reactive oxygen species scavengers revealed that singlet oxygen and superoxide radical anion were involved in the 8-OHdG formation induced by microsomal BaP metabolites and UVA. Two specific BaP metabolites, benzo[a]pyrene-r-7,t-8-dihydrodiol-t-9,10-epoxide (+/-) (anti) (BPDE) and BaP-7,8-dione, were further tested for synergism with UVA. BaP-7,8-dione showed an effect on 8-OHdG formation induced by UVA radiation that was similar to that of the parent BaP, whereas BPDE exhibited significantly higher induction of 8-OHdG than BaP. At as low as 0.5 microM, BPDE plus UVA radiation substantially increased 8-OHdG levels about 25-fold over the parent BaP. BPDE increased the formation of 8-OHdG levels in both BPDE concentration- and UVA dose-dependent manners. Additionally, singlet oxygen was found to play a major role in 8-OHdG induction by BPDE and UVA. These results suggest that BaP metabolites such as BPDE synergize with UVA radiation to produce ROS, which in turn induce DNA damage.

Genistein is a soybean isoflavone with diverse biological activities. It is a potent antioxidant, a specific inhibitor of protein tyrosine kinase, and a phytoestrogen. In recent years, increasing evidence has accumulated that this natural ingredient shows preventative and therapeutic effects for breast and prostate cancers, postmenopausal syndrome, osteoporosis, and cardiovascular diseases in animals and humans. In the past decade we have conducted a series of studies and demonstrated that genistein has significant antiphotocarcinogenic and antiphotoaging effects. Genistein substantially inhibits skin carcinogenesis and cutaneous aging induced by ultraviolet (UV) light in mice, and photodamage in humans. The mechanisms of action involve protection of oxidative and photodynamically damaged DNA, downregulation of UVB-activated signal transduction cascades, and antioxidant activities. In this article, we review the biological activities of genistein, as well as published and unpublished research from our laboratory. In addition, we discuss the potential application of genistein to clinical dermatology.

Benzo[a]pyrene (BaP) is an ubiquitous environmental pollutant with potential carcinogenecity. It was shown that BaP, upon irradiation by UV A, enhanced the formation of 8-hydroxy-2'-deoxyguanosine in purified DNA and in cultured cells. The purpose of this present study was to determine whether BaP and UV radiation synergistically generate reactive oxygen species (ROS) that consequently result in the oxidation of DNA bases. In this study, the levels of H(2)O(2) were measured as an indicator of ROS in A431 cells and primary human keratinocytes treated with BaP plus UV radiation. Production of H(2)O(2) significantly increased from cells treated with BaP plus UVB or UVA, with the latter having a much greater effect. The responses of A431 cells and primary human keratinocytes to BaP and UVA irradiation were similar in generation of extracellular H(2)O(2). Also, H(2)O(2) production proportionally correlated with UVA and UVB dose, but was independent of time or BaP concentration. Treatment with catalase and general ROS scavengers significantly decreased H(2)O(2) production from cells treated with BaP plus UVA, whereas scavengers of *O2-, *OH, and (1)O(2) had minimal effects. These results demonstrate that BaP synergistically enhances the production of H(2)O(2) from cultured cells by UVA and, to a lesser extent, by UVB, supporting the hypothesis that interaction of BaP and UVA can generate ROS and further substantiate oxidative DNA damage that may lead to carcinogenesis., (Copyright 2003 Elsevier Science (USA))

Polycyclic aromatic hydrocarbons, including benzo[a]pyrene (BaP), are ubiquitous environmental carcinogens. BaP is metabolized in vivo to reactive intermediates that become covalently bound to DNA and form BaP-DNA adducts, an initial event in carcinogenesis. Ultraviolet A (UVA) synergizes with BaP to significantly enhance genetic damage and accelerate carcinogenic processes. This study was initiated to investigate in vivo cellular changes related to carcinogenesis induced by repeated exposures to BaP plus UVA. Simulated chronic exposure to an environmental carcinogen and sunlight was conducted through biweekly topical application of BaP followed 2 h later by UVA exposure over a 10 week period. BaP diol epoxide (BPDE)-DNA adducts were measured in vivo by immunohistochemistry using an anti-BPDE-DNA monoclonal antibody. Oxidative DNA damage was measured by the detection of 8-hydroxy-2'-deoxyguanosine (8-OHdG) formation using high-performance liquid chromatography. Alterations in the cell cycle that were relevant to carcinogenesis were revealed by changes in p53, as identified in vivo using a polyclonal anti-p53 antibody. We found that cells containing BPDE-DNA adducts and nuclear p53 expression significantly increased between 2 and 10 weeks of BaP-UVA treatment, whereas neither BPDE-DNA adducts nor significant changes in p53 were observed in untreated skin. Using regression analysis, oxidative 8-OHdG damage also showed a parallel increase over 2-10 weeks (r = 0.80). These results indicate that genetic damage caused by exposures to BaP plus UVA accumulates with time and increases the potential for inductive events leading to carcinogenesis and tumor formation.

Lycopene, an acyclic hydrocarbon carotenoid found in tomatoes and their products, is a well-established potent antioxidant, and its anticancer properties have been shown in cultured cells and animal models. We investigated the protective effects of two concentrations of topical lycopene against acute ultraviolet B (UVB)-induced photodamage. Application of lycopene dose dependently inhibited UVB-induced ornithine decarboxylase (P < 0.05) and myeloperoxidase (P < 0.05) and significantly reduced bifold skin thickness (P < 0.05). Immunohistochemical staining revealed increased active caspase-3 of apoptotic pathway in the UVB-exposed group compared with the unexposed control. Application of topical lycopene prevented the cleavage of caspase-3. UVB irradiation completely diminished proliferating cell nuclear antigen (PCNA), and the untreated skin maintained positively stained cells throughout the basal epidermis. Topical application of lycopene significantly reversed UVB-induced PCNA inhibition, and normal PCNA staining was restored in the lycopene-treated skin. Our results suggest that topical lycopene is able to exert its protective effects against acute UVB-induced photodamage. Furthermore, it may act as a preventative agent via inhibition of epidermal ornithine decarboxylase activity, reducing inflammatory responses, maintaining normal cell proliferation, and possibly preventing DNA damage as indicated by blocking the necessitating step of apoptosis following UVB injury.

Long-term psoralen plus ultraviolet A radiation (PUVA) therapy is associated with an increased risk of squamous cell carcinoma and malignant melanoma. Genistein (4',5,7-trihydroxyisoflavone), a major isoflavone in soybeans and a specific inhibitor of protein tyrosine kinase, has been shown to inhibit UVB induced skin carcinogenesis in hairless mice. For this study we examined the protective effects of topical genistein on PUVA-induced photodamage. In two separate experiments, genistein in a dimethyl sulfoxide/acetone (1:9) solution was applied to SKH-1 female mice 1 h post 8-methoxy-psoralen dosing and 1 h prior to UVA irradiation. Application of genistein significantly decreased PUVA-induced skin thickening, and greatly diminished cutaneous erythema and ulceration in a dose-dependent manner. Histological examination showed that PUVA treatment of mouse skin induced dramatic inflammatory changes throughout the epidermis; topical genistein prevented these changes without noticeable adverse effects. Cells containing cleaved poly(ADP-ribose) polymerase (PARP) and active caspase-3 were significantly increased in PUVA-treated skin (P < 0.05 and P < 0.0001, respectively) as compared with unexposed control skin. Topical genistein completely inhibited cleavage of PARP and caspase-3. Proliferating cell nuclear antigen (PCNA) positive cells were observed in suprabasal areas of the epidermis and were significantly decreased in PUVA-treated skin compared with both control samples and samples treated with PUVA plus topical genistein (P < 0.005). These results indicate that genistein protects the skin from PUVA-induced photodamage.