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Abstract: Pheomelanin and pheomelanin precursors have been implicated as risk factors for induction of melanoma by ultraviolet radiation. The pheomelanin precursor, 5-S-cysteinyldopa, has been shown to sensitise DNA to oxidative damage by ultraviolet radiation. We here show that 5-S-cysteinyldopa significantly protects melanocytes from membrane damage (permeability) induced by ultraviolet A radiation. Thus, 5-S-cysteinyldopa, may at the same time sensitise DNA and protect membranes from damage induced by ultraviolet radiation. [Copyright &y& Elsevier]

The dominant skin pigment melanin is believed to protect human skin against several harmful effects of ultraviolet radiation. It is not clear, however, how melanin located inside melanin-producing melanocytes modulates the effect of ultraviolet radiation on melanocytes themselves. We have determined membrane damage in pigmented and unpigmented albino mouse melanocytes after ultraviolet A radiation, which is suspected to induce melanoma. Unpigmented cells were much more susceptible to ultraviolet-A-induced membrane permeability than pigmented cells. Unpigmented cells were also more susceptible to ultraviolet-A-induced lipid peroxidation than strongly pigmented cells. Furthermore, unpigmented cells were much more susceptible to ultraviolet-A-induced depletion of glutathione than pigmented cells. Reduced glutathione is known to be a major antioxidant of unpigmented skin cells such as fibroblasts and keratinocytes. To examine whether or not glutathione is also a major antioxidant in melanocytes, melanocytes were depleted of glutathione by means of buthionine sulfoximine. We found that depletion of glutathione in pigmented melanocytes did not change lipid damage induced by ultraviolet A radiation. In unpigmented melanocytes, however, depletion of glutathione significantly increased lipid damage induced by ultraviolet A radiation. Thus, pigmented melanocytes apparently contain antioxidants more potent than glutathione, protecting them from ultraviolet-A-induced membrane damage. [ABSTRACT FROM AUTHOR]

Highly pigmented, dark skin is more resistant to the harmful effects of solar ultraviolet radiation than light-colored human skin. The extent to which tanning protects skin from harmful effects including induction of skin cancer is not known, however. We have investigated whether the skin pigment, melanin, sensitizes or protects isolated DNA or nuclear DNA in melanoma cells from the induction of the premutagenic oxidative DNA base damage, 8-hydroxy-deoxyguanosine, by ultraviolet A irradiation. Synthetic eumelanin sensitized isolated DNA to induction of the oxidative DNA base damage by ultraviolet A, but it also induced the oxidative DNA base damage in the dark. To study the role of natural melanin in mammalian melanoma cells in the induction of oxidative DNA base damage, melanin synthesis was modulated 5–7-fold in the human melanoma cells GLL19 and IGR1 (which contain both pheomelanin and eumelanin) as well as in the mouse melanoma cells B16 (which contain mainly eumelanin). Increased melanin synthesis clearly did not protect against ultraviolet A-induced oxidative DNA base damage in cells. On the contrary, the human melanoma cells with high melanin content accumulated two times more 8-hydroxy-deoxyguanosine after ultraviolet A irradiation than cells with low melanin content. Furthermore, preirradiation of the human melanoma cells, IGR1, with ultraviolet A 4 h before a second ultraviolet A exposure produced an altered amount of induced 8-hydroxy-deoxyguanosine dependent on the melanin content of the cells. We conclude that stimulation of melanin synthesis, but probably not melanin itself, increases the susceptibility of human melanoma cells to induction of premutagenic oxidative DNA base damage by ultraviolet A irradiation. [ABSTRACT FROM AUTHOR]

The level of endogenous photosensitiser, protoporphyrin IX (PPIX), can be enhanced in the cells by 5-aminolevulinic acid (ALA). We investigated the effect of critical parameters such as growth state of the cells and availability of intracellular iron in modulating the level of PPIX, in human primary cultured skin fibroblasts (FEK4) maintained either in exponentially growing or growth-arrested phase, following treatment with ALA. The addition of ALA to exponentially growing cells increased the level of PPIX 6-fold relative to control cells; however, in growth-arrested cells the same treatment increased the level of PPIX up to 34-fold. The simultaneous addition of the hydrophilic iron-chelator Desferal with ALA, boosted the level of PPIX up to 47-fold in growing cells and up to 42-fold in growth-arrested cells, suggesting that iron is limiting under the latter conditions. The strict dependence of PPIX enhancement on free available iron levels was examined by the level of activation of iron regulatory protein in band shift assays. This analysis revealed that the basal level of iron regulatory protein in growth-arrested cells was 6-fold higher than in growing cells, reflecting the influence of the free available iron pool in exponentially growing cells. Interestingly, the same ratio was found between the basal level concentration of PPIX in growing and growth-arrested cells. We propose that iron regulatory protein activation could serve as a marker for developing photodynamic therapy protocols because it identifies cells and tissues with a propensity to accumulate PPIX and it is therefore likely to predict the effectiveness of such therapies. [ABSTRACT FROM AUTHOR]

Abstract: 7,8-dihydro-8-oxo-guanine (8-oxoG) is a mutagenic DNA lesion that is induced by ultraviolet A (UVA) radiation. 8-oxoG results in increased frequency of GC→TA transversion mutations. UVA-induced mutant frequency was measured in the guanine phosphoribosyl transferase (gpt) gene of Chinese hamster ovary cells (AS52) that were stably transfected to overexpress the hOGG1 protein, the human DNA repair glycosylase for 8-oxoG. This mutant frequency was compared with ultraviolet A-induced mutant frequency in AS52 cells stably transfected with the same vector without the hOGG1 gene. The mutant frequency was significantly decreased in the hOGG1 overexpressing cells irradiated with 300 and 400kJ/m2 ultraviolet A radiation, corresponding to 25% and 10% cell survival, respectively. The hOGG1 overexpressing cells repaired oxidative DNA lesions three times faster than the vector only cells as measured by a semi-automated alkaline elution assay using FPG enzyme, the bacterial OGG1 analogue, to cut DNA at oxidative base modifications. Thus, the lower mutation frequency in UVA-induced mutations of the hOGG1 overexpressing cells may be related to the increased repair of 8-oxoG. No GC→TA mutations were detected in the UVA-irradiated hOGG1 overexpressing cells. The results suggest a link between the 8-oxoG lesion and UVA-induced mutagenesis. We propose that hOGG1 has a role in maintaining genomic stability in mammalian cells after oxidative stress. [Copyright &y& Elsevier]