36 B. E. Johnson repair process by which uvr-induced thymine dimers are removed from DNA (46). This is perhaps the first report to show that DNA is a direct target for skin reactions to uvr and it may be inferred from the results that thymine dimers are the molecular lesions involved. In addition, fluorescent antibody techniques, using antibodies raised against uv-irradiated native DNA, have been used to demonstrate damage in DNA of skin irradiated in vivo (47, 48). As thymine dimers are the most prominent of the DNA photo- products, these studies can be interpreted as demonstrating, although again indirectly, that thymine dimers are formed in skin irradiated in vivo. Wavelength dependency studies of this reaction show that it is restricted to the sunburn region of the uv-spectrum (49). Changes in intensity and distribution of the staining reaction for acid phosphatase activity in uv-irradiated skin have been interpreted as evidence that the lysosome mem- brane is a primary target for uvr damage and that lysosome enzymes released as a con- sequence of this damage may be involved in the secondary processes of sunburn (39, 50). The decrease in keratinosome number reported by Wilgram et al. (37) appears to support this hypothesis, but Honigsmann et al. (51) could find no evidence of early lysosome membrane disruption in epidermal cells of uv-irradiated skin using ultra- structural techniques. BIOPHYSICS AND BIOCHEMISTRY A specific free radical signal is produced in lightly pigmented skin on exposure to sun- burning uvr (52) but the chemical nature of this excited molecular state has not been identified. Lipid peroxide, possibly derived from free radical induced reactions in un- saturated fatty acids and relatively long lived, has been identified in uv-irradiated skin (53, 54). Lipid peroxides have considerable biological activity which includes the inhi- bition of respiration and glycolysis, oxidation of sulphydryl groups and destruction of anti-oxidants such as vitamin E. Moreover, lysosome membranes are particularly sus- ceptible to labilisation by lipid peroxides which have been incriminated in cell necrosis in carbon tetrachloride damaged liver and in inflammation of the intestinal mucosa. Peroxides isolated from uv-irradiated skin have been shown to be potent carcinogens (55) but the role, if any, of these compounds in uvr inflammation has still to be defined. Under ideal conditions, the action spectrum for a photochemical reaction should fit the absorption spectrum for the molecule involved. In the photosensitive porphyrias the action spectrum for the abnormal response fits the soret band absorption characteristic of the porphyrins. The finding of a peak of erythemal activity at around 260 nm and general activity at longer wavelengths has focused attention on nucleic acids and proteins as the primary target molecules for skin reactions to uvr. However, skin is so heterogeneous a medium, uvr energy may be absorbed and dissipated as harmless quantities of heat or as fluorescence, that such a treatment is hopeful if not naive. The action spectrum presented by Everett et al. (19) might well be interpreted as representing an absorbance in un- saturated fatty acids to initiate the reaction, deviations from the shape of a smooth absorption curve being explained in terms of the filtering effects of the horny layer. Nonetheless, DNA appears to be the major molecular target for uvr lethality and muta- genesis in bacteria and mammalian cells in culture (56, 57, 58). Although DNA-protein binding may be involved in these effects, the most prominent and contributory molecular lesion is the cyclobutane type dimer formed between adjacent thymines on a single strand of the DNA double helix. Inhibition of DNA synthesis, cell death and mutation may result directly from the presence of thymine dimers or indirectly through faulty

Sunburn and mechanisms of protection 37 repair. Excision repair has already been mentioned and is the best known mechanism in both bacteria and mammalian cells. It is a multi-enzyme process in which a segment of the DNA strand containing a dimer is removed and repair synthesis, based on the tem- plate of the sister strand, fills in the gap. The process is completed when the end of the newly synthesised segment is joined into the parent strand. A second repair mechanism, termed recombinational or post-replication repair (59, 60) involves the filling in of gaps left in daughter strand DNA after replication in which thymine dimers in the template are by-passed by DNA polymerase. A third form, photoreactivation, is a uv-A stimulated, enzymic splitting of the dimers. Although its action is well established in bacteria and most animal cells, higher mammals do not appear to possess the typical photoreactiva- tion enzyme (61). Photoreactivation does not occur in higher mammalian cells in culture but some form of uv-A induced recovery of sunburn damage has been demonstrated, and Sutherland (62) has isolated a photoreactivation type enzyme from human leukocytes. Until the late 1960's and the autoradiographic studies of Fukuyama, Epstein and Epstein (63), Epstein, Fukuyama and Epstein (64) and Epstein, Fukuyama and Epstein (45), few studies of the possible involvement of nucleic acids in the reactions of skin to uvr had been attempted. Tickner (64), although demonstrating a decrease in phospho- lipid content, a finding which drew attention to the uvr effects on cellular membranes, could find no change in DNA or RNA content of mouse skin exposed in vitro to massive doses of uvr. This is perhaps not surprising in view of the now known photochemistry of the nucleic acids. Baden and Pearlman (66) foreshadowed the autoradiographic studies by using chemical extraction methods to demonstrate that synthesis of RblA and protein is inhibited in mammalian epidermis shortly after uv-irradiation in vivo, recovering by 24 h and being increased at 72 h after exposure. A similar study of DNA synthesis (67) shows it to be inhibited shortly after exposure, the degree of inhibition and its duration depending on the exposure dose. In 1970, Pathak and his co-workers reported the first isolation of thymine dimers from uv-irradiated skin, confirming this report later and showing that the effect was restricted to the sunburning wavelengths of uvr (68, 69). Bowden et al. (70) isolated thymine dimers from uv-irradiated mouse epidermis and appear to have demonstrated a form of repair replication to take place, although excision repair in rodent cells is thought to operate only at very low efficiency, if at all. Cooke and Johnson (71) have also isolated thymine dimers from mouse skin exposed to uvr in vivo but could find little evidence of excision repair, the dimers being retained in the skin for at least 24 h. Cooke's extensive studies of DNA involvement in mouse skin reactions to uvr have failed to demonstrate a clear cut relationship between thymine dimer formation and inhibition of DNA syn- thesis or the vascular reactions produced. For example, the dimer yield after exposure to 'sunlamp' fluorescent tube irradiation, peak emission around 315 nm, is very low but severe skin reactions are elicited and inhibition of DNA synthesis appears relatively efficient. The involvement of DNA in human-skin reactions to uvr might be inferred from the findings that repair processes may be deficient in cells from patients with the various forms of Xeroderma Pigmentosum (72, 73, 74). The major feature of this autosomal recessive disease is an abnormal skin sensitivity to sunlight. Increased epidermal damage and accelerated development of the chronic effects of exposure such as premature ageing and cancer of the skin appear to be magnified normal skin responses to uvr, and for these re- actions DNA may well be the primary molecular target. However, the vascular reactions