Masami Yamaji

8-Methoxypsoralen plus UVA induces the 72-kDa heat shock protein in organ-cultured normal human skin

Abstract The proteins induced by heat and other stressors, called heat shock proteins (HSP) or stress proteins, are considered to play a general role in protection from cellular injury. Exposure to UVA (320400 nm) following application of 8‐methoxypsoralen (8‐MOP), termed PUVA is commonly used in the field of dermatology. In order to understand the induction of HSP in PUVA‐treated human skin, indirect immunofluorescence using a monoclonal antibody specific for the 72 kDa HSP (HSP 72) was carried out in organ‐cultured normal human skin that was treated with PUVA. When the organ‐cultured skin was treated at 37°C for 1 h with 8‐MOP at a final concentration of 10 or 100 μg/mL and exposed to UVA (51.3 kJ/m2), nuclear immunofluorcscence of HSP 72 was detected in the epidermal cells 12 h after UVA irradiation. In contrast, the induction of HSP 72 was not detected either by UVA irradiation or 8‐MOP treatment. These results suggest that PUVA treatment is one of the stressors for human skin, and DNA damage caused by PUVA induces HSP 72.

Induction and repair of UVB-induced cyclobutane pyrimidine dimers and (6-4) photoproducts in organ-cultured normal human skin

SummaryTo examine the induction and repair of UV-induced DNA damage, indirect immunofluorescence was performed on UVB-irradiated organ-cultured normal human skin using monoclonal antibodies specific for either cyclobutane pyrimidine dimers or (6-4) photoproducts. Nuclear immunofluorescence of cyclobutane pyrimidine dimers and (6-4) photoproducts were observed in a dosedependent manner after UVB irradiation. The intensity of nuclear immunofluorescence of the upper epidermal layers was stronger and clearer than that of the lower epidermal layers. DNA repair time-course studies showed that both types of DNA damage could be repaired within 24 h after UVB irradiation.

Melanin reduces ultraviolet-induced DNA damage formation and cell killing rate in cultured human melanoma cells

Epidermal melanin pigment is believed to prevent development of ultraviolet (UV)-induced skin cancer by shielding cell nuclei and reducing DNA damage formation. It has not been experimentally proved, however, whether melanin reduces UV-induced DNA damage, because published experiments have been inconclusive. The present study was carried out to determine whether intracellular melanin protected cultured cells against UV-induced DNA damage and killing. Three human melanoma cell lines containing different amounts of melanin were used. Absorption spectrum, subcellular localization of melanin, and melanin concentration were examined in the three cell lines. Two types of DNA damage, cyclobutane pyrimidine dimers and (6-4)photoproducts, were detected by an enzyme-linked immunosorbent assay (ELISA) with monoclonal antibodies specific for these photolesions. We found that melanin reduced the induction rates of both types of DNA damage in pigmented cells irradiated with low doses of UV in a melanin concentration-dependent manner. Almost no differences in repair capacity for the two types of photolesions were observed among the three melanoma cell lines. We also found that the more highly melanotic melanoma cell lines were more UV resistant than the less melanotic melanoma cell lines. These results suggest that intracellular melanin plays an important role in preventing UV-induced cell killing by reducing the formation of two types of DNA damage.