Effects of all-trans retinoic ac id on tyrosinase metabolism and Cu/Zn superoxide dismutase mRNA expression in A375 cells irradiated by ultraviolet B

Abstract

Objective To evaluate the effects of all-trans retinoic acid (ATRA) on melanin content,activity and protein expression of tyrosinase,mRNA expression of Cu/Zn superoxide dismutase (SOD) in A375 cells irradiated with ultraviolet B (UVB).Methods Cultured A375 cells were classified into 6 groups:ATRA+UVB group treated with ATRA after UVB irradiation,hydroquinone+UVB group treated with hydroquinone after UVB irradiation,UVB group and ATRA group treated with UVB irradiation and ATRA respectively,negative control group receiving no treatment.Melanin content and tyrosinase activity were determined by NaOH solubilization assay and dopa-oxidation assay respectively at 24,48 and 72 hours after the addition of ATRA into medium.Western blot was performed to detect the protein expression of tyrosinase,and real-time quantitative PCR to measure the mRNA expressions of tyrosinase and Cu/Zn SOD in A375 cells after 24-hour culture with ATRA.Results The melanin content and tyrosinase activity decreased in UVB-irradiated cells after being treated with ATRA for 24,48 and 72 hours.The protein (gray scale) and mRNA (2-△△Ct value) expression levels of tyrosinase were 0.72 ± 0.070 and 1.400 ± 0.135 respectively at 24 hours after UVB irradiation,decreased to 0.42 ± 0.056 (P ＜0.01) and 0.810 ± 0.062 (P ＜ 0.01 ) respectively after additional treatment with ATRA.The mRNA expression level of Cu/Zn SOD was 0.323 ± 0.066 in A375 cells at 24 hours after UVB irradiation,and increased to 0.625 ±0.103 (P ＜ 0.01 ) after additional treatment with ATRA.Conclusion ATRA can suppress UVB-induced increase in melanin synthesis and elevate Cu/Zn SOD level in A375 cells,likely through tyrosinase pathway. Key words: Ultraviolet rays; Cell line, tumor; Tretinoin; Melanins; Monophenol monooxygenase; Superoxide dismutase