8-甲氧补骨脂素对小鼠黑素瘤细胞黑素生成的调节及相关信号转导研究

Study on Regulation of Melanogenesis Induced by 8-Methoxypsoralen and Related Signal Transduction Pathways in Murine Melanoma Cells

目的研究光化疗法中光敏剂8 -甲氧补骨脂素(8 -MOP)在诱导白癜风色素减退皮损复色过程中所起的作用及其分子机制。方法用体外培养的B16F10鼠黑素瘤细胞作模型 ,观察了不同浓度(10～500μmol/L)8 -MOP对B16F10细胞的形态、酪氨酸酶活性和黑素含量的影响。结果50或100μmol/L浓度的8 -MOP处理细胞48h后 ,B16F10细胞的树状突明显增多延长 ,胞浆内出现较多的棕褐色颗粒。8 -MOP并能以浓度依赖方式提高酪氨酸酶活性和黑素含量 ,较对照组高2～3倍(P<0.01),且在50、100μmol/L浓度时这种作用呈现最强。此外还发现8 -MOP尚能浓度依赖地抑制细胞生长 ,100μmol/L时与对照组相比细胞抑制高达60 % ,500μmol/L时细胞几乎不能生长。用蛋白激酶A或C的激活剂 ,异丁基甲基黄嘌呤 (IBMX)和佛波酯 (TPA)与8 -MOP联合处理细胞发现8 -MOP诱导黑素生成作用能被IBMX加强 ,被长时程作用的TPA抑制。结论8 -MOP能以浓度依赖方式提高酪氨酸酶活性和黑素含量,在体外直接诱导鼠黑素瘤细胞黑素生成 ,蛋白激酶A和蛋白激酶C信号途径参与8 -MOP诱导黑素生成作用的调节。

Objective To get more insight on the molecular mechanisms involved in psoralen induced melanogenesis during repigmentation of the hypomelanotic lesions of vitiligo vulgaris under PUVA treatment. Methods The effects of various concentrations of 8 methoxypsoralen (8 MOP)(10～500μmol/L) upon cellular morphologic changes, tyrosinase activity, and melanin contents of cultured B16F10 murine melanoma cells were examined in vitro. Results 8 MOP caused a striking change in cellular morphology with many elongate cell dendrites and deposits of brown granular materials in cytoplasm, these effects were observed clearly at final concentrations between 50 and 100μmol/L of 8 MOP after treating for 48 hours. In a range of final concentrations(10～500μmol/L), there were a dose dependent stimulation of tyrosinase activity and melanin contents with a 2～3 fold increase over untreated cells ( P< 0.01 ), the maximum action was obtained at 50 and 100μmol/L. It was also found that there was a dose related inhibition of cell growth with a 60%reduction of growth at a final concentration of 100μmol/L and no growth of cells at the concentration of 500μmol/L. Furthermore, isobutyl methylxanthine(IBMX), a protein kinase A (PKA) activator, potentiated the 8 MOP upregulation of tyrosinase activity and melanin contents, however, 12 O tetradecanoyl phorbol 13 acetate (TPA),a potent activator of PKC, inhibited the upregulative effect of 8 MOP when treated for a long duration. Conclusion These results demonstrate that in murine melanoma cells 8 MOP could induce melanogenesis by increasing tyrosinase activity and melanin contents. At least, PKA and PKC-dependent signaling pathways appear to play an important role in the melanogenic effect of 8 MOP.