黄芩苷对H2O2氧化损伤人黑素细胞的保护机制研究

目的:探讨黄芩苷对H2O2氧化损伤正常人黑素细胞的保护作用及其机制研究。方法:构建H2O2氧化损伤黑素细胞模型，通过观察不同浓度的黄芩苷处理H2O2氧化损伤正常人黑素细胞，采用倒置显微镜观察黄芩苷对黑素细胞形态的影响；cell counting kit-8（CCK-8）法检测黄芩苷对黑素细胞增殖活力作用；乳酸脱氢酶（lactate dehydrogenase,LDH）法检测黄芩苷对黑素细胞LDH表达的影响, 并通过流式细胞仪检测活性氧（ROS）水平，荧光实时定量RT-PCR检测过氧化氢酶（catalase,CAT）、谷胱甘肽过氧化物酶1（peroxidase 1,GPX1）mRNA表达情况，通过ELISA试剂盒检测黄芩苷和H2O2处理黑素细胞后分泌的热休克蛋白70（heat shock protein 70,HSP70）。结果:镜下观察黄芩苷给药后细胞较对照组形态明显改变，细胞数增多；黄芩苷作用黑素细胞后增殖率较对照组升高（P<0.05）；黄芩苷明显抑制黑素细胞中LDH表达，呈剂量-效应关系（P<0.05）。黄芩苷能显著促进CAT和GPX1 mRNA表达，且明显抑制HSP70分泌。结论:黄芩苷对H2O2氧化损伤的正常人黑素细胞具有明显保护作用，可为临床白癜风的治疗提供参考。

Protective effects of Baicalin to the human melanocyte cocultured with H2O2

AIM: To study the mechanism of anti-oxidant and melanocyte protection by Baicalin. METHODS: Different concentration of Baicalin andH₂O₂ were cocultured with normal human melanocyte. Cell proliferation was detected by cell counting kit-8 assay, morphology of the human melanocyte was observed by ordinary inverted microscope, and LDH assay was used to detect cell viability. For mechanism research, DCFH-DA ROS level was assayed by flow cytometry. Induced HSP70 from melanocyte was detected using HSP70 high sensitivity ELISA kit. By RT-PCR, mRNA levels of CAT and GPX1 were dectected. RESULTS: Baicalin has obvious protective effects on melanocytes damage caused byH₂O₂. Compared with control group,in Baicalin groups, cell shape was changed and increased. In 12.5, 25, 50 μmol/L Baicalin groups, the effects of on melanocyte proliferation rate were higher than the control group (P<0.05). Baicalin inhibited the expression of LDH in black pigment cells in a dose-dependent manner, there was statistically significant difference (P<0.05). After H₂O₂ oxidative damage of melanocytes, Baicalin could significantly promote the expression of CAT and GPX1 mRNA, and the secretion of HSP70 was significantly increased. CONCLUSION: Baicalin protects human melanocytes from oxidative damage by inhibitingH₂O₂-induced apoptosis. It is referential for the treatment of vitiligo.