雷公藤内酯醇联合热疗对肝癌细胞增殖、凋亡及p62蛋白表达的影响

Effects of TPL and hyperthermia on proliferation,apoptosis,and expression of p62 protein of human hepatoma cells

目的观察雷公藤内酯醇(TPL)联合热疗对肝癌细胞增殖、凋亡及p62蛋白表达的影响。方法将对数生长期的肝癌HepG2细胞随机分为4组,对照组不处理,TPL组加入终浓度20 nmol/L的TPL作用24 h,热疗组于43℃加热150 min,联合组加入TPL后热疗处理。取各组细胞,采用MTT法测算细胞增殖抑制率,流式细胞仪测算细胞凋亡率,Western blotting法检测自噬相关蛋白p62。结果联合组细胞增殖抑制率高于TPL组、热疗组(P均<0. 05),TPL组、热疗组细胞增殖抑制率差异无统计学意义。细胞凋亡率对照组<TPL组、热疗组<联合组(P均<0. 05),TPL组、热疗组细胞凋亡率差异无统计学意义。细胞中p62蛋白相对表达量对照组<TPL组、热疗组<联合组(P均<0. 05),TPL组、热疗组细胞中p62蛋白相对表达量差异无统计学意义。结论 TPL联合热疗较单一治疗可更有效地抑制肝癌细胞增殖并促进其凋亡,该作用有可能与其上调p62蛋白表达抑制细胞自噬有关。

Objective To study the effects of triptolide(TPL)and hyperthermia on proliferation,apoptosis,and the expression of p62 protein of human hepatoma cell line HepG2.Methods HepG2 cells were divided into four groups:the control group without any treatment,TPL group which was added with TPL at a final concertration of 20 nmol/L for 24 h,hyperthermia group which was heated at 43℃for 150 min,and the combination group which was added with TPL and then received hyperthermia.The 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide(MTT)assay was adopted to detect the proliferation rates of cells.The apoptosis was observed by flow cytometry(FCM).The expression of p62 protein was detected by Western blotting.Results The proliferative inhibition rate in the combination group was higher than those of the TPL group and hyperthermia group(P<0.05).There was no statistically significant difference in the cell proliferation inhibition rate between the TPL group and hyperthermia group.The apoptosis rate was in the following order:the control group<TPL group,hyperthermia group<the combination group(P<0.05).No statistically significant difference was found in the apoptosis rate between the TPL group and hyperthermia group.The relative expression of p62 protein was in the following order:the control group<TPL group,hyperthermia group<the combination group(P<0.05),and there was no significant difference in the expression of p62 protein between the TPL group and hyperthermia group.ConclusionTPL combined hyperthermia can significantly accelerate apoptosis and inhibit the proliferation of hepatoma cells by up-regulating the expression of p62 protein and inhibiting the cell autophagy.