组蛋白去乙酰化酶抑制剂丙戊酸肝细胞毒性相关作用机制研究

Study on the hepatotoxicity mechanism of histone deacetylase inhibitor-valproic acid

目的研究丙戊酸肝细胞毒性相关机制。方法使用人肝癌细胞系HepG2,VPA、TSA处理,检测细胞活性、凋亡情况、线粒体膜电位、PI3K/AKT/mTOR通路活性、Bcl-2、Bax凋亡通路和自噬激活情况。结果5 mmol/L、10 mmol/L VPA处理24 h,细胞活性明显降低;5μmol/L TSA处理24 h,细胞活性明显降低。因此选用10 mmol/L VPA、5μmol/L TSA。10 mmol/L VPA处理24 h或5μmol/L TSA处理24 h,细胞凋亡率显著升高,细胞线粒体膜电位降低,细胞PI3K/AKT/mTOR通路活性降低,细胞Bcl-2活性降低,Bax活性升高,cleaved-caspase-3比例升高,细胞LC3Ⅱ表达增加,p62表达降低,VPA的作用强于TSA。结论 PI3K/Akt/mTOR通路抑制以及线粒体异常引起的凋亡在VPA相关肝毒性中发挥重要作用,而且VPA的肝毒性也与其HDAC抑制剂活性具有一定关联。

Objective To study the hepatotoxicity mechanism of valproic acid.Methods Human hepatoma cell lines HepG2,VPA,and TSA were used to detect the cell viability,apoptosis,mitochondrial membrane potential,activity of PI3K/AKT/mTOR pathway,Bcl-2,Bax apoptosis pathway and autophagy activation.Results After being treated with 5 mmol/L and 10 mmol/L VPA for 24 hours,the cell viability of HepG2 was significantly reduced.After being treated with 5 μmol/L TSA for 24 hours,the cell viability of HepG2 was significantly reduced.Therefore,10 mmol/L VPA and 5 μmol/L TSA were chosen.After being treated with 10 mmol/L VPA for 24 hours or 5 μmol/L TSA for 24 hours,the apoptotic rate of cells was significantly increased;the mitochondrial membrane potential was decreased;the activity of PI3K/AKT/mTOR pathway and Bcl-2 was decreased;the activity of Bax was increased;the ratio of cleaved-caspase-3 and the expression of LC3II were increased;the expression of p62 was decreased;the effect of VPA was better than that of TSA.Conclusion PI3K/Akt/mTOR pathway inhibition,as well as apoptosis caused by mitochondrial abnormalities,plays an important role in VPA-associated hepatotoxicity,and hepatotoxicity of VPA is also associated with its HDAC inhibitor activity.