组蛋白去乙酰化酶抑制剂伏立诺他抑制肝癌细胞生长的机制

The mechanism of histone deacetylase inhibitor vorinostat inhibiting the growth of hepatocellular carcinoma cells

目的:探究核酸外切酶1(exonuclease 1,EXO1)在肝癌中的作用及组蛋白去乙酰化酶(histone deacetylase,HDAC)抑制剂伏立诺他抑制肝癌细胞生长可能的作用机制。方法:构建敲降或过表达EXO1的肝癌细胞系,通过细胞表型实验探讨EXO1对肝癌细胞增殖、迁移、侵袭能力的影响。通过同源重组报告实验、免疫荧光染色和蛋白质印迹法检测EXO1在同源重组修复过程中的作用及对肝癌细胞电离辐射敏感性的影响。使用伏立诺他处理肝癌细胞,通过CCK-8法、流式细胞术和蛋白质印迹法检测伏立诺他对EXO1表达的影响及抑制肝癌细胞生长的可能机制。结果:EXO1在肝癌细胞中的高表达促进了肝癌细胞增殖、迁移和侵袭,其通过同源重组途径促进DNA双链断裂修复,降低肝癌细胞对电离辐射的敏感性。此外,EXO1表达水平升高使肝癌细胞对伏立诺他更加不敏感,而敲降EXO1可增加肝癌细胞对伏立诺他的敏感性。且伏立诺他可通过抑制EXO1的表达从而抑制肝癌细胞同源重组修复能力,进而抑制肝癌细胞生长。结论:伏立诺他可通过降低EXO1的表达水平抑制肝癌细胞同源重组修复能力。EXO1可作为肝癌的潜在治疗靶点。

Objective:To investigate the mechanism of exonuclease 1(EXO1)in hepatocellular carcinoma(HCC)and the possible mechanism of the histone deacetylase(HDAC)inhibitor vorinostat inhibiting the growth of HCC cells.Methods:HCC cell lines with knockdown or overexpression of EXO1 were constructed.And the effects of EXO1 on the proliferation,migration and invasion in HCC were investigated by cell phenotype assays.The role of EXO1 in homologous recombination(HR)repair and its effect on ionizing radiation sensitivity of cells were detected by HR reportor assay,immunofluorescence and western blots.Additionally,HCC cells were treated with vorinostat.The effect of vorinostat on EXO1 expression and the possible mechanism of inhibiting the growth of HCC cells were detected by CCK-8,flow cytometry and Western blotting.Results:High level of EXO1 promoted the proliferation,migration and invasion of HCC cells.EXO1 also promoted DNA double-strand break repair through the HR pathway and reduced cell sensitivity to ionizing radiation.In addition,elevated EXO1 levels were less sensitive to vorinostat,whereas knockdown of EXO1 increased the sensitivity of hepatocellular carcinoma cells to vorinostat.Vorinostat further inhibited the HR repair ability of HCC cells by suppressing EXO1 expression,thereby inhibiting cell growth.Conclusion:Vorinostat can inhibit HR repair ability by decreasing the expression level of EXO1,which may serve as a potential therapeutic target for HCC.