白花丹素对肝癌细胞增殖迁移凋亡能力的影响及其作用机制研究

Effect and Mechanism of Plumbagin on Proliferation Migration and Apoptosis of Hepatoma Cells

本研究旨在探究天然产物白花丹素体外抗肝癌作用及其机制。选择人源和鼠源肝癌细胞SMMC-7721和Hepa 1-6为研究对象,探究白花丹素对肝癌细胞的增殖、迁移和凋亡的影响,并揭示白花丹素诱导肝癌细胞凋亡的作用机制。在本研究中,不同浓度的白花丹素处理SMMC-7721细胞和Hepa 1-6细胞,采用MTT法、平板克隆、划痕和流式细胞术等实验技术和方法测定细胞生长抑制率、增殖能力、细胞迁移能力和细胞凋亡情况。采用蛋白质免疫印迹法(Western blot)检测了细胞凋亡通路相关蛋白表达。实验结果显示,与对照组相比,白花丹素处理组肝癌细胞生长抑制率显著升高(P <0.05),迁移能力显著降低(P <0.05),且呈浓度依赖性。在IC;浓度时,流式结果显示肝癌细胞凋亡率显著上升(P <0.05),且呈时间依赖性;Western blot结果显示两种细胞的凋亡抑制蛋白Bcl-2表达量显著降低,且呈时间依赖性;SMMC-7721细胞凋亡蛋白Bax表达量、Caspase-3激活量和Caspase-9激活量在24h达到最大值;Hepa 1-6细胞的凋亡蛋白Bax表达量和Caspase-9激活量在12 h时达到最大值,Caspase-3激活量在24 h达到最大值。因此,白花丹素能抑制肝癌细胞的增殖和迁移,并能诱导细胞凋亡,其作用机制与激活Bax、Caspase-3和Caspase-9线粒体凋亡通路密切相关。

In this study,the anti-hepatocellular carcinoma of plumbagin(PLB) and its mechanism in vitro were investigated.Human hepatocellular carcinoma cell(HCC) SMMC-7721 and murine HCC Hepa 1-6 were selected to explore the effects of PLB on the proliferation,apoptosis and migration of HCC,and to reveal the mechanism of HCC apoptosis induced by PLB.SMMC-7721 and Hepa 1-6 were treated with different concentrations of PLB.MTT assay,plate cloning,scratch and flow cytometry were used to determine the cell growth inhibition rate,proliferation ability,cell migration ability and cell apoptotic effect.Western blot was used to detect the expression of proteins related to apoptosis pathway.The results showed that the growth inhibition rate of HCC in the group treated with PLB was significantly increased(P <0.05) and the migration ability was significantly decreased(P <0.05) in a concentration-dependent,compared with the DMSO group.Flow cytometry results showed that the apoptosis rate of HCC cells increased significantly at IC50 concentration(P <0.05) and was time dependent;Western blot results showed that the expression of Bcl-2,an inhibitor of apoptosis protein,was significantly decreased in a timedependent manner.The expression of apoptosis-associated protein Bax,the activation of Caspase-3 and Caspase-9 in SMMC-7721 cells reached the maximum at 24 h.The expression of Bax and the activation of Caspase-9 in Hepa 1-6 cells reached the maximum at 12 h,and the activation of Caspase-3 reached the maximum at 24 h.Therefore,PLB could inhibit the proliferation and migration of HCC cells and induce apoptosis,and its mechanism was closely related to the activation of Bax,Caspase-3 and Caspase-9 apoptotic pathways.