洛伐他汀联合顺铂对人肝癌HepG2细胞生物学特性的影响及其机制

Effect and Its Mechanism of Lovastatin Combined with Cisplatin on Biological Characteristics of Human Hepatic Cancer Cells HepG2

关键词:  洛伐他汀  肝癌  克隆形成  迁移  侵袭  凋亡

Key words:  lovastatin  hepatocellular carcinoma  clone formation  migration  invasion  apoptosis

摘要:

目的 研究洛伐他汀单用或与化疗药物顺铂联用对人肝癌HepG2细胞生物学特性的影响，初步探索其抗肿瘤作用机制。方法 不同浓度洛伐他汀、洛伐他汀联合顺铂处理细胞48 h后，CCK-8法检测HepG2细胞的增殖抑制作用，金氏公式计算联合应用效果；平板克隆形成试验评价药物作用于肝癌细胞的远期效应；划痕试验检测药物对细胞迁移能力的影响；Transwell小室法检测药物对细胞侵袭能力的影响；流式细胞术检测药物处理后细胞周期和凋亡情况；Western blotting检测凋亡相关蛋白Bcl-2、Bax、caspase-3的表达水平变化。结果 洛伐他汀呈浓度依赖性抑制HepG2细胞的增殖，金氏公式计算结果显示洛伐他汀可协同增强顺铂的抗肿瘤作用；平板克隆形成试验结果表明洛伐他汀能显著抑制HepG2细胞克隆形成率；划痕试验提示洛伐他汀能显著降低肝癌细胞的迁移率；Transwell小室侵袭实验结果发现洛伐他汀能明显减少穿膜细胞数量；流式细胞检测发现洛伐他汀可引起G₀/G₁期细胞增加，S期细胞减少，细胞凋亡率增加；Western blotting检测结果显示洛伐他汀可下调Bcl-2蛋白表达，同时上调Bax和caspase-3蛋白表达。结论 洛伐他汀可明显抑制HepG2细胞增殖、迁移和侵袭能力，与顺铂联用可增强顺铂体外抗肿瘤效果，通过线粒体途径诱导肿瘤细胞凋亡是其可能的抗肿瘤作用机制之一。

Abstract:

OBJECTIVE To investigate the effect of lovastatin separately or in combination with cisplatin on biological characteristics of human hepatic cancer cells HepG2 and to explore the molecular mechanisms. METHODS After 48 h of treatment cells with different concentrations of lovastatin and lovastatin combined with cisplatin, the proliferation inhibitory effect on HepG2 cells was assessed by CCK-8 assay. Jin's Q value was adopted to judge the efficacy of the joint-medicinal application. The long-term effects of drugs on hepatoma cells was studied by plate cloning test. The motile ability was detected by wound healing test and invasion ability was determined by Transwell assay. The cell cycle changes and apoptotic ratio was detected by flow cytometry. The expression of Bcl-2, Bax, and caspase-3 was examined with Western blotting. RESULTS Lovastatin inhibited the proliferation of HepG2 cells in a concentration-dependent manner. The results of Jin's Q value showed that lovastatin could enhance the anti-tumor effect of cisplatin. Plate cloning test showed that lovastatin could significantly inhibit HepG2 cells clone formation rate. Wound healing assay showed that lovastatin can significantly reduce the mobility of hepatocellular carcinoma cells. Transwell cell invasion test showed that lovastatin significantly reduced the number of perforating cells. The flow cytometry showed that lovastatin caused cell cycle arrest at G₀/G₁ phase with the reduction of S phase cell, the apoptotic ratio was increased by lovastatin. Western blotting assay confirmed that lovastatin could decrease the expression of Bcl-2, and increase the expression of Bax and caspase-3. CONCLUSION Lovastatin can inhibit the ability of proliferation, migration and invasion of HepG2 cells. The combination of lovastatin and cisplatin can enhance the anti-tumor effect of cisplatin in vitro. The mechanisms may be induce apoptosis through mitochondrial pathway.