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目的:研究法尼基转移酶抑制剂Manumycin对人肝癌HepG2细胞的抗肿瘤作用,并探讨其诱导凋亡的分子机制。方法:采用MTT(Methythiazolyltetrazolium)法观察法尼基转移酶抑制剂Manumycin对肝癌细胞株HepG2细胞增殖的抑制作用,荧光显微镜、DNA凝胶电泳、流式细胞术等技术检测细胞凋亡,应用Westernblot方法检测bcl-2、p53、bax的蛋白水平变化。结果:法尼基转移酶抑制剂Manumycin能明显抑制HepG2细胞的生长且呈浓度依赖性,其IC50为(17.65±0.58)μmol/L。荧光显微镜检查显示Manumycin处理的HepG2细胞DAPI染色后,细胞核内可见浓染致密的颗粒荧光,典型细胞可见新月型改变,固缩或片段化的核。DNA凝胶电泳可见典型的DNA梯形带。流式细胞DNA直方图上出现典型的亚二倍体“凋亡峰”,细胞凋亡与Manumycin作用的时间和浓度相关。Manumycin能时间依赖性地诱导HepG2细胞发生G2/M期阻滞。Manumycin处理HepG2细胞后,Westernblot检测结果显示p53蛋白表达明显增加,而bcl-2蛋白和bax蛋白表达无明显变化。结论:法尼基转移酶抑制剂Manumycin对人肝癌细胞株HepG2有强烈的细胞毒作用,其分子机制可能是诱导HepG2细胞凋亡。Manumycin诱导HepG2细胞凋亡与bcl-2蛋白和bax蛋白表达水平无关,而p53蛋白表达水平的上调可能在此过程中起了一定
Objective: To study the anti-tumor effect of farnesyl transferase inhibitor Manumycin on HepG2 cells and to explore the molecular mechanism of its induction of apoptosis. METHODS: MTT (Methythiazolyltetrazolium) method was used to observe the inhibitory effect of farnesyltransferase inhibitor Manumycin on the proliferation of hepatocellular carcinoma cell line HepG2. The apoptosis was detected by fluorescence microscopy, DNA gel electrophoresis and flow cytometry. Western blot method was used. The protein levels of bcl-2, p53 and bax were detected. Results: The farnesyl transferase inhibitor Manumycin significantly inhibited the growth of HepG2 cells in a concentration-dependent manner, and its IC50 was (17.65±0.58) μmol/L. Fluorescent microscopy showed that after DAPI staining of Manumycin-treated HepG2 cells, dense and dense particle fluorescence was observed in the nucleus, typical cells could be seen with crescent-type changes, pyknosis or fragmented nuclei. DNA gel electrophoresis shows a typical DNA ladder. A typical sub-diploid “apoptotic peak” appears on the flow cytometric DNA histogram, and apoptosis is related to the time and concentration of Manumycin action. Manumycin induces G2/M arrest in HepG2 cells in a time-dependent manner. After Manumycin treatment of HepG2 cells, Western blot showed that the expression of p53 protein was significantly increased, while the expression of bcl-2 protein and bax protein had no significant changes. Conclusion: The farnesyltransferase inhibitor Manumycin has a strong cytotoxic effect on human hepatocellular carcinoma cell line HepG2, and its molecular mechanism may be to induce HepG2 cell apoptosis. The apoptosis of HepG2 cells induced by Manumycin is not related to the expression of bcl-2 protein and bax protein, but the up-regulation of p53 protein expression may play a role in this process.