Effect of adriamycin combined with Mcl-1 inhibitor Marinopyrrole A on drug resistance and apoptotic protein expression in hepatocellular carcinoma cells
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摘要 目的: 探究阿霉素（ADM）与髓样细胞白血病-1（Mcl-1）选择性抑制剂Marinopyrrole A联合使用抗肝癌活性及相关分子机制。方法: 以肝癌Huh7细胞和HepG2细胞为研究对象，采用MTT法检测ADM、Marinopyrrole A及两者联用对肝癌细胞的抑制作用；光学显微镜下观察细胞形态学变化；流式细胞仪检测不同处理组细胞的凋亡率，蛋白质免疫印迹（western blotting）法检测细胞多药耐药蛋白1（MDR1）、穹窿主体蛋白（MVP）、Mcl-1、剪切型多聚ADP核糖聚合酶（Cleaved-PARP）/PARP、Bcl-2、Bax等耐药和凋亡相关蛋白的表达。结果: ADM对HepG2和Huh7细胞的半数抑制浓度（IC50）分别为（3.557±0.640）μmol/L、（1.178±0.127）μmol/L，HepG2细胞对ADM的敏感性低于Huh7细胞（P＜0.01）。HepG2细胞的MDR1、MVP和Mcl-1蛋白表达水平均高于Huh7细胞（均P＜0.01）；与control组和单独ADM处理组比较，Marinopyrrole A联合ADM可使HepG2细胞密度减少，细胞皱缩变小，细胞凋亡率显著升高，MDR1、MVP、Mcl-1蛋白表达下调，Bcl-2/Bax比值减小，Cleaved-PARP/PARP比值增加（均P＜0.05）。结论: Mcl-1抑制剂Marinopyrrole A可增加肝癌细胞对ADM的敏感性，可能与下调耐药相关蛋白MDR1、MVP和抗凋亡蛋白Mcl-1的表达有关。
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关键词
- 肝癌 /
- 阿霉素 /
- Marinopyrrole A /
- 髓样细胞白血病-1抑制剂 /
- 联合用药
Abstract Objective: To explore the anti-hepatoma activity and related molecular mechanism of adriamycin (ADM) combined with the selective inhibitor Marinopyrrole A of myeloid cell leukemia-1 (Mcl-1). Methods: Hepatocellular carcinoma Huh7 cells and HepG2 cells were used as research objects. The inhibitory effect of ADM, Marinopyrrole A and Marinopyrrole A combined with ADM on hepatocellular carcinoma cells were detected by the methyl thiazolyl tetrazolium (MTT) assay, and the morphological changes of the cells were observed under light microscope. The apoptosis rates of cells in different treatment groups were detected by flow cytometry. The expression of multidrug resistance protein 1 (MDR1), major vault protein (MVP), Mcl-1, Cleaved poly ADP ribose polymerase (Cleaved-PARP)/PARP, Bcl-2, Bax and other drug resistance and apoptosis-related proteins was detected by western blotting. Results: The half inhibitory concentrations (IC50) of ADM on HepG2 and Huh7 cells were (3.557 ±0.640) μmol/L and (1.178 ±0.127) μmol/L, respectively. HepG2 cells were less sensitive to ADM than Huh7 cells (P＜0.01). The protein expression levels of MDR1, MVP and Mcl-1 in HepG2 cells were higher than those in Huh7 cells (all P＜0.01). Compared with the control group and ADM treatment group alone, Marinopyrrole A combined with ADM decreased the density and shrinkage of HepG2 cells, and significantly increased the rate of apoptosis, down-regulated the expression of MDR1, MVP, and Mcl-1 proteins, decreased the Bcl-2/Bax ratio, and increased the Cleaved-PARP/PARP ratio (all P＜0.05). Conclusion: The Mcl-1 inhibitor Marinopyrrole A can increase the sensitivity of hepatocellular carcinoma cells to ADM, which may be related to down-regulating the expression of drug resistance-related proteins MDR1, MVP and anti-apoptotic protein Mcl-1. -
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