甘草酸通过线粒体膜去极化诱导HPV18阳性的人宫颈癌HeLa细胞凋亡的作用研究

Study on Effect of Glycyrrhizin on Apoptosis of HPV18+ Human Cervical Carcinoma HeLa Cells Induced by Mitochondrial Membrane Depolarization

关键词:  宫颈癌  人乳头瘤病毒  甘草酸  HeLa细胞  细胞凋亡

Key words:  cervical cancer  human papilloma virus  glycyrrhizin  HeLa cells  apoptosis

摘要:

目的 分析甘草酸对人乳头瘤病毒（human papilloma virus，HPV）18阳性的人宫颈癌HeLa细胞的抗增殖和促凋亡特性及其可能的作用方式，探讨甘草酸在预防HPV感染的作用和治疗宫颈癌的潜力。方法 MTT法检测甘草酸在caspase-8，caspase-9，caspase-3抑制剂存在的情况下对HeLa细胞增殖的抑制作用，相差显微镜观察细胞形态学，4’，6-二脒基-2-苯基吲哚染色检测细胞核凝结和Hoechst 33342染色检测核形态，Annexin V-FITC/PI法分析细胞凋亡，采用caspase-8，caspase-9和caspase-3比色试剂盒测定宫颈癌细胞中caspase的活性，通过Mito Tracker Red CMX Ros染色评估线粒体膜电位（ΔΨm）。结果 与对照组相比，甘草酸可显著降低HeLa细胞活力，并伴随着核浓缩和DNA碎片的增加，呈剂量依赖性。甘草酸可通过线粒体去极化诱导宫颈癌HeLa细胞凋亡，并诱导caspase-8，caspase-9和caspase-3活化。结论 甘草酸对宫颈癌细胞的抗增殖和促凋亡特性可能是通过诱导线粒体膜电位的破坏，激活细胞内和细胞外途径的caspases活性，甘草酸可作为预防和控制宫颈癌的一种辅助手段。

Abstract:

OBJECTIVE To analyze the anti-proliferative and apoptotic properties of glycyrrhizin on human papilloma virus(HPV)18-positive HeLa cells in cervical cancer and its possible mode of action, and to explore the potential of glycyrrhizin in the prevention of HPV infection and treatment of cervical cancer. METHODS MTT assay was used to detect the inhibitory effect of glycyrrhizin on HeLa cell proliferation and the inhibition of caspase-8, caspase-9 and caspase-3 inhibitors. Cell morphology was observed by phase contrast microscopy. Nuclear condensation was detected by 4’,6-diamidino-2-phenylindole staining and nuclear morphology was detected by Hoechst 33342 staining. Apoptosis was analyzed by Annexin V-FITC/PI method. The activity of caspase in cervical cancer cells was determined by caspase-8, caspase-9 and caspase-3 colorimetric kits, and the mitochondrial membrane potential(ΔΨm) was evaluated by Mito Tracker Red CMX Ros staining. RESULTS Compared with control group, exposure to glycyrrhizin significantly reduced HeLa cell viability, accompanied by nuclear condensation and increased DNA fragmentation, in a dose-dependent manner. Glycyrrhizin could induce apoptosis by mitochondrial depolarization and could induce caspase-8, caspase-9 and caspase-3 activation in cervical cancer HeLa cells. CONCLUSION The anti-proliferative and apoptotic properties of glycyrrhizin on cervical cancer cells may be through the induction of disruption of mitochondrial membrane potential and activation of caspases activity in intracellular and extracellular pathways. Glycyrrhizin can be used as an adjunct prevent and manage cervical cancer.