羟喜树碱通过诱导自噬抗宫颈癌的机制研究

Antitumor Mechanism of Hydroxycamptothecin on Cervical Cancer by Inducing Cell Autophagy

目的探讨抗肿瘤药物羟喜树碱(HCPT)对宫颈癌HeLa细胞作用的机制。方法 CCK8检测Hela细胞增殖,Western blot检测自噬和凋亡相关蛋白的表达,GFP-LC3 shRNA转染和Hoechst染色后,荧光显微镜下观察细胞自噬特异小体量的改变以及凋亡形态学的变化。结果 HCPT抑制Hela细胞生长呈浓度依赖性(P<0.05),IC503μmol/L HCPT作用Hela细胞后,自噬相关蛋白Beclin1、p62的表达以及LC3-Ⅱ/LC3-Ⅰ比值发生改变,差异有显著性意义(P均<0.05);凋亡相关蛋白Bax、cleaved caspase-3、Bcl-2表达也发生明显改变(P均<0.05);荧光显微镜下观察Hela细胞带有GFP-LC3的自噬体增加,凋亡细胞也增多(P均<0.05)。结论 HCPT能够诱导HeLa细胞中自噬相关基因Beclin1、p62以及LC3的表达增强,从而激活细胞自噬发生;且HCPT能够激活宫颈癌HeLa细胞发生自噬,进而诱导细胞凋亡来达到抗肿瘤目的。

Objective To investigate the antineoplastic mechanism of hydroxycamptothecin （ HCPT） on cervical cancer HeLa cell line.Methods CCK8,western blot and fluorescence microscopy were used to detect cellular morphology ,autophagy-related proteins and apoptosis-related proteins and the volume and shape of autophagosome after the Hela cell was treated with HCPT,and being transfected with GFP-LC3 shRNA and stained by Hoechst .Results HCPT inhibited the growth of Hela cells in a concentration-dependent manner （P〈0.05） with IC503μmol/L.After 3μmol/L HCPT being used on Hela cells,the expression of autophagy-related proteins Beclin1 and p62 were heighten obviously and LC 3-Ⅱ/LC3-Ⅰ ratio was changed apparently （ P〈0.05）.The expression of apoptosis-related proteins Bax,cleaved caspase-3 and Bcl-2 were also varied significantly （P〈0.05）. HeLa cells with GFP-LC3 showed an increase of autophagy and apoptotic cells observed by fluorescence microscopy （P〈0.05）. Conclusion HCPT could induce the expression of autophagy-related genes Beclin1,p62 and LC3,thereby to activate cellular au-tophagy and apoptosis .Besides,HCPT could activate the autophagy in cervical cancer HeLa cells ,thereby inducing the apoptosis to achieve the purpose of anti-tumor.