miR-222-3p对人乳头瘤病毒感染的宫颈癌细胞活性及糖酵解的作用及其机制

Effect of miR-222-3p on the activity and glycolysis of positive cervical cancer cells infected with human papillomavirus and related mechanism

目的:探讨miR-222-3p对人乳头瘤病毒(HPV)感染阳性宫颈癌细胞活性及糖酵解的作用及其机制。方法:将Siha细胞分为宫颈癌组(未转染的HPV感染Siha细胞)、miR-222-3p-NC组(HPV感染Siha细胞转染miR-222-3p-NC)、miR-222-3p mimics组(HPV感染Siha细胞转染miR-222-3p mimics)及miR-222-3p siRNA组(HPV感染Siha细胞转染miR-222-3p siRNA)。qPCR检测各组Siha细胞miR-222-3p相对表达;CCK-8检测各组Siha细胞活性;流式细胞仪检测各组Siha细胞凋亡率;Transwell小室检测各组Siha细胞侵袭数目;划痕实验检测各组Siha细胞迁移距离;试剂盒检测各组Siha细胞葡萄糖及乳酸水平;免疫印迹检测各组丙酮酸激酶M2型(PKM2)、乳酸脱氢酶A(LDHA)、磷脂酰肌醇-3-激酶(PI3K)、蛋白激酶B(AKT)蛋白表达;荧光素酶证实miR-222-3p对PI3K、AKT的调控作用。结果:宫颈癌组、miR-222-3p-NC组、miR-222-3p mimics组及miR-222-3p siRNA组的Siha细胞中miR-222-3p的相对表达分别为1.00±0.00、0.96±0.02、1.33±0.09及0.60±0.04,差异有统计学意义。与miR-222-3p-NC组相比,miR-222-3p mimics组细胞增殖、侵袭及迁移、乳酸、葡萄糖、PKM2、LDHA、PI3K、AKT升高,凋亡率减少;与miR-222-3p mimics组相比,miR-222-3p siRNA组增殖、侵袭、迁移、乳酸、葡萄糖、PKM2、LDHA、PI3K、AKT均降低,凋亡率升高。荧光素酶证实抑制miR-222-3可抑制野生型PI3K、AKT表达。结论:抑制miR-222-3p可显著降低HPV感染的宫颈癌细胞增殖、迁移及恶性侵袭,促进凋亡,抑制肿瘤细胞糖酵解,作用机制可能与降低PI3K/AKT活性相关。

Objective:To investigate effect of miR-222-3p on the activity and glycolysis of positive cervical cancer cells infected with human papillomavirus and related mechanism.Methods:Siha cells were divided into cervical cancer group(untransfected HPV infected Siha cells),miR-222-3p-NC group(HPV infected Siha cells were transfected with miR-222-3p-NC),lactate dehydrogenase A group(HPV infected Siha cells were transfected with miR-222-3p mimics and miR-222-3p siRNA group(HPV infected Siha cells transfected with miR-222-3p siRNA).qPCR was used to detect the relative expression of miR-222-3p in Siha cells of each group.Siha cell activity was detected by CCK-8.The apoptosis rate of Siha cells was detected by flow cytometry.The invasion number of Siha cells in each group was detected by Transwell chamber.The migration distance of Siha cells in each group was detected by scratch test.Glucose and lactic acid levels of Siha cells were detected by kit.The protein expressions of pyruvate kinase M2(PKM2),lactate dehydrogenase A(LDHA),phosphatidylinositol 3-kinase(PI3K)and protein kinase B(AKT)were detected by Western blotting.Luciferase was used confirmed the regulation of miR-222-3p on PI3K and AKT.Results:The relative expression of miR-222-3p in Siha cells of cervical cancer group,miR-222-3p-NC group,miR-222-3p mimics group and miR-222-3p siRNA group were 1.00±0.00,0.96±0.02,1.33±0.09 and 0.60±0.04,respectively.Compared with the miR-222-3p-NC group,cell proliferation,invasion and migration,lactic acid,glucose,PKM2,LDHA,PI3K and AKT were increased in the miR-222-3p mimics group,and the apoptosis rate was decreased.Proliferation,invasion,migration,lactic acid,glucose,PKM2,LDHA,PI3K and AKT were decreased in miR-222-3p siRNA group,and apoptosis was increased.Luciferase confirmed that inhibition of miR-222-3 could inhibit the expression of wild-type PI3K and AKT.Conclusion:Inhibition of miR-222-3p can significantly reduce the proliferation,migration and malignant invasion of HPV-infected cervical cancer cells,promote apoptosis,and inhibit the glycolysis of tumor cells.The mechanism of action may be related to the decrease of PI3K/AKT activity.