miR-126-3p过表达对宫颈癌HeLa细胞生物学行为的影响及作用机制

Effect of miR-126-3p overexpression on biological behaviors of cervical cancer HeLa cells and its mechanism

目的探究miR-126-3p过表达对宫颈癌细胞生物学行为的影响及其作用机制。方法研究样本为来自北京生物技术有限公司的宫颈癌HeLa细胞,培养后随机分为对照组(n=9)及miR-126-3p过表达组(n=9),对照组细胞正常培养,miR-126-3p过表达组采用miR-126-3p模拟物转染。使用细胞增殖实验检测两组样本细胞活力,使用流式细胞术检测两组细胞周期,使用划痕实验、Wright染色法检测两组细胞迁移量和细胞迁移率,使用蛋白印迹检测两组Bax、Casepase-3、Bcl-2、p85β、p-PDK1、p-AKT蛋白表达,使用DAPI染色观察两组细胞凋亡小体表达。结果miR-126-3p过表达组miR-126-3p相对表达量比对照组高(3.14±0.28 vs 1.04±0.11,P=0.000),模型构建成功。miR-126-3p过表达组HeLa细胞存活率显著低于对照组(P<0.05)。DAPI染色结果显示,miR-126-3p过表达组细胞显示出凋亡小体,对照组未显示出明显的细胞凋亡。miR-126-3p过表达组G0/G1期细胞占比显著高于对照组,S期细胞占比显著低于对照组(P<0.05);miR-126-3p过表达组细胞迁移量和细胞迁移率显著低于对照组(P<0.05)。与对照组相比,miR-126-3p过表达组侵袭细胞数减少;转染24,48 h后miR-126-3p过表达组平均细胞侵袭率显著低于对照组(P<0.05)。miR-126-3p过表达组Bax、Casepase-3蛋白相对表达量显著高于对照组,Bcl-2、p85β、p-PDK1、p-AKT蛋白相对表达显著低于对照组(P<0.05)。结论miR-126-3p过表达可能通过PI3K/PDK1/AKT蛋白通路调控HeLa细胞生物学行为,包括降低癌细胞活力、迁移和侵袭,诱导宫颈癌细胞周期停滞。外源性miR-126-3p可能是宫颈癌患者治疗的新靶点之一。

Objective To investigate the effect of miR-126-3p overexpression on the biological behavior of cervical cancer cells and its mechanism.Methods Cervical cancer HeLa cells were from Beijing Biotechnology Co.,LTD.After culture,they were randomly divided into control group(n=9)and miR-126-3p overexpression group(n=9).HeLa cells in control group were cultured normally,while the cells in miR-126-3p overexpression group were transfected with miR-126-3p mimics.Cell proliferation assay was used to detect the cell viability,flow cytometry was used to detect the cell cycle,and scratch assay and Wright staining method were used to detect the cell migration volume and the cell mobility.The protein expression levels of Bax,Casepase-3,Bcl-2,p85β,p-PDK1 and p-AKT were detected by Western blotting,and the apoptotic corpussome expression was observed by DAPI staining.Results After transfection,the relative expression level of miR-126-3p in miR-126-3p overexpression group was higher than that in control group(3.14±0.28 vs 1.04±0.11,P=0.000),and the model was successfully constructed.The survival rate of HeLa cells in miR-126-3p overexpression group was significantly lower than that in control group(P<0.05).The apoptotic bodies were found in miR-126-3p overexpression group by DAPI staining,while there was no obvious apoptotic bodies in control group.The proportion of G0/G1-stage cells in miR-126-3p overexpression group was significantly higher than that in control group,and the proportion of S-stage cells was significantly lower than that in control group(P<0.05).The cell migration volume and the cell migration rate in miR-126-3p overexpression group were significantly lower than those in control group(P<0.05).Compared with control group,the number of invasive cells in miR-126-3p overexpression group was decreased,and the average cell invasion rate in miR-126-3p overexpression group was significantly lower than that in control group at 24 h and 48 h after transfection(P<0.05).The relative protein expression levels of Bax and Casepase-3 in miR-126-3p overexpression group were significantly higher than those in control group,while the relative protein expression levels of Bcl-2,p85β,p-PDK1 and p-AKT in miR-126-3p overexpression group were significantly lower than those in control group(P<0.05).Conclusion Overexpression of miR-126-3p may regulate the cell biological behavior of HeLa cells through the PI3K/PDK1/AKT pathway,reduce the cervical cancer cell activity,migration and invasion,and induce the cell cycle arrest of cervical cancer cells.Exogenous miR-126-3p may be one of the new therapeutic targets for cervical cancer patients.