摘要
目的 探讨miR-424-5p对宫颈癌SiHa细胞增殖、迁移及凋亡的影响及其作用机制。方法 将宫颈癌SiHa细胞分为四组:mimics-NC组(转染mimics-NC)、miR-424-5p mimics组(转染miR-424-5p mimics)、inhibitor-NC组(转染inhibitor-NC)、miR-424-5p inhibitor组(转染miR-424-5p inhibitor);采用实时荧光定量聚合酶链反应(qRT-PCR)检测miR-424-5p表达水平,用扫描电子显微镜(SEM)和激光扫描共聚焦显微镜(LSCM)观察转染后宫颈癌SiHa细胞的形态学变化,用细胞增殖-毒性检测(CCK-8)法检测细胞的增殖情况,用细胞划痕愈合实验检测细胞的迁移率,用流式细胞术检测细胞的周期,用免疫蛋白印迹(WB)实验检测PI3K信号通路相关蛋白的表达,并进行统计分析。结果 miR-424-5p mimics组miR-424-5p表达水平与mimics-NC组相比明显升高(t=-5.386,P<0.05),miR-424-5p inhibitor组miR-424-5p表达水平与inhibitor-NC组相比明显降低(t=50.408,P<0.01)。miR-424-5p会导致宫颈癌SiHa细胞微绒毛及伪足明显减少,细胞凋亡增加,细胞长势变差。将四组宫颈癌SiHa细胞转染1、2、3、4天后,其细胞增殖的OD值比较差异均有统计学意义(F值分别为345.313、1 469.439、2 146.425、6 536.024,P<0.01)。四组宫颈癌SiHa细胞的迁移率比较差异有统计学意义(F=114.014,P<0.01)。miR-424-5p阻碍细胞周期的进展,使G0/G1期细胞增多,而S期、G2/M期细胞减少(F值分别为592.930、380.590、1 272.137,P<0.01)。miR-424-5p能明显下调与磷脂酰肌醇-3激酶/蛋白激酶B/西罗莫司靶蛋白(PI3K/AKT/mTOR)通路相关的蛋白分子磷酸化蛋白激酶B(p-AKT)、磷酸化西罗莫司靶蛋白(p-mTOR)的表达(F值分别为36.901、53.399,P<0.01)。结论 miR-424-5p可通过调控PI3K/AKT/mTOR信号通路有效抑制宫颈癌的增殖、迁移及细胞周期,并诱导细胞凋亡,有望成为宫颈癌治疗的新靶点。
Objective To investigate the effect and mechanism of miR-424-5 p on proliferation, migration and apoptosis of cervical cancer SiHa cells.Methods Cervical cancer SiHa cells were divided into four groups, including mimics-NC group(transfected with mimics-NC),miR-424-5 p mimics group(transfected with miR-424-5 p mimics),inhibitor-NC group(transfected with inhibitor-NC),miR-424-5 p inhibitor group(transfected with miR-424-5 p inhibitor).Quantitative real-time polymerase chain reaction(qRT-PCR) was used to assess the expression level of miR-424-5 p.Changes of the morphology of cervical cancer SiHa cells after transfection were observed under scanning electron microscope(SEM) and laser scanning confocal microscopy(LSCM).The proliferation of cells was detected by Cell Counting Kit-8(CCK-8),the migration rate of cells was detected by scratch wound healing assay, cell cycle was detected by flow cytometry, and the expression levels of proteins related to PI3 K signaling pathway was verified by Western blot(WB) assay.Statistical analysis was performed.Results The expression level of miR-424-5 p in the miR-424-5 p mimics group was significantly higher than that in the mimics-NC group(t=-5.386,P<0.05),while the expression level of miR-424-5 p in the miR-424-5 p inhibitor group was significantly lower than that in the inhibitor-NC group(t=50.408,P<0.01).The miR-424-5 p decreased the microvilli and pseudopod on the surface of cervical cancer SiHa cells, increased the number of apoptosis, and led to poor cells growth.There were significant differences in the OD values of proliferation ability of cervical cancer SiHa cells in the four groups at 1 d, 2 d, 3 d and 4 d after transfection(F=345.313,1 469.439,2 146.425 and 6 536.024,respectively, P<0.01).There were statistically significant differences in the migration rates of cervical cancer SiHa cells among the four groups(F=114.014,P<0.01).The miR-424-5 p inhibited the process of cell cycle, increased cells in G0/G1 phase, and decreased cells in S phase and G2/M phase(F=592.930,38
作者
王佳佳
常明智
罗强
张凡
WANG Jiajia;CHANG Mingzhi;LUO Qiang;ZHANG Fan(Department of Obstetrics and Gynecology,Affiliated Hospital of Youjiang Medical University for Nationalities,Guangxi Baise 533000,China;Life Science Research Center,Hebei North University,Hebei Zhangjiakou 075000,China;Department of Pathology,The First Affiliated Hospital of Hebei North University,Hebei Zhangjiakou 075000,China)
出处
《中国妇幼健康研究》
2022年第5期7-13,共7页
Chinese Journal of Woman and Child Health Research
基金
河北省财政厅专科能力建设和专科带头人培养资助项目(361009)。
