摘要
目的探讨微小RNA-93(miRNA-93)对子宫内膜癌细胞增殖的影响,检测同源丢失性磷酸酶-张力蛋白(PTEN)/磷脂酰肌醇-3-激酶(PI3K)/蛋白激酶B(AKT)/哺乳动物雷帕霉素靶蛋白(m TOR)信号通路相关蛋白,分析miRNA-93对该通路的调控作用,探讨miRNA-93在子宫内膜癌发生、发展中可能的作用及机制。方法实验起止时间:2022年3月—2023年8月。将人子宫内膜癌Ishikawa细胞分为5组,分别为空白对照组(NC组)、miRNA-93激动剂组(miRNA-93 mimic组)、miRNA-93激动剂阴性对照组(miRNA-93 mimic NC组)、miRNA-93抑制剂组(miRNA-93 inhibitor组)、miRNA-93抑制剂阴性对照组(miRNA-93 inhibitor NC组),进行定量逆转录聚合酶链反应(qRT-PCR),比较各组转染后miRNA-93表达水平。采用CCK-8法检测过表达和抑制miRNA-93对人子宫内膜癌Ishikawa细胞增殖的影响。采用Western blot法检测PTEN、PI3K、AKT、p-AKT、m TOR等通路相关蛋白表达水平。结果转染人子宫内膜癌Ishikawa细胞后,NC组、miRNA-93mimic组、miRNA-93 mimic NC组、miRNA-93 inhibitor组、miRNA-93 inhibitor NC组miRNA-93相对表达水平分别为(1.00±0.11)、(197.25±23.32)、(0.43±0.03)、(0.24±0.02)、(0.46±0.04)。miRNA-93 mimic组miRNA-93相对表达水平高于NC组、miRNA-93 mimic NC组(均P<0.05),说明miRNA-93过表达组成功建立。miRNA-93 inhibitor组miRNA-93相对表达水平低于NC组、miRNA-93 inhibitor NC组(均P<0.05),说明miRNA-93抑制组成功建立。与miRNA-93 inhibitor组吸光度(OD值)[24 h(0.47±0.01)、48 h(0.65±0.01)、72 h(0.86±0.04)]相比,miRNA-93 mimic组OD值[24 h(0.66±0.01)、48h(1.20±0.05)、72 h(1.41±0.03)]均显著升高(均P<0.05),表明过表达miRNA-93显著提高人子宫内膜癌Ishikawa细胞增殖活性。与miRNA-93 inhibitor组[PTEN(1.17±0.08)、p-AKT(0.29±0.02)、p-AKT/AKT比值(0.33±0.01)、m TOR(0.40±0.03)]相比,miRNA-93 mimic组PTEN蛋白表达量(0.50±0.10)显著降低,p-AKT蛋白表达量、p-AKT/AKT比值、m TOR蛋白表达量[(0.97±0.04)、(1.17±0.08)、(1.20±0.09)]均显著升高(均P<0.05),表明过表达miRNA-93可抑制PTEN表达,增加p-AKT、m TOR表达,提高p-AKT/AKT比值,促进AKT磷酸化。结论miRNA-93可以通过调控PTEN/PI3K/AKT/m TOR信号通路促进子宫内膜癌细胞增殖。
Objective To explore the effect of microRNA-93(miRNA-93)on proliferation of endometrial cancer cells,detect the related proteins of phosphatase and tensin homologue(PTEN)/phosphatidylinositol-3-kinase(PI3K)/protein kinase B(AKT)/mammalian target of rapamycin(mTOR)signaling pathway,analyze the regulatory role of miRNA-93 in this pathway,and investigate the possible role and mechanism of miRNA-93 in the occurrence and development of endometrial cancer.Methods The duration time of the experiment was from March 2022 to August 2023.Human endometrial cancer Ishikawa cells were divided into 5 groups:NC group,miRNA-93 agonist group(miRNA-93 mimic group),miRNA-93 agonist negative control group(miRNA-93 mimic NC group),miRNA-93 inhibitor group(miRNA-93 inhibitor group),and miRNA-93 inhibitor negative control group(miRNA-93 inhibitor NC group),quantitative reverse transcription polymerase chain reaction(qRT-PCR)was conducted,and the expression levels of miRNA-93 after transfection in each group were compared.The effects of overexpression and inhibition of miRNA-93 on proliferation of human endometrial cancer Ishikawa cells were detected by CCK-8 method.The expression levels of pathway-related proteins such as PTEN,PI3K,AKT,p-AKT,and mTOR were detected by Western blot method.Results After transfection of human endometrial cancer Ishikawa cells,the relative expression levels of miRNA-93 in NC group,miRNA-93 mimic group,miRNA-93 mimic NC group,miRNA-93 inhibitor group,and miRNA-93 inhibitor NC group were(1.00±0.11),(197.25±23.32),(0.43±0.03),(0.24±0.02),and(0.46±0.04),respectively.The relative expression level of miRNA-93 in miRNA-93 mimic group was higher than those in NC group and miRNA-93 mimic NC group(both P<0.05),indicating the successful establishment of miRNA-93 overexpression group.The relative expression level of miRNA-93 in miRNA-93 inhibitor group was lower than those in NC group and miRNA-93 inhibitor NC group(both P<0.05),indicating the successful establishment of miRNA-93 inhibition group.Compared with optical densities(OD values)in miRNA-93 inhibitor group[24 hours(0.47±0.01),48 hours(0.65±0.01),72 hours(0.86±0.04)],OD values in miRNA-93 mimic group[24 hours(0.66±0.01),48 hours(1.20±0.05),72 hours(1.41±0.03)]increased significantly(all P<0.05),indicates that overexpression of miRNA-93 significantly enhanced proliferation activity of human endometrial cancer Ishikawa cells.Compared with miRNA-93 inhibitor group[PTEN(1.17±0.08),p-AKT(0.29±0.02),the ratio of p-AKT/AKT(0.33±0.01),mTOR(0.40±0.03)],the expression level of PTEN protein in miRNA-93 mimic group(0.50±0.10)significantly decreased,the expression levels of p-AKT protein,the ratio of p-AKT/AKT,and the expression level of mTOR protein[(0.97±0.04),(1.17±0.08),(1.20±0.09)]significantly increased(all P<0.05),indicating that overexpression of miRNA-93 could inhibit the expression of PTEN,increase the expressions of p-AKT and mTOR,raise the ratio of p-AKT/AKT,and promote phosphorylation of AKT.Conclusion miRNA-93 can promote proliferation of endometrial cancer cells by regulating PTEN/PI3K/AKT/mTOR signaling pathway.
作者
陈玲玲
刘小梅
何嘏娜
林煜
CHEN Ling-ling;LIU Xiao-mei;HE Gu-na;LIN Yu(Fuzhou Second General Hospital,Fuzhou,Fujian 350007,China)
出处
《中国妇幼保健》
2025年第21期4028-4032,共5页
Maternal and Child Health Care of China
基金
福建省福州市卫生健康中青年科学研究项目(2021-S-wq18)。
