顺铂通过调节FTO的表达诱导卵巢颗粒细胞凋亡  

Cisplatin induces apoptosis of ovarian granulosa cells by regulating FTO expression

作　　者：何晶晶 洪名云[1] 戴志俊[1] 朱凯[2] 刘炯炯 HE Jingjing;HONG Mingyun;DAI Zhijun;ZHU Kai;LIU Jiongjiong(Reproductive Medicine Center,Hefei Maternal and Child Health Hospital,Hefei 230001,Anhui,China)

机构地区：[1]合肥市妇幼保健院生殖医学中心,安徽合肥230001 [2]中国科学技术大学附属第一医院安徽省立医院儿外科,安徽合肥230001 [3]中国科学技术大学附属第一医院安徽省立医院国际医疗部,安徽合肥230001

出　　处：《皖南医学院学报》2024年第5期418-421,425,共5页Journal of Wannan Medical College

基　　金：安徽省卫生健康委科研项目立项项目(AHWJ2021b127);安徽省级临床重点专科建设项目(皖卫函2023-320号)。

摘　　要：目的:观察顺铂处理人卵巢颗粒细胞KGN后,细胞中脂肪和肥胖相关基因(FTO)的表达水平和细胞凋亡的变化,探讨FTO基因对化疗后KGN细胞凋亡的调控。方法:分别使用0、5、10、15、20μmol/L顺铂处理KGN细胞24、48、72 h, CCK-8法分析细胞增殖能力变化,流式细胞术检测处理48 h后细胞的凋亡水平,Western blot(WB)法检测细胞中FTO、Bcl-2和Bax蛋白的表达水平。KGN细胞转染针对FTO基因的小干扰RNA(siRNA)(siFTO组),并设立细胞对照组(Control组,只含有脂质体)和siRNA对照组(siNC组,转染阴性对照序列);KGN细胞转染FTO真核表达质粒(FTO组),并设立细胞对照组(Control组,只含有脂质体)和空载体对照组(Vector组,转染空载体)。转染后,同时使用5μmol/L顺铂处理细胞48 h;分别采用qRT-PCR和WB法检测各组FTO、Bcl-2和Bax的表达水平。结果:随着顺铂浓度的增加,各处理时间组KGN细胞的增殖能力均逐渐下降(P<0.05)。细胞凋亡水平随着顺铂浓度的增加逐渐升高(P<0.05),FTO的表达水平逐渐下降(P<0.05)。使用siRNA下调FTO后,抑凋亡蛋白Bcl-2的表达水平下降(P<0.001),促凋亡蛋白Bax的表达水平升高(P<0.001);使用真核表达质粒上调FTO后,抑凋亡蛋白Bcl-2的表达升高(P<0.01),促凋亡蛋白Bax的表达下降(P<0.001)。结论:顺铂能够下调KGN细胞中FTO的表达水平,并通过抑制抑凋亡蛋白Bcl-2表达和上调促凋亡蛋白Bax表达,促进KGN细胞的凋亡,提示化疗可能通过FTO表达促进卵巢颗粒细胞凋亡,引起卵巢早衰。Objective:To observe the expression of fat mass and obesity-associated protein(FTO)and the level of apoptosis in human ovarian granulocytes treated with cisplatin for investigating the effect of FTO gene on granulocyte apoptosis after chemotherapy.Methods:KGN cells were treated with cisplatin by concentration at 0,5,10,15 and 20μmol/L for 24,48 and 72 h,respectively.The changes in cell proliferation ability were analyzed using CCK-8 method,the level of apoptosis was detected by flow cytometry after 48 hours of treatment.Western blot was used to measure the expression levels of FTO,Bcl-2 and Bax proteins in the cells.KGN cells were transfected with small interfering RNA(siRNA)targeting the FTO gene(siFTO group),and a cell control group(Control group,only containing liposomes)and a siRNA control group(siNC group,transfected with negative control sequences)were established.KGN cells were then transfected with FTO eukaryotic expression plasmids(FTO group),and a cell control group(Control group,only containing liposomes)and an empty vector control group(Vector group,transfected with empty vectors)were established.After transfection,cells were simultaneously treated with 5μmol/L cisplatin for 48 h.finally,the expression levels of FTO,Bcl-2,and Bax in each group were detected using qRT-PCR and Western blot.Results:The proliferation ability of KGN cells gradually decreased in each treatment time group with added concentration of cisplatin(P<0.05),and the level of apoptosis was gradually increased with cisplatin concentration addition(P<0.05),yet FTO expression was gradually downregulated(P<0.05).The relative expression levels of Bcl-2 decreased and Bax were increased after downregulating FTO using siRNA(both P<0.001),and the relative expression levels of Bcl-2 was increased(P<0.01),yet Bax was decreased after upregulating FTO using eukaryotic expression plasmids(P<0.001).Conclusion:Cisplatin can downregulate the expression level of FTO in KGN cells and promote apoptosis of KGN cells by inhibiting the expression of apop

关 键 词：卵巢早衰 肥胖相关蛋白 卵巢颗粒细胞 小干扰核糖核酸 细胞凋亡 

分 类 号：R11.75[医药卫生—公共卫生与预防医学] R969