摘要
目的 探究FOXO3a对子宫内膜癌细胞增殖、凋亡和细胞周期生物学行为的调控作用和分子机制。方法 选择2019年3月至2021年12月陕西省人民医院妇科收治的子宫内膜癌确诊患者45例,收集患者的子宫内膜癌组织及与病灶邻近(距离>3 cm)的正常子宫内膜组织,采用qRT-PCR检测FOXO3a基因的表达水平,采用免疫组织化学染色法和Western blot检测FOXO3a蛋白表达水平。正常培养人子宫内膜癌细胞系HEC-1B和人子宫内膜上皮细胞HEECs,采用qRT-PCR检测HEC-1B和HEECs细胞中FOXO3a的mRNA表达水平,采用Western blot检测FOXO3a蛋白表达水平。将HEC-1B细胞分为5组:对照组、si-NC组、si-FOXO3a组、pcDNA-NC组和pcDNA-FOXO3a组。采用Western blot检测Bim、FasL、TRAIL、p27、p21、c-myc、PI3K、p-PI3K、AKT和p-AKT蛋白表达水平;采用MTT法和平板克隆形成实验检测细胞增殖水平;采用流式细胞术检测细胞凋亡和细胞周期分布。结果 与正常子宫内膜组织比较,子宫内膜癌组织中FOXO3a的mRNA和蛋白表达水平均较低(P<0.01)。与HEECs细胞比较,HEC-1B细胞中FOXO3a的mRNA和蛋白表达水平均较低(P<0.01)。与对照组和si-NC组比较,si-FOXO3a组HEC-1B细胞增殖水平和细胞克隆数均升高(P<0.05),细胞处于G1期和G2/M期的比例均降低(P<0.05),S期细胞数量升高(P<0.05),细胞凋亡水平降低(P<0.05)。与对照组和pcDNA-NC组比较,pcDNA-FOXO3a组HEC-1B细胞增殖水平和细胞克隆数均降低(P<0.05),细胞处于G1期的比例升高(P<0.05),S期细胞数量降低(P<0.05),细胞凋亡水平升高(P<0.05)。与对照组和si-NC组比较,si-FOXO3a组HEC-1B细胞中Bim、FasL、TRAIL、p27和p21蛋白相对表达量均降低(P<0.05),c-myc、p-PI3K和p-AKT蛋白相对表达量均升高(P<0.05)。与对照组和pcDNA-NC组比较,pcDNA-FOXO3a组Bim、FasL、TRAIL、p27和p21蛋白相对表达量均升高(P<0.05),c-myc、p-PI3K和p-AKT蛋白相对表达量均降低(P<0.05)。结论 FOXO3a通过负向调控抑制PI3K/AKT�
Objective To investigate the regulatory effect and its molecular mechanism of FOXO3a on the proliferation,the apoptosis and the cell cycle of endometrial carcinoma cells.Methods A total of 45 patients with endometrial carcinoma admitted to the department of gynecology in Shaanxi Provincial People’s Hospital were collected from March 2019 to December 2021.And endometrial carcinoma tissues and normal endometrial tissues adjacent to the cancer lesion(distance>3 cm)were isolated and collected to detect the expression levels of FOXO3a by qRT-PCR,immunohistochemical staining and Western blot.Human endometrial cancer cell line HEC-1B and human endometrial epithelial cell line HEECs were cultured by conventional culture methods.The mRNA expression level of FOXO3a in HEC-1B and HEECs cells was detected by qRT-PCR,and the protein expression level of FOXO3a was detected by Western blot.The HEC-1B cells were divided into five groups:control group,si-NC group,si-FOXO3a group,pcDNA-NC group and pcDNA-FOXO3a group.The protein expression levels of Bim,FasL,TRAIL,p27,p21,c-myc,PI3K,p-PI3K,AKT and p-AKT were detected by Western blot.The cell proliferation level was detected by MTT assay and plate clone formation assay.The cell apoptosis and the cell cycle distribution were detected by flow cytometry.Results Compared with normal tissues,the mRNA and protein expression levels of FOXO3a were decreased in endometrial carcinoma tissues(P<0.01).Compared with HEECs cells,the mRNA and protein expression levels of FOXO3a in HEC-1B cells were decreased(P<0.01).Compared with control group and si-NC group,the HEC-1B cell proliferation and the number of cell clones were increased in si-FOXO3a group(P<0.05),the proportions of cells in G1 phase and G2/M phase were decreased(P<0.05),the number of cells in S phase was increased(P<0.05),and the level of cell apoptosis was decreased(P<0.05).Compared with control group and pcDNA-NC group,the HEC-1B cell proliferation and the number of cell clones were decreased in pcDNA-FOXO3a group(P<0.05),the pro
作者
李帆
唐阳芳
吴利英
LI Fan;TANG Yangfang;WU Liying(Department of Gynecology,Shaanxi Provincial People’s Hospital,Xi’an 710068,China;Department of Gynecology,First Affiliated Hospital of Xi’an Medical University;Department of Gynecology and Obstetrics,Xi’an People’s Hospital,Xi’an Fourth Hospital)
出处
《山西医科大学学报》
CAS
2023年第8期1029-1037,共9页
Journal of Shanxi Medical University
基金
陕西省重点研发计划项目(2022SF-553)。
