Yes相关蛋白对人宫颈癌SiHa细胞生物学行为的影响  

Effect of Yes-associated proteins on biological behaviors of human cervical cancer SiHa cells

作　　者：赵芳 李珍玲[1] 朴丽花 韩龙哲[1] 崔银姬[1] 权春姬[1,4] 金雪梅 ZHAO Fang;LI Zhenling;PIAO Lihua;HAN Longzhe;CUI Yinji;QUAN Chunji;JIN Xuemei(Department of Pathology,Affiliated Hospital,Yanbian University,Yanji 133000,China;Department of Pathology,Municipal Hospital,Heze City,Shandong Province,Heze 274000,China;Department of Histology and Embryology,College of Medical Sciences,Yanbian University,Yanji 133002,China;Department of Pathology,Hainan Provincial Women and Children’s Medical Center,Haikou 570100,China)

机构地区：[1]延边大学附属医院病理科,吉林延吉133000 [2]山东省菏泽市立医院病理科,山东菏泽274000 [3]延边大学医学院组织学与胚胎学教研室,吉林延吉133000 [4]海南省妇女儿童医学中心病理科,海南海口570100

出　　处：《吉林大学学报(医学版)》2025年第1期68-75,共8页Journal of Jilin University:Medicine Edition

基　　金：国家自然科学基金地区基金项目(81860043);国家自然科学基金青年基金项目(81902790);吉林省科技厅科技发展计划项目(20200201571JC);吉林省教育厅“十三五”科学技术项目(JJKH20200525KJ)。

摘　　要：目的:探讨Yes相关蛋白(YAP)沉默对人宫颈癌(CC)SiHa细胞增殖、迁移和侵袭能力的影响。方法:对人CC细胞株SiHa细胞进行体外培养,慢病毒YAPshRNA转染到SiHa细胞,建立稳定转染的YAP-shRNA实验组(sh-YAP组)和空载质粒对照组(对照组),采用Western blotting法检测YAP沉默效果;免疫荧光法检测2组细胞中肌动蛋白(F-actin)微丝数量及形态变化;CCK-8法、Transwell小室实验和细胞划痕实验检测2组细胞存活率、迁移及侵袭细胞数以及细胞划痕愈合率;Western blotting法检测2组细胞中上皮-间质转化(EMT)相关标记物[E钙黏蛋白(E-cadherin)和锌指转录因子(Snail)]、DNA损伤修复相关蛋白γ(γ-H2AX)和凋亡相关蛋白[c-MYC和B细胞淋巴瘤2(Bcl-2)]蛋白表达水平。结果:慢病毒YAPshRNA转染SiHa细胞后,SiHa细胞中YAP蛋白表达水平明显降低(P<0.05)。免疫荧光实验,YAP沉默后SiHa细胞中F-actin微丝稀疏且排列规则,细胞数量减少、细胞呈现蜷缩的状态。CCK-8法,与对照组比较,sh-YAP组培养24和48 h后SiHa细胞存活率明显降低(P<0.01);Transwell小室实验和细胞划痕实验,与对照组比较,sh-YAP组的SiHa细胞迁移和侵袭细胞数明显减少(P<0.01),细胞划痕愈合率明显降低(P<0.05);Western blotting法,与对照组比较,sh-YAP组细胞中E-cadherin蛋白表达水平升高(P<0.05),c-MYC、Bcl-2和γ-H2AX蛋白表达水平降低(P<0.05或P<0.01)。结论:YAP基因沉默导致人CC SiHa细胞F-actin的解聚,并调控细胞凋亡和DNA损伤修复,可能会逆转EMT进程,从而抑制肿瘤细胞增殖和迁移。Objective:To discuss the effect of Yes-associated protein(YAP)silencing on the proliferation,migration,and invasion capabilities of the human cervical cancer(CC)SiHa cells.Methods:The human CC SiHa cells were cultured in vitro,and the lentiviral YAP shRNA was transfected into the SiHa cells to establish stably transfected YAP-shRNA experimental group(sh-YAP group)and empty plasmid control group(control group).Western blotting method was used to detect the silencing effect of YAP;immunofluorescence method was used to detect the microfilament number and morphology of actin filaments(F-actin)in the cells in both groups;CCK-8 method was used to detect the survival rates of the cells in two groups;Transwell chamber assay and wound healing assay were used to detect the numbers of migration and invasion cells and scratch healing rates of the cells in two groups;Western blotting method was used to detect the expression levels of epithelial-mesenchymal transition(EMT)markers(E-cadherin and Snail),DNA damage repair-related proteins(γ-H2AX),and apoptosis-related proteins[c-MYC and B-cell lymphoma-2(Bcl-2)]in the cells in two groups.Results:The results of lentiviral YAP shRNA transfection into SiHa cells showed that the expression level of YAP protein in the SiHa cells was significantly decreased(P<0.05).The immunofluorescence results showed that after YAP silencing,the F-actin in SiHa cells was sparse and regularly arranged,with a reduced number of cells and a shriveled appearance.The CCK-8 results showed that compared with control group,the survival rate of the SiHa cells in sh-YAP group was significantly decreased cultured for 24 and 48 h(P<0.01).The results of Transwell chamber assay and the wound healing assay showed that compared with control group,the numbers of migration and invasion SiHa cells in sh-YAP group were significantly decreased(P<0.01),and the cell scratch healing rates were signifiantly decreased(P<0.05).The Western blotting results showed that compared with control group,the expression level of E-cadher

关 键 词：宫颈肿瘤 Yes相关蛋白 细胞增殖 细胞迁移 

分 类 号：R737.32[医药卫生—肿瘤] R363.21[医药卫生—临床医学]