紧密连接蛋白Occludin通过ANKRD37调控膀胱癌自噬的机制研究  

Study on the mechanism of tight junction protein Occludin regulating autophagy in bladder cancer through ANKRD37

作　　者：杨帆 何建忠 刘野 YANG Fan;HE Jian-zhong;LIU Ye(Department of Pathology,The Fifth Affiliated Hospital,Sun Yat-sen University,Zhuhai 519000,China)

机构地区：[1]中山大学附属第五医院病理科,广东珠海519000

出　　处：《中国实验诊断学》2021年第12期1739-1746,共8页Chinese Journal of Laboratory Diagnosis

基　　金：广东省科技厅自然科学基金面上项目(No.2018A030313428)。

摘　　要：目的探讨紧密连接蛋白家族成员Occludin(OCLN)在膀胱癌中的表达情况及调控膀胱癌细胞自噬的机制和临床意义。方法 (1)分析膀胱癌中OCLN在mRNA和蛋白水平的表达变化。(2)在膀胱癌T24细胞系中,敲降或过表达OCLN后,利用蛋白质印迹法检测自噬相关蛋白LC3B,P62表达变化,同时利用qRT-PCR检测与自噬相关的ANKRD37的mRNA变化,并采用独立样本t检验的方法进行组间差异分析。结果 (1)OCLN的mRNA和蛋白表达量随着膀胱癌病理分期的升高逐渐上调;(2)通过蛋白印迹法检测分析发现,与对照组相比,在敲降T24细胞系OCLN后,shOCLN-1组和shOCLN-2组的LC3BⅡ/Ⅰ蛋白比值明显提高(P=0.0107,P=0.0008),P62蛋白的表达水平显著降低(P=0.0382,P=0.0327);相反,过表达OCLN后,LC3BⅡ/Ⅰ的比值明显降低(P=0.0238),P62蛋白的表达水平显著升高(P=0.0049);(3)通过qRT-PCR检测发现,与对照组相比,在敲降OCLN的T24细胞中,shOCLN-1组和shOCLN-2组ANKRD37的mRNA表达量下降(P=0.0108,P=0.0183),在过表达OCLN的T24细胞系中表达量显著升高(P=0.0238)。结论 OCLN在膀胱癌中高表达,抑制OCLN的表达促进膀胱癌细胞的自噬,ANKRD37是OCLN调控膀胱癌自噬的重要靶基因。Objective To explore the expression of Occludin(OCLN),a member of the tight junction protein family,in bladder cancer,and to explore the mechanism of regulating autophagy in bladder cancer cells and its clinical significance.Methods (1)Analysis the mRNA and protein expression levels of OCLN in bladder cancer.(2)In the bladder cancer T24 cell line,after knocking down or overexpressing OCLN,the expression changes of autophagy-related proteins LC3 Band P62 were detected by Western blot,and the mRNA changes of ANKRD37 related to autophagy were detected at the same time.The method of sample T test was used to analyze the difference between groups.Results (1)The mRNA and protein expression level of OCLN gradually overexpressed with the increase of the pathological stage of bladder cancer;(2)Through Western blot detection analysis,it was found that compared with the control group,knockdown the expression of OCLN in T24 cell line,the ratio of LC3 BⅡ/I protein increased significantly(P=0.0107,P=0.0008),and the expression level of P62 protein decreased significantly(P=0.0382,P=0.0327);On the contrary,after overexpression of OCLN,the ratio of LC3 BII/I decreased significantly(P=0.0238),the expression level of P62 protein was significantly increased(P=0.0049);(3)Through qRT-PCR detection,it was found that ANKRD37 mRNA expression in T24 cells that after knocked down OCLN was decreased(P=0.0108,P=0.0183)and the ANKRD37 mRNA expression level in overexpressing OCLN T24 cell line was significantly increased(P=0.0238).Conclusion OCLN is highly expressed in bladder cancer,inhibiting the expression of OCLN promotes autophagy in bladder cancer cells.ANKRD37 is an important target gene for OCLN to regulate autophagy in bladder cancer.

关 键 词：膀胱癌 OCCLUDIN ANKRD37 自噬 

分 类 号：R737.14[医药卫生—肿瘤]