巨噬细胞分泌TNF-α调控CLDN10通路促进膀胱癌细胞上皮-间质转化  

Macrophages promote EMT of bladder cancer cells through secreting TNF-a to regulate CLDN10/β-catenin pathway

作　　者：韩娜 陈函 张海燕 HAN Na;CHEN Han;ZHANG Haiyan(Health Examination and Oncology Screening Center,Chongqing University Cancer Hospital,Chongqing,400030,China;Department of Urological Oncology,Chongqing University Cancer Hospital,Chongqing,400030,China)

机构地区：[1]重庆大学附属肿瘤医院健康体检与肿瘤筛查中心,重庆400030 [2]重庆大学附属肿瘤医院泌尿肿瘤科,重庆400030

出　　处：《陆军军医大学学报》2023年第16期1741-1747,共7页Journal of Army Medical University

摘　　要：目的探讨巨噬细胞调控膀胱癌细胞上皮-间质转化(epithelial-mesenchymal transformation,EMT)发生的作用机制。方法收集2000-2015年在重庆大学附属肿瘤医院泌尿肿瘤科行根治性膀胱切除术的137例患者膀胱癌(bladder cancer,BC)组织切片,采用免疫组化(immunohistochemical,IHC)实验检测其CLDN10(claudin10)的表达,分析CLDN10的表达与患者临床资料的相关性。SiRNA干扰技术敲低RAW264.7细胞的TNF-α后,将鼠源膀胱癌细胞MB49与鼠源巨噬细胞RAW264.9共培养,运用Western blot实验检测MB49细胞内EMT相关蛋白的表达变化。采用免疫荧光实验检测MB49细胞内CLDN10与β-catenin蛋白的结合变化。用平板克隆形成实验检测MB49细胞与RAW264.7细胞内EMT相关蛋白的表达变化。结果在137例标本中,CLDN10高表达68例,CLDN10高表达与肿瘤分期(P=0.014)、淋巴结转移(P=0.005)呈正相关,与生存时间(P=0.048)、无复发生存时间(P=0.008)呈负相关。外源性TNF-α作用膀胱癌细胞后,免疫荧光结果显示CLDN10在膀胱癌细胞的表达增加,Western blot结果显示EMT相关蛋白Vimentin、Snail、Twist的表达上调(P<0.05);抑制巨噬细胞TNF-α后与膀胱癌细胞共培养,发现膀胱癌细胞内Vimentin、Snail、Twist的表达下调(P<0.05)。将RAW264.7细胞与MB49细胞共同培养后,MB49细胞内Vimentin、Snail、Twist的表达上调,E-cadherin表达下调(P<0.05)。结论巨噬细胞通过分泌TNF-α调控CLDN10/β-catenin信号通路促进膀胱癌细胞的增殖和EMT发生。Objective To explore the mechanism that macrophages promote epithelial-mesenchymal transformation(EMT)in bladder cancer cells.Methods A total of 137 tissues of bladder cancer(BC)were collected from Department of Urological Oncology of Chongqing University Cancer Hospital between 2000 and 2015.Immunohistochemical(IHC)assay was used to detect the expression of CLDN10 in 137 bladder cancer specimens,and statistical analysis was used to investigate the association in CLDN10 expression and clinical features.In mouse RA W264.7 macrophages and bladder cancer MB49 cells,siRNA technology was used to knockdown the expression of TNF-cu in RAW264.7 cells.Western blot assay was performed to detect the levels of related protein expression of EMT in MB49 ells,while immunofluorescence assay was used to detect the combination of CLDN10 andβ-catenin.P late clone formation assay was performed to detect the levels of related protein expression of EMT in MB49 cells and RAW264.7 cells.Results In 137 samples,the proportion of overexpression of CLDN10 was 49.6%(68/137).Overexpression of CLDNI0 was positively correlated with tumor(P=0.014)and lymphatic metastasis(P=0.005),and was negatively correlated with survival time(P=0.048)and recurrence-free survival(P=0.008).Treatment of exogenous TNF-cx led to accumulation of CLDNI0 in BC cell nucleus.The results of W estern botting showed the levels of EMT related proteins Vimentin,Snail and Twist were up-regulated(P<0.05).Adversely,repression of TNF-ax in macrophages resulted in decrease of the levels of EMT related proteins,Vimentin,Snail and Twist(P<0.05).The levels of expression of Vimentin,snail and Twist were increased while the level of E-cadherin was decreased(P<0.05)in MB49 cells after co-culturing of RAW264.7 cells and MB49 cells.Conclusion Macrophages can modulate BC cell proliferation and EMT progression by secreting TNF-c to regulate CLDN10/β-catenin mediated signaling pathway.

关 键 词：巨噬细胞 CLDN10 TNF-Α 膀胱癌 

分 类 号：R392.11[医药卫生—免疫学] R730.23[医药卫生—基础医学] R737.14