骨髓间充质干细胞来源外泌体抑制高糖诱导的腹膜间皮细胞EMT  

Bone marrow mesenchymal stem cell-derived exosome inhibits high glucose-induced EMT of peritoneal mesothelial cells

作　　者：赵俊丽 朱君君 詹秋楠 刘苗 ZHAO Junli;ZHU Junjun;ZHAN Qiunan;LIU Miao(Department of Nephrology,Zhoupu Hospital of Pudong New Area,Shanghai University of Medicine&Health Sciences Affiliated Zhoupu Hospital,Shanghai 201318,China)

机构地区：[1]上海市浦东新区周浦医院,上海健康医学院附属周浦医院,肾内科,上海201318

出　　处：《基础医学与临床》2024年第8期1149-1156,共8页Basic and Clinical Medicine

基　　金：上海市浦东新区科技发展基金(PKJ2021-Y34)。

摘　　要：目的探讨骨髓间充质干细胞外泌体(BMSCs-Exo)对高糖腹膜透析液(PDF)作用下人腹膜间皮细胞系(HMrSV5)发生上皮-间充质转化(EMT)的影响及其作用机制。方法采用透射电镜、纳米颗粒追踪分析仪和Western blot对BMSCs-Exo进行鉴定;将HMrSV5细胞进行分组:(1)对照组;(2)不同浓度PDF(1.5%、2.5%、4.25%)组;(3)siNLRP3组;(4)siNC组;(5)BMSCs-Exo组。Western blot检测EMT标志物及NLRP3炎性体通路蛋白的表达,RT-qPCR检测mRNA表达,ELISA检测细胞上清中炎性因子表达量。结果BMSCs-Exo呈双层膜结构的圆形囊泡,直径40~150 nm,外泌体特异标记物CD9、CD81、TSG101和Alix阳性;与对照组相比,PDF组细胞中α-SMA、vimentin、NLRP3、pro-caspase-1及pro-IL-1β表达明显上调,而E-cadherin表达下调(P<0.05);PDF组细胞上清中TGF-β1、IL-1β和IL-18表达量显著提高(P<0.05)。siNLRP3组细胞α-SMA表达下降,而E-cadherin表达明显上调。与4.25%PDF组比较,BMSCs-Exo组细胞E-cadherin表达上调,而vimentin、α-SMA、NLRP3、pro-caspase-1、pro-IL-1β蛋白表达下调(P<0.05);ELISA结果显示,BMSCs-Exo可下调4.25%PDF诱导的细胞上清IL-1β、IL-18、TGF-β1的水平(P<0.05)。结论高糖PDF通过激活NLRP3炎性体信号途径诱导腹膜间皮细胞EMT,BMSCs-Exo可通过抑制该通路改善腹膜间皮细胞EMT。Objective To investigate the effects of bone marrow mesenchymal stem cell-derived exosomes(BMSCs-Exo)on the regulation of epithelia-mesenchymal transition(EMT)in human peritoneal mesothelial cells(HPMCs)treated with glucose-based peritoneal dialysis fluid(PDF).Methods BMSCs-Exo were verified by transmission electron microscopy(TEM),nanoparticle tracking analyzer(NTA)and Western blot.Cultured HPMCs(HMrSV5)were divided into 5 groups:control group,high glucose-based PDF(1.5%,2.5%,and 4.25%)group,siNLRP3 group,siNC group and BMSCs-Exo treated group.Expression of E-cadherin,vimentin,α-smooth muscle actin(α-SMA)and NLRP3 inflammasome-related proteins were detected by Western blot.Real time RT-PCR was used to detected the expression ofα-SMA,E-cadherin and TGF-β1 mRNAs in HMrSV5 cells.The concentration of TGF-β1,IL-1βand IL-18 in the culture supernatant was determined by ELISA.Results The exosomes isolated were spherical and double-membrane vesicles with 40-150 nm in diameter,which expressed CD9,CD81,TSG101 and Alix protein.Our results showed that the level ofα-SMA and vimentin were significantly up-regulated and E-cadherin(epithelial marker)was significantly decreased in HMrSV5 cells treated with high glucose PDF compared with the normal HMrSV5 cells.The expression of NLRP3,pro-caspase-1 and pro-IL-1βwere also significantly up-regulated in HMrSV5 cells treated with high glucose PDF compared with the normal HMrSV5 cells.The level of TGF-β1,IL-1βand IL-18 in high glucose PDF treated HMrSV5 cells culture supernatant was up-regulated in a dose dependent manner.The protein level ofα-SMA was decreased and E-cadherin level was increased by an NLRP3 siRNA to inhibit the activation of NLRP3.Compared with 4.25%PDF treated cells,E-cadherin expression was up-regulated,while the expression ofα-SMA and vimentin were down-regulated in BMSCs-Exo treatment cells(P<0.05).Furthermore,the protein expression of NLRP3,pro-caspase-1 and pro-IL-1βin 4.25%PDF-treated HMrSV5 cells were significantly reduced by BMSCs-Exo.BMSCs-Exo al

关 键 词：腹膜间皮细胞 上皮-间充质转化(EMT) NLRP3 间充质干细胞外泌体 

分 类 号：R692.5[医药卫生—泌尿科学]