脂肪间充质干细胞外泌体对人子宫内膜异位症细胞纤维化形成的影响  

Effect of adipose derived stem cell exosomes on fibrogenesis of human endometriosis cells

作　　者：李冰冰 丁美琪 孙新慧 王荣 LI Bingbing;DING Meiqi;SUN Xinhui;WANG Rong(Jining Medical College,Jining 272000,China;Beijing University of Chinese Medicine,Beijing 100029,China)

机构地区：[1]济宁医学院,山东济宁272000 [2]北京中医药大学,北京100029

出　　处：《药物评价研究》2024年第9期2032-2040,共9页Drug Evaluation Research

基　　金：国家自然科学基金委员会项目(82104920);山东省自然科学基金委员会项目(ZR2021QH129);山东省中医药重点学科方剂学(鲁卫中医药科教字[2022]4号)。

摘　　要：目的探讨脂肪间充质干细胞外泌体(ADSCs-Exos)对子宫内膜异位症细胞(12Z)增殖以及纤维化相关因子表达的影响。方法采用超速离心法提取ADSCs-Exos,并通过透射电镜(TEM)、纳米颗粒跟踪分析(NTA)、Western blotting鉴定;采用PKH26标记外泌体法观察12Z细胞对外泌体的吞噬内化情况;采用CCK-8法检测ADSCs-Exos(12.5、25.0、50.0μg·mL^(-1))对12Z细胞增殖的影响;将12Z细胞分为:对照组、模型组、ADSCs-Exos(12.5、25.0、50.0μg·mL^(-1))组,除对照组外,其余各组采用TGF-β1(5 ng·mL^(-1))诱导细胞纤维化,给药24 h后采用实时荧光定量PCR(qRT-PCR)检测纤维化因子基因mRNA的表达。结果提取的ADSCs-Exos呈现圆形颗粒,清晰可见;总萃取物的平均粒径为122.6 nm,浓度为1.8×10^(6)颗粒·mL^(-1);CD9、CD63和TSG101在ADSCs-Exos均有表达,且Calnexin蛋白未表达,符合外泌体的特征。12Z细胞可以成功将ADSCs-Exos吞噬内化;CCK-8检测结果显示,与对照组相比,采用25μg·mL^(-1)ADSCs-Exos干预24 h后12Z细胞的相对存活率显著降低(P<0.01)。qRT-PCR结果显示,与对照组相比,模型组细胞的CTGF、COL1A1、α-SMA mRNA表达水平显著升高(P<0.001);与模型组比较,12.5、25.0μg·mL^(-1)ADSCs-Exos组CTGF、COL1A1、α-SMA mRNA表达水平降低(P<0.05、0.01、0.001);50.0μg·mL^(-1)ADSCs-Exos组α-SMA mRNA表达水平明显降低(P<0.001)。结论ADSCs-Exos可被12Z细胞所吞噬,抑制12Z细胞的增殖,并且降低细胞纤维化相关因子基因的表达水平。Objective To investigate the effects of adipose-derived stem cell-derived exosomes(ADSCs-Exos)on the proliferation and fibrosis-related factor expression of endometriosis cells(12Z).Methods The ADSCs-Exos were extracted by ultracentrifugation,identified by transmission electron microscopy,nanoparticle tracking analysis(NTA),and Western blotting.The uptake of exosomes by 12Z cells was observed by PKH26 labeling exosome method.The effects of ADSCs-Exos(12.5,25.0,50.0μg·mL^(-1))on the proliferation of 12Z cells were detected by CCK-8 assay.The 12Z cells were divided into four groups:control group,model group,and ADSCs-Exos(12.5,25.0,50.0μg·mL^(-1))group.Except for the control group,the other groups were induced to fibrosis by TGF-β1(5 ng·mL^(-1))for 24 h,and the expression of fibrosis-related gene mRNA was detected by real-time quantitative PCR(qRT-PCR)after drug treatment.Results The ADSCs-Exos extracted showed round particles that were clearly visible.The average particle size of the total extract was 122.6 nm,and the concentration was 1.8×10^(6)particles·mL^(-1).CD9,CD63,and TSG101 were expressed in ADSCs-Exos,and Calnexin protein was not expressed,consistent with the characteristics of exosomes.12Z cells successfully internalized and phagocytosed ADSCs-Exos.The CCK-8 assay results showed that compared with the control group,the relative survival rate of 12Z cells was significantly lower after 24 h of treatment with 25.0μg·mL^(-1) ADSCs-Exos(P<0.01).The qRT-qPCR results showed that compared with the control group,the expression levels of CTGF,COLIAl,andα-SMA mRNA in the model group were significantly incresed(P<0.001).Compared with the model group,the expression levels of CTGF,COLIAl,andα-SMA mRNA in the 12.5,and 25.0μg·mL^(-1) ADSCs-Exos groups were lower(P<0.05,0.01,0.001),and the expression level ofα-SMA mRNA in the 50.0μg·mL^(-1) ADSCs-Exos group was significantly lower(P<0.001).Conclusion Adipose mesenchymal stem cell exosomes can be swallowed by endometriosis cells,which can further inhibit

关 键 词：脂肪间充质干细胞 外泌体 子宫内膜异位症 纤维化 吞噬 

分 类 号：R965[医药卫生—药理学]