腹膜透析流出液中人腹膜间皮细胞的培养及转分化特征  

Epithelial-mesenchymal transition of human peritoneal mesothelial cells isolated from effluent fluid in dialysis

作　　者：周循[1] 凌光辉[1] 邹莎琳[1] 段绍斌[1] 肖力[1] 孙林[1] 刘伏友[1] 

机构地区：[1]中南大学湘雅二医院肾内科,湖南长沙410011

出　　处：《中国现代医学杂志》2010年第17期2634-2638,2642,共6页China Journal of Modern Medicine

基　　金：国家自然科学基金面上项目(No:30871169)

摘　　要：目的建立从腹膜透析(PD)流出液中分离、培养人腹膜间皮细胞(HPMC)的方法,并检测上皮黏附连接蛋白E-cadherin、紧密连接蛋白claudin-1及间充质细胞标志物α-平滑肌肌动蛋白(α-SMA)的表达水平。方法从25例持续不卧床腹膜透析(CAPD)患者PD流出液中分离HPMC,并在体外进行原代培养。根据开始透析的时间分为新开管组(10例)和透析半年以上组(15例)。采用倒置相差显微镜、免疫组织化学染色和扫描电镜对培养的细胞进行鉴定;采用实时定量PCR检测细胞E-cadherin、claudin-1及α-SMAmRNA表达。结果 22例患者(新开管组10例,半年以上组12例)PD流出液分离的细胞在体外培养成活,经鉴定均具有HPMC的特征。实时定量PCR结果显示,半年以上组较新开管组分离的HPMC细胞E-cadherin、claudin-1 mRNA表达降低,α-SMA mRNA表达增高(P<0.05),提示出现了间充质细胞转分化(EMT)。结论该研究成功建立了PD流出液分离培养原代HPMC的方法,并证实长期PD患者HPMC存在EMT倾向。该研究的发现为进一步研究腹膜纤维化(PF)和超滤衰竭(UFF)提供了新的方法与实验依据。ObjectiveTo establish a culture model of human peritoneal mesothelial cells（HPMCs） in vitro isolated from effluent fluid in dialysis,and to observe the expressions of E-cadherin,claudin-1 and α-SMA in HPMCs.MethodsHPMCs were isolated from effluent fluid of dialysis in 25 patients undergoing continuous ambulatory peritoneal dialysis（CAPD）.Ten patients were newly undergoing PD,while fifteen patients were undergoing PD for over six months.The isolated cells were identified by morphology and immunohistochemistry.The expressions of E-cadherin,claudin-1 and α-SMA were detected by real-time PCR.ResultsCells from 22 patients（10 newly undergoing PD,12 undergoing PD for over 6 months） survived.The confluent cells showed characteristics of HPMC by morphology,immunohistochemistry and electron microscopy.In cultured cells from patients undergoing PD for over 6 months,mRNA expressions of E-cadherin and claudin-1 were decreased while α-SMA increased,compared to new PD group（P 0.05）.It confirmed epithelial-mesenchymal transition（EMT） of HPMC during PD.ConclusionThe culture model of HPMC isolated from effluent fluid in dialysis has been successfully established and that EMT tendency of HPMC during PD has been revealed.It will provide the research basis of peritoneal fibrosis（PF） and ultrafiltration failure（UFF）.

关 键 词：腹膜透析 间皮细胞 细胞培养 上皮间充质转分化 

分 类 号：R459.5[医药卫生—治疗学]