含糖腹膜透析液诱导人腹膜间皮细胞自噬机理研究  

Mechanism of Autophagy Induced by Peritoneal Dialysate Solution Containing Glucose in HMrSV5 Cells

作　　者：段仁全[1] 丁佑铭[1] 汪长华 DUAN Renquan;DING Youming;WANG Changhua(Dept. of Hepatobiliary ＆Laparoscopic Surgery, Renmin Hospital of Wuhan University, Wuhan 430060, China;Dept. of Pathophysiology, School of Basic Medical Sciences, Wuhan University, Wuhan 430071 , China)

机构地区：[1]武汉大学人民医院肝胆腔镜外科,湖北武汉430060 [2]武汉大学基础医学院病理生理学教研室,湖北武汉430071

出　　处：《武汉大学学报（医学版）》2017年第6期968-972,977,共6页Medical Journal of Wuhan University

基　　金：国家科技支撑计划资助项目(编号:2012BAD31B00)

摘　　要：目的:研究含糖腹膜透析液对人腹膜间皮细胞株(HMrSV5)生长影响,探讨引发的线粒体氧自由基的产生、细胞氧自由基增加,以及炎症和自噬发生的可能机理。方法:用1.5%、2.5%和4.25%浓度的含糖低钙腹膜透析液及阳性对照鱼藤酮(Rotenone)刺激人腹膜间皮细胞48h,用流式细胞术检测人腹膜间皮细胞线粒体氧自由基产生和细胞内氧自由基变化、蛋白印迹检测NLRP3和LC3表达变化及透射电镜观察细胞器变化。结果:含糖低钙腹膜透析液在不同的浓度和不同的作用时间情况下,对HMrSV5线粒体及细胞内活性有不同的影响。随糖浓度的增加线粒体氧自由基含量明显增加。随糖浓度升高和时间延长,胞内氧自由基也明显增加;含糖低钙腹膜透析液诱导NLRP3与LC3的表达,并呈糖浓度与时间依赖性;透射电镜观察到细胞线粒体出现空泡及自噬小体的出现。结论:高糖低钙腹膜透析液可刺激HMrSV5细胞的线粒体氧自由基产生并泄露到胞质中,引发细胞炎症自噬的产生。这为基础研究及临床防治腹膜纤维化提供了一条新途径。Objective： To study the effect of of peritoneal dialysate solution（PDS） containing different concentrations of glucose on peritoneal mesothelium cell lines （HMrSVS） growth, cellular reac- tive oxygen species（ROS） production and leakage from mitochondrial, and the occurrence of in- flammation and autophagy. Methods： Peritoneal mesothelium cells was stimulated with low-calci- um peritoneal dialysate solution containing 1.5%, 2.5% or 4.25% concentrations of glucose and rotenone （a positive control） for 48 hours. Flow cytometry was used to detect the level of mito- chondrial reactive oxygen species in human peritoneal mesothelium cell and the change of intracel- lular reactive oxygen species; Western blot was used to measure the expressions of NLRP3 and LC3; transmission electron microscope was used to observe the organelles change. Results： Low calcium peritoneal dialysate solution containing glucose had significant effects on mitochondrialand intracellular activities in HMrSV5 ceils. With the increase of glucose content, mitochondrial ROS was significantly increased, but it was decreased at high concentration. The level of cyto- plasm ROS increased with the increase of glucose concentration and stimulation time. NLRP3 and LC3 were activated by low-calcium peritoneal dialysate solution containing glucose in a glucose concentration-dependent and time-dependent manner. In addition, mitochondrial cavitation and autophagosome appearance were observed by transmission electron microscopy. Conclusion. Low- calcium peritoneal dialysate solution containing high glucose can stimulate mitochondrial ROS production and leakage into cytoplasm, induce inflammation and cell autophagy. These results provide a feasible way for peritoneal fibrosis in basic research and clinical treatment.

关 键 词：腹膜透析液 腹膜间皮细胞 氧自由基 炎症 自噬 

分 类 号：R692.5[医药卫生—泌尿科学]