黄芪多糖对高糖腹透液诱导HMrSV5细胞凋亡的影响  

Effect of Astragalus Polysaccharide on Apoptosis of HMrSV5 Induced by Peritoneal Dialysis Solution

作　　者：赵君谊 单云 朱晓琳 史俊[1] 俞曼殊 李正红[1] 盛梅笑[1] ZHAO Junyi;SHAN Yun;ZHU Xiaolin;SHI Jun;YUN Manshu;LI Zhenghong;SHENG Meixiao(The Affiliated Hospital of Nanjing University of Chinese Medicine,Nanjing 210029,Jiangsu,China)

机构地区：[1]南京中医药大学附属医院肾内科,江苏南京210029

出　　处：《中华中医药学刊》2020年第10期113-117,I0032,I0033,共7页Chinese Archives of Traditional Chinese Medicine

基　　金：国家自然科学基金(81774253);江苏省自然科学基金(BK20171514);江苏省中医药领军人才项目(SLJ0205);江苏省中医院高峰学术人才项目(y2018rc08)。

摘　　要：目的观察黄芪多糖(Astragalus polysaccharide,APS)对高糖腹透液(peritoneal dialysis solution,PDS)诱导人腹膜间皮细胞(HMrSV5)凋亡的影响及其抗凋亡机制。方法①体外培养HMrSV5细胞株,CCK-8法检测不同浓度PDS(1.5%、2.5%、3.0%、3.5%、4.25%PDS)及不同浓度APS(50、75、100、200、300、400、500、600、800μg·mL^-1)对细胞活力的影响,根据CCK-8检测结果筛选用于后续实验PDS及APS的合适浓度;②将实验分为4组:空白组、APS(100μg·mL^-1)组、PDS(3.0%)组、PDS+APS(3.0%+100μg·mL^-1)组,相差显微镜观察细胞形态变化,通过JC-1法测定线粒体膜电位的去极化;TUNEL技术检测各组细胞凋亡情况;③Western Blot检测Bcl-2、Bax、Caspase-3、Cleaved Caspase-3蛋白的表达。结果①不同浓度PDS作用下HMrSV5细胞活力均下降,呈浓度依赖(P<0.01);25~100μg·mL^-1的APS能提高PDS对HMrSV5细胞的抑制作用,且100μg·mL^-1 APS组最显著(P<0.01)。②与空白组相比,PDS模型组细胞形态变小、拉长,细胞内出现空泡样变,线粒体膜电位下降,细胞凋亡率增加;与PDS模型组比较,PDS+APS组细胞形态有所恢复,细胞内空泡样变减轻,线粒体膜电位水平升高,细胞凋亡率降低。③与空白组相比,PDS模型组Bax、Cleaved Caspase-3蛋白表达显著增加,Bcl-2蛋白表达降低(P<0.05);与PDS模型组相比,PDS+APS组Bax、Cleaved Caspase-3蛋白表达降低,Bcl-2蛋白表达升高(P<0.05)。结论APS能改善高糖腹透液对HMrSV5细胞诱导的凋亡,其机制可能是影响与线粒体凋亡相关的Bcl-2、Bax和Cleaved Caspase-3蛋白表达。Objective To evaluate the effects of Astragalus polysaccharide(APS)on high glucose peritoneal dialysis solution(PDS)inducing apoptosis of human peritoneal mesothelial cell(HMrSV5)and its possible anti-apoptosis mechanism.Methods HMrSV5 cell line was cultivated in vitro.CCK-8 test was used to detect cell viability after different concentrations of glucose PDS(1.5%,2.5%,3.0%,3.5%,4.25%)and different concentrations of APS(50,75,100,200,300,400,500,600,800μg·mL-1)so as to determine proper concentrations of glucose aseptic fluid and APS applied in the experiment.Cells were divided into four groups including control group,APS(100μg·mL^-1)group,PDS(3.0%)model group and PDS+APS(3.0%+100μg·mL^-1)group.The phase contrast microscope was used to observe cellular morphological change.JC-1 test was used to determine the depolarization of mitochondrial membrane potential(MMP).TUNEL technique was used for cell apoptosis and Western Blot for protein expressions of Bcl-2,Bax,Caspase-3 and Cleaved Caspase-3.Results①Compared with those of the control group,PDS at different concentrations lowered HMrSV5 viability dose-dependently(P<0.01).APS at 25~100μg·mL^-1 significantly relieved PDS’s inhibitory effects on HMrSV5,with concentration of 100μg·mL^-1 demonstrating the most statistical significance(P<0.01).②Compared with that of the control group,cells in PDS model group turned smaller and stretched with vacuoloid changes.It demonstrated decreased MMP and increased apoptosis rate,while cells in PDS+APS group displayed certain cellular morphological recovery with few vacuoloid changes,and enjoyed increased MMP and decreased apoptosis rate.③Compared with those of the control group,expressions of Bax and Cleaved Caspase-3 in APS group were significantly increased while Bcl-2 obviously decreased(P<0.05).Compared with those of the model group,expressions of Bax and Cleaved Caspase-3 in PDS+APS group were significantly decreased while Bcl-2 obviously increased(P<0.05).Conclusion APS can relieve high glucose PDS inducing cel

关 键 词：黄芪多糖 高糖腹透液 凋亡 人腹膜间皮细胞 线粒体 

分 类 号：R285.5[医药卫生—中药学]