雷帕霉素对糖尿病肾病小鼠足细胞自噬的改善作用  

Rapamycin attenuates podocyte autophagy in mice with diabetic nephropathy

作　　者：肖堂利[1] 孙蕾[1] 管旭[1] 杨可[1] 何婷[1] 徐新丽[1] 黄云剑[1] 赵景宏[1] 

机构地区：[1]第三军医大学新桥医院肾内科,重庆400037

出　　处：《第三军医大学学报》2013年第23期2530-2535,共6页Journal of Third Military Medical University

基　　金：国家自然科学基金面上项目(81270290);重庆市自然科学基金重点项目(CSTC2013jjB10023)~~

摘　　要：目的探讨雷帕霉素(Rapamycin)对糖尿病肾病小鼠肾小球足细胞自噬和损伤的影响,以及对糖尿病肾病的改善作用,阐明糖尿病肾病可能的发生机制。方法健康雄性Balb/c小鼠48只,采用完全随机设计分组法分为柠檬酸对照组(n=12)、雷帕霉素对照组(n=12)、链脲佐菌素(streptozotocin,STZ)组(STZ组,n=12)和STZ+雷帕霉素组(n=12)。采用腹腔注射STZ制备糖尿病动物模型,成模后即给予小鼠腹腔注射雷帕霉素2 mg/(kg·48 h),雷帕霉素注射0、2、4、8、12周时分别测定各组小鼠血糖和体质量,12周时留取24 h尿标本,取肾脏称质量、检测24 h尿蛋白和肌酐,计算尿蛋白/肌酐值(UACR)和肾质量/体质量值(KW/BW)。PAS染色观察肾组织病理改变,透射电镜观察肾小球足细胞自噬体大小和数量、细胞器损伤情况,Western blot检测足细胞自噬标志物LC3蛋白的表达变化,nephrin免疫荧光染色观察足细胞数量。结果与对照组相比,STZ组小鼠血糖、UACR、KW/BW明显增高(P<0.05),PAS染色发现肾小球基膜增厚,系膜基质增多,电镜提示足细胞足突增宽、融合,足细胞和自噬体数量减少,自噬活化标志物LC3-Ⅱ/LC3-Ⅰ值明显下降(P<0.01)。与STZ组比,雷帕霉素处理后STZ小鼠肾小球病理变化有所改善,足细胞和自噬体数量增多,细胞器损伤减轻,LC3-Ⅱ/LC3-Ⅰ值上调,UACR和KW/BW下降(P<0.05),但血糖无统计学差异。结论糖尿病小鼠腹腔注射雷帕霉素可以减轻肾小球病理改变和足细胞损伤,其作用可能与促进足细胞自噬有关,而维持足细胞自噬平衡可能是延缓糖尿病肾病进展的有效措施。Objective To determine the effect of rapamycin on podocyte autophagy and impairment in diabetic mice, and its ameliorative role in diabetic nephropathy. Methods A total of 48 healthy male Balb/c mice were randomly divided into 4 groups, that is, control group treated with citric acid （Control, n = 12 ）, control mice treated with rapamycin （Control ＋ R group, n = 12）, STZ group （n = 12 ）, streptozotocin （STZ） mice intraperitoneal injected with rapamycin group （STZ ＋ R group, n = 12）. Diabetic animal models were induced by intraperitoneal injection of STZ into the STZ mice. After the model establishment, mice were given rapamycin at 2 rag/（ kg·48 h） through intraperitoneal injection. Serum glucose and body weight of each group were measured in 0, 2, 4, 8 and 12 weeks after treatment. At the 12th week, 24-hour urinary specimens wet~ collected to determine albumin and creatinine levels for calculation of urinary albumin/creatinine ratio （UACR）, and the kidneys were taken and weighted to calculate kidney weight/body weight ratio （KW/BW）. Renal pathological changes were observed by PAS staining. Size and number of podocyte autophagy and organ- elles injury were investigated by transmission electron microscopy （TEM）. LC3 protein, a podocyte autophagy marker, were detected by Western blotting. Podocyte count was carried through nephrin immunofluoreseence staining. Results Compared with control group, blood glucose, UACR and KW/BW in STZ group were increased significantly （P 〈 0. 05 ）. PAS staining showed thickened glomerular basement membrane and increased mesangial matrix. TEM indicated podocyte foot processes were broadened and fused, and the number of podoeytes and autophagosomes were decreased. LC3-Ⅱ/LC3-Ⅰ protein ratio was significantly decreased （P 〈0.01 ）. Compared with STZ group, the mice from STZ ＋ R group showed ameliorated glomerular pathological changes and increase in number of podocytes and autophagosomes after rapamycin treatment, alleviated organe

关 键 词：雷帕霉素 自噬 足细胞 糖尿病肾病 

分 类 号：R587.2[医药卫生—内分泌] R692[医药卫生—内科学]