还原型谷胱甘肽对糖尿病大鼠肾组织糖基化及转化生长因子β_1表达的影响

Effects of reduced glutathione on nonenzymatic glycosylation and expression of TGF-β_1 in the kidney of diabetic rats

目的观察还原型谷胱甘肽及盐酸氨基胍对链脲佐菌素诱导的糖尿病大鼠肾组织非酶糖化及转化生长因子β1(TGF-β1)表达的影响,以发现更为有效的防治糖尿病肾病及减少终末期肾病发病率的方法。方法应用链脲佐菌素建立大鼠糖尿病模型,随机分为正常对照组、糖尿病对照组(DM组)、氨基胍治疗组(DM+AG组)、还原型谷胱甘肽治疗组(DM+GLUT组)、氨基胍+还原型谷胱甘肽治疗组(DM+AG+GLUT组)。第8周留取双侧肾脏,应用反转录聚合酶链反应(RT-PCR)和免疫组织化学等方法研究5组大鼠肾脏TGF-β1mRNA及蛋白质表达、肾皮质糖基化终末产物(AGEs)含量、24 h尿白蛋白、肾组织学形态,测定肾重/体重比值、血糖、糖化血红蛋白、血肌酐水平,计算肌酐清除率。结果(1)与正常对照组相比,各组糖尿病大鼠TGF-β1mRNA表达明显上调(P<0.01);肾皮质匀浆及血清TGF-β1蛋白量均明显高于正常对照组(P<0.01);肾皮质匀浆脂相、蛋白质相荧光值即AGEs含量均明显增高(P<0.01);血糖、糖化血红蛋白、尿白蛋白排泄量、内生肌酐清除率及肾重/体重比值均明显增高(P<0.01);(2)与DM组相比,DM+AG组、DM+GLUT组及DM+AG+GLUT 3组TGF-β1mRNA表达增加被明显逆转(P<0.05或P<0.01);肾皮质匀浆及血清TGF-β1蛋白量均明显降低(P<0.01或P<0.05);AGEs含量均明显降低(P<0.01或P<0.05);尿白蛋白排泄量、内生肌酐清除率及肾重/体重均明显降低(P<0.01);(3)PAS、PAM、Masson染色显示:DM组肾小球可见轻度系膜增殖,未见系膜细胞增生、局灶节段性硬化及肾小球基底膜(GBM)增厚;正常对照组、DM+AG组、DM+GLUT组、DM+AG+GLUT组肾小球无明显异常。结论还原型谷胱甘肽及盐酸氨基胍可以减轻糖尿病大鼠肾组织的非酶糖化过程,并减轻肾小球肥大及肾肥大,延缓其肾病发展,对糖尿病肾脏具有保护作用,二者对肾脏的保护作用至少部分与抑制TGF-β1在肾脏的过度表达有关。

Objective To investigate the effects of reduced glutathione （ GLUT ） and aminoguanidine （ AG ） on nonenzymatic glycosylafion and expression of transforming growth factor - β1 （ TGF - β1 ） in the kidneys of diabetic rats. Methods Male wister diabetic rats were induced by intraperitoneal injection of streptosotocin （ 60 mg/kg）. They were randomly divided into five groups： control group, DM group, DM ＋ AG group, DM ＋ GLUT group and DM ＋ AG ＋ GLUT group. After eight weeks the kidneys of each rats were taken. The mRNA and protein expression of TGF - β1 was detected by quantitative RT - PCR and enzyme-linked immunesorbont assay. Advanced glycation end products（ AGEs） of renal cortical tissue, urinary albumin excretion, the renal histology were detected by immunohistochemistry staining. Kideny weight/body weight ratio, blood glucose, glycosylated hemoglobin A1 c, blood creatinine levels and creatin/ne clearance rate were tested. Results （ 1 ） Compared with those in the kidneys of control group,the expression of TGF - β1 in the diabetic rats is significantly upregnlated （ P 〈0. 01 ） ,the fluorescence of renal cortical tissue as well as the TGF - β1 protein in both renal cortical tissue and serum is significantly increased （ P 〈 0. 01 ）, blood glucose, hemoglobin A1c, urinary albumin excretion, creatinine clearance rate, kidney weight/body weight ratio are markedly increased （ P 〈 0.01 ）. （ 2 ） Compared with those in diabetic group, among DM ＋ AG group, DM ＋ GLUT group and DM ＋ AG ＋ GLUT group there is a significant reversion in the expression of TGF - β1 （ P 〈0. 05 ,P 〈0. 01 ）. The TGF - β1 protein in both renal cortical tissue and serum is significantly decreased（ P 〈0. 01 ） ,AGEs in renal cortical tssue is also decreased signifieanfly （ P 〈 0. 01, P 〈 0. 05 ） ,urinary albumin excretion,creatinine clearance rate, kidney weight/body weight ratio are decreased significantly（P〈0.01 ）. （3） PAS,PAM,Masson staining display light degree mesangial proliferation,no mesangial cells proliferation,focal segmental glomerular sclerosis and GBM incrassation in DM group. There is no obvious glomerular abnormality in the control group, DM ＋ AG group, DM ＋ GLUT group and DM ＋ AG ＋ GLUT group. Conclusions Reduced glutathione and aminoguanidine can decrease the expression of TGF - β1 in the kidney tissue of diabetic rats induced by streptozotocin and prevent progression of diabetic nephropathy by suppressing overproduction of AGEs.