氧化应激介导糖尿病大鼠肾小管上皮细胞转分化及普罗布考的干预作用

目的 研究氧化应激在糖尿病肾病（DN）大鼠肾小管上皮细胞转分化中的作用，探讨抗氧化剂普罗布考对大鼠DN的肾脏保护作用。 方法 30只雄性SD大鼠随机分为正常对照组、DN组和普罗布考干预组（1％普罗布考饮食），每组10只。分别于第3周、第8周及第12周检测24 h尿蛋白（UTP）；12周末检测各组大鼠血糖、血脂（胆固醇、三酰甘油）、Scr、肌酐清除率（Ccr）、肾脏组织匀浆液丙二醛（MDA）含量及谷胱甘肽过氧化物酶（GSH-Px）活性。肾组织病理切片行 HE和Masson染色；采用免疫组化和Western印迹检测肾组织核转录因子Sp1、α平滑肌肌动蛋白（α-SMA）及E钙黏蛋白（E-cadherin）表达。 结果 与正常对照组比较，DN组血糖、Scr、肾组织匀浆MDA和24 h UTP水平显著增高（均P < 0.01），Ccr显著降低（P < 0.01）；肾组织肾小管损伤分数、α-SMA和 Sp1蛋白表达水平明显增高（均P < 0.01）；肾组织E-cadherin蛋白表达明显下调。肾组织MDA含量分别与α-SMA及Sp1蛋白表达呈正相关（r ＝ 0.896，P < 0.01；r ＝ 0.862，P < 0.01），与E-cadherin蛋白表达呈负相关（r = -0.673， P < 0.01）。普罗布考干预组Scr、24 h UTP、肾组织MDA、肾小管损伤分数及肾组织α-SMA、 Sp1蛋白表达水平较DN组均明显降低（均P < 0.01）；Ccr和肾组织E-cadherin蛋白表达水平较DN组均明显增加（均P < 0.01）。 结论 氧化应激在DN大鼠肾小管上皮细胞转分化中起重要作用。普罗布考可能通过抗氧化、下调肾组织Sp1蛋白表达及抑制肾小管上皮细胞转分化延缓DN大鼠肾脏病变进展。

Epithelial-to-mesenchymal transdifferentiation of renal tubular epithelial cells mediated by oxidative stress and intervention effect of probucol in diabetic nephropathy rats

Objective To explore the role of oxidative stress in the epithelial-tomesenchymal transition (EMT) of tubular epithelial cells and the protective effect of probucol in rat model with diabetic nephropathy (DN). Methods Thirty SD rats were randomly divided into normal control group, DN group, probucol treatment group (supplemented 1% probucol dietary). Twentyfour hours urinary protein excretion (UTP) was measured at the 3rd, the 8th and the 12th week respectively. The biochemical indicators including blood glucose (BG), lipids triglyceride (TG), total cholesterol (TC)], low-density lipoprotein (LDL), serum creatinine (Scr), creatinine clearance rate (Ccr),kidney tissue malondialdehyde (MDA) level and glutathione peroxidase (GSH-Px) activity were assessed at the end of the 12th week in all groups. The renal pathological changes were evaluated by hematoxylin & eosin (HE) and Masson staining. The protein expression of specificity protein 1 (Sp1), α-smooth muscle actin (α-SMA) and E-cadherin was also detected and analyzed by immunohistochemistry and Western blotting. Results Compared with the normal control group,the BG, TC, LDL, Scr, 24 h UTP and MDA level of renal tissue increased significantly and the Ccr reduced in the rats of DN group (all P＜0.01). The pathological scores and the expression of Sp1 and α-SMA in renal tissue were higher in the DN animals than that in the other animals (all P＜0.01), the expression of E-cadherin downregulated significantly in the DN animals (P＜0.01). The MDA level of renal tissue was positively correlated to the expression of α-SMA and Sp1 protein in DN group (r=0.896, P＜0.01; r=0.862, P＜0.01, respectively), and negatively correlated to the expression of E-cadherin protein (r=-0.673, P＜0.01). In the diabetic animals treated with probucol, the Scr, 24 h UTP, pathological scores, MDA content,expression of Sp1 and α-SMA in renal tissue were lower than those in the diabetic animals (all P＜0.01). The Ccr and the expression of E-cadherin upregulated obviously (all P＜0.01). Conclusion Oxidative stress plays an important role in the EMT process of tubular epithelial cells. Probucol can slow down renal disease progression in DN rats through anti-oxidant, downregulating the expression of Sp1 protein and inhibiting the renal tubular EMT.