论文部分内容阅读
[目的]探讨高糖对肾小管上皮细胞转分化影响的分子机制。[方法]体外培养的大鼠近端肾小管上皮细胞(NRK52E细胞)分为4组:①甘露醇组;②高糖组;③高糖+TGF-β1中和抗体组;④高糖+IgG1对照组。ELISA检测细胞培养上清中TGF-β1的浓度,细胞免疫化学方法检测p-Smad2/3核表达水平,RT-PCR和Western blot方法分别检测α-SMA和E-cadherin mRNA和蛋白的表达。[结果]高糖以时间依赖方式上调α-SMA和下调E-cadherin mRNA表达。与刺激前和甘露醇比较,高糖可以刺激NRK52E细胞合成和分泌内源性TGF-β1,上调p-Smad2/3核表达水平(P﹤0.01)。TGF-β1中和抗体能抑制高糖介导的p-Smad2/3核表达,下调高糖介导的α-SMA蛋白表达,及上调E-cadherin蛋白表达(P﹤0.01)。[结论]高糖通过TGF-β/Smad信号通路介导肾小管上皮细胞转分化。
[Objective] To investigate the molecular mechanism of high glucose on the transdifferentiation of renal tubular epithelial cells. [Methods] Rat proximal renal tubular epithelial cells (NRK52E cells) cultured in vitro were divided into 4 groups: ① mannitol group; ② high glucose group; ③ high glucose + TGF-β1 neutralizing antibody group; ④ high glucose + IgG1 Control group. The concentration of TGF-β1 in the cell culture supernatant was detected by ELISA. The nuclear expression of p-Smad2 / 3 was detected by immunocytochemistry. The mRNA and protein expressions of α-SMA and E-cadherin were detected by RT-PCR and Western blot respectively. [Result] High glucose up-regulated α-SMA and down-regulated E-cadherin mRNA expression in a time-dependent manner. Compared with pre-stimulation and mannitol, high glucose could stimulate NRK52E cells to synthesize and secrete endogenous TGF-β1 and up-regulate the expression of p-Smad2 / 3 (P <0.01). The neutralizing antibody of TGF-β1 could inhibit the expression of p-Smad2 / 3 induced by high glucose, the expression of α-SMA induced by high glucose and the expression of E-cadherin (P <0.01). [Conclusion] High glucose mediates renal tubular epithelial cell transdifferentiation through TGF-β / Smad signaling pathway.