高糖对小鼠足细胞向间充质细胞转分化的影响

Impact of high-glucose on epithelial-mesenchymal transition in mouse podocytes

目的通过观察高糖环境下小鼠足细胞表达NEPH1、desmin、TGF-β1和Smad7的变化,探讨高糖损伤足细胞的机制。方法将培养成熟的小鼠足细胞用RandA 1.0软件完全随机分为高糖1、2、3组(葡萄糖浓度分别为15、25和30mmol/L)和对照组(葡萄糖浓度5mmol/L),倒置显微镜下观察高糖干预前后足细胞的形态变化,采用RT—PCR和Western blot技术分别检测足细胞NEPH1、desmin、TGF-β1和Smad7的mRNA和蛋白表达变化。结果与对照组相比,高糖环境下培养48h后,足细胞形态发生不同程度变化。NEPH1和Smad7的mRNA表达较对照组显著减少(P<0.05或0.01),desmin和TGF-β1的mRNA表达较对照组增加(P<0.05或0.01)。其中高糖2组的NEPH1和Smad7的蛋白表达较对照组显著减少(均P<0.01),而desmin和TGF—β1的蛋白表达较对照组显著增加(P<0.01)。小鼠足细胞在高糖培养下NEPH1 mRNA和desmin mRNA的表达呈显著负相关(r=-0.993,P<0.01);NEPH1 mRNA和TGF—β1 mRNA的表达呈显著负相关(r=-0.989,P<0.01);TGF-β1 mRNA的表达和Smad7 mRNA的表达呈显著负相关(r=-0.996,P<0.01)。结论高糖可能通过激活TGF—β/Smad信号通路诱导小鼠足细胞发生表型转化。

Objective To investigate the effect of high-glucose on epithelial-mesenchymal transition in mouse podocytes. Methods Matured mouse podocytes were cultured and randomly divided into control group （5mmol/L D-glucose） and different high glucose groups （15, 25 and 30 mmol/L D-glucose, respectively） by means of RandA1.0 randomized software. Mor- phological changes of podocytes were examined with inverted microscope and expression changes of NEPH1, desmin, TGF- 15 1 and Smad7 mRNA and protein were detected by RT-PCR and Western blot, respectively. Results Matured podocytes incubated with high glucose were converted from arborized celrs into cobblestone appearance after 48h culture. Compared with the control group, the expression of NEPH1 and Smad7 mRNA in pedocytes cultured with different high glucose were significantly decreased （P〈0.05 or 0.01）; meanwhile, the expressions of desmin and TGF- 15 lmRNA in pedocytes cultured with different doses of high glu- cose were significantly increased in comparison with those of control group （P〈0.05 or 0.01）. The protein expressions of NEPH 1 and Smad7 were significantly decreased while those of desmin and TGF- [3 1 were significantly increased in 25mmol/L glucose group compared with control group （P〈0.01）. The expression of NEPH1 mRNA was negatively correlated with that of desmin mRNA （r=-0.993, P〈0.01） and TGF- 13 1mRNA（r=-0.989, P〈0.01）. The expression of TGF- 15 1 mRNA was negatively correlated with that of Smad7 mRNA in mouse podocyte exposed to high glucose （r=-0.996, P〈0.01）. Conclusion The phenotypic transformation of mouse podocytes can be possibly induced by high glucose via activating the TGF- 15/Smad signal pathway.