Wnt/β-catenin信号通路在高糖诱导足细胞转分化中的作用机制研究

Effect Mechanism of Wnt/β-catenin Signaling Pathway in Podocyte Epithelial-mesenchymal Transition Induced by High Glucose

目的探讨Wnt/β-catenin信号通路在高糖诱导的足细胞转分化中的作用机制。方法收集原代培养的大鼠足细胞,设置正常糖组(D-葡萄糖5 mmol/L)、高糖组(D-葡萄糖15 mmol/L、30 mmol/L)、高渗对照组(甘露醇25 mmol/L+D-葡萄糖5 mmol/L)。观察细胞形态并采用间接免疫荧光法检测nephrin和Wt-1的表达;免疫印迹半定量检测nephrin、α-SMA、活化β-catenin及Wnt-1的表达。结果 nephrin以颗粒状及线状表达于足细胞胞膜、胞质及胞核周围,Wt-1表达于胞核。48 h后,正常糖组nephrin相对表达量为(0.967±0.065),15 mmol/L高糖组为(0.771±0.068),30 mmol/L高糖组为(0.112±0.042),高渗对照组为(1.096±0.104),差异有统计学意义(F=106.848,P=0.000);正常糖组α-SMA相对表达量为(0.360±0.023),15 mmol/L高糖组为(0.430±0.008),30 mmol/L高糖组为(0.524±0.040),高渗对照组为(0.320±0.030),差异有统计学意义(F=18.149,P=0.001)。高糖作用12 h后,足细胞Wnt-1及活化β-catenin表达开始升高,24 h时达到高峰。DKK1有助于增加高糖诱导的nephrin的表达,降低α-SMA的表达。结论高糖可诱导足细胞发生表型转化,而Wnt/β-catenin信号通路可能参与了高糖诱导的足细胞表型转化。

Objective To investigate the effect mechanism of Wnt/β - catenin signaling pathway in podocyte epithelial-mesenchymal transition （EMT） induced by high glucose. Methods The glomerular podocytes of rats were randomly dividedinto three groups： the normal glucose group （5 mmo/L） , high glucose group （15 mmo/L, 30 mmo/L） and mannitolgroup. Morphology method were used to observe the characteristic of the glomerular podocytes. The expression of Wt -1 and neph-rin were detected by indirect immunofluorescence technique,and the expression of nephrin,α - SMA,active β - catenin andWnt - 1 were detected by immunoblotting. Results The expression of nephrin and Wt - 1 were observed in the glomerular podo-cytes by the microscope. The nephrin expression level of high glucose group was significantly lower than that of normal glucosegroup and mannitol group〔5 mmo/L ： （0. 967 ±0. 065） ; mannitol ： （1. 096 ±0. 104） ; 15 mmo/L ： （0. 771 ±0. 068 ） ; 30mmol/L： （0. 112 ±0. 042） ; F = 106. 848,P =0. 000〕,the α - SMA expression level of high glucose group was significantlyhigher than that of normal glucose group and mannitol group [5 mmo/L： （0. 360 ±0. 023） ; mannitol： （0. 320 ±0. 030） ; 15mmo/L： （0. 430 ± 0. 008） ; 30 mmol/L： （0. 524 ± 0. 040） ; F = 18. 149 , P = 0. 001 ] . The Wnt - 1 and active β - catenin expression level of high glucose group increased at 12 hours, and peaked at 24 hours （P =0. 001 ） . The DKK1 may help to in-crease nephrin expression level in podocytes induced by high glucose,and may help to decrease α- SMA expression level inpodocytes induced by high glucose. Conclusion Phenotypic changes of podocytes can be induced by high glucose,and Wnt/ β -catenin signaling pathway may involve in this process.