太灵丹含药血清调控PI3K/AKT/GSK3β通路减轻高糖诱导的足细胞转分化的机制研究

Tailing Pills-containing Serum Inhibits Podocyte Hyperglycemia-induced Epithelial Mesenchymal Transition Via PI3K-AKT-GSK3βPathway

目的:观察太灵丹(TLD)含药血清对高糖诱导的足细胞转分化及PI3K/AKT/GSK3β通路的影响及分子机制。方法:将小鼠肾脏足细胞MPC5分为空白对照组、高糖刺激组(模型组)、太灵丹血清组(高、中、低浓度)。CCK-8检测足细胞的活力,流式细胞检测足细胞的凋亡,免疫组化检测desmin、FSP-1的蛋白表达,Western blot和QPCR法检测足细胞podocin、CD2-AP、FSP-1、PI3K、Akt、p-Akt、GSK3β蛋白及mRNA表达。结果:与正常组相比,模型组足细胞活力下降(P<0.01),凋亡增加(P<0.01),免疫组化结果显示,desmin及FSP-1的蛋白表达较正常组升高(P<0.05),podocin、CD2-AP、PI3K、Akt、p-Akt的蛋白及mRNA表达明显降低(P<0.05),而desmin、FSP-1、GSK3β的蛋白及mRNA表达明显升高(P<0.05)。与模型组相比,太灵丹含药血清各组足细胞活力显著升高(P<0.01),凋亡减少(P<0.01),podocin、CD2-AP、PI3K、Akt、p-Akt的蛋白及mRNA表达均明显上调(P<0.05),desmin、FSP-1、GSK3β的蛋白及mRNA表达均明显下调(P<0.05)。结论:太灵丹含药血清更能够减轻高糖诱导的足细胞转分化,其部分作用机制是通过调控PI3K/AKT/GSK3β通路实现的。

Objective:To observe the effects of drug-containing serum of Tailing Pills on glomerular podocyte epithelial-mesenchymal transition(EMT)and PI3K/AKT/GSK3βpathway under high glucose conditions,and to explore the protection mechanism for podocyte injury.Methods:Cultured differentiated mouse podocytes were treated with a drug-containing serum of Tailing Pills.After treating 48 hours,podocytes viability was determined by CCK-8 assay,the markers of podocyte EMT and the molecules related to PI3K/AKT/GSK3βpathway were analyzed by real-time PCR and Western Blot.Results:Compared with normal group,the model group’s gene and protein expressions of Podocin,CD2-AP,PI3K,Akt and p-Akt of podocytes induced by hyperglycemia in model group were decreased(P<0.05).The gene and protein expressions of Desmin,FSP-1 and GSK3βwere increased(P<0.05).The podocyte viability was decreased(P<0.01)and apoptosis was increased(P<0.01).Compared with model group,the Tailing Pills group’s gene and protein expressions of Podocin,CD2-AP,PI3K,Akt and P-Akt were significantly increased(P<0.05),and the gene and protein expressions of Desmin,FSP-1 and GSK3βwere decreased(P<0.05).The podocyte viability was increased(P<0.01)and apoptosis was decreased(P<0.01).Conclusion:Drug-containing serum of Tailing Pills can inhibit hyperglycemia-induced podocyte EMT,and the underlying mechanism is partially through regulating PI3K/AKT/GSK3βpathway.