TFP5通过NGF/SIRT1轴抑制Cdk5/p25活性干预糖尿病肾病足细胞损伤机制的研究

Podocyte damage and loss is a common feature in Diabetic Nephropathy (DN), which causes proteinuria and glomerolosclerosis. Previous studies showed that high glucose (HG) induce p25 expression; Cdk5/p25 overactivity causes podocyte hypertrophied, actin protein reorganized, morphological changed, and cell apoptosis. The mechanisms were correlated with oxidative stress, inflammatory factors, and signaling pathway regulation induced by glucose toxicity. SIRT1 is one of Cdk5 substrates; it plays an important protective role in cell differentiation, survival, anti-oxidation, anti-aging. High glucose causes Cdk5 hyperphosphorylation of SIRT1 and lost its protective function. Recent study demonstrated that NGF is a new cellular regulation of biological targets; and also showed that NGF can increase SIRT1 expression and activity and protect cells from damage. It suggests that NGF/SIRT1 axis may serve as a treatment target for cell damage. Our study in neuronal cells and islet β cells has revealed that Cdk5 inhibits peptides, TFP5 specifically inhibits Cdk5/p25 activity, reduces inflammatory responses and oxidative stress, and protects cells from damage,in turn, reduces cell apoptosis. We ask whether TFP5 have the same protective effects in podocyte which damaged by Cdk5/p25 pathway and whether this protective function works by regulation of the NGF/SIRT1 axis? These are still little known. In this study, we will investigate the therapeutic effects and mechanism of TFP5 on podoctes damage in DN by studying the TFP5 regulation of NGF/SIRT1 axis in vitro and in vivo. The aim of this study is to exploring the new therapeutic approach for DN treatment targeting on Cdk5.

肾小球足细胞损伤是糖尿病肾病肾小球硬化始动和进展主要原因。我们前期研究(81160093)发现高糖裂解p35产生p25，Cdk5/p25过度活性通过氧化应激、炎症因子引起足细胞损伤。SIRT1是Cdk5底物之一，在抗氧化、抗炎症中起重要保护作用。高糖环境Cdk5过度磷酸化SIRT1使其失去上述作用。研究发现NGF可以上调SIRT1的表达和活性，保护细胞免于损伤。NGF/SIRT1轴可能成为新的干预细胞损害治疗靶点。我们的研究(81460161) 证实Cdk5抑制短肽TFP5在神经细胞和胰岛β细胞抑制高糖诱导的Cdk5/p25活性，抑制炎症反应和氧化应激，减少细胞凋亡。TFP5是否在足细胞有同样的保护作用？是否通过NGF/SIRT1轴起作用？这些问题鲜为人知。本课题就以上问题用体内外实验，研究TFP5通过NGF/SIRT1轴对Cdk5/p25引起足细胞损伤机制的干预，寻找治疗糖尿病肾病新靶点

足细胞损伤是糖尿病蛋白尿产生和肾脏病进展的重要因素，长期高血糖可引起足细胞损伤，细胞数目减少和进行性肾小球硬化，CDK5/P25异常活化导致的微炎症反应、氧化应激是足细胞损伤的主要原因。我们的研究发现，TFP5 可有效地抑制 Cdk5/p25 的活性，抑制氧化应激反应，减少炎症因子的产生，故应用体内外实验，进一步从分子水平探讨 Cdk5/p25 通过NGF/SIRT1 轴在糖尿病肾病足细胞损伤的机制，以 Cdk5 为靶点，寻找2型糖尿病肾病的防治靶点。具体结果如下,1.在慢性葡萄糖毒性实验中,高糖可促进CDK5表达及活性增加，TFP5可通过抑制 Cdk5/p25 的活性对 NGF/ SIRT1 轴起调节作用，减少炎症因子分泌，抑制氧化应激反应，缓解足细胞凋亡。2.应用db/db小鼠模型完成TFP5抑制的体内实验，结果发现对db/db小鼠给予TFP短肽治疗后，肾脏CDK5、P35/25表达水平降低，NGF、Sirt1表达水平升高，足细胞损伤减少，炎症标记物IL-1、I-L6、IL-1β及氧化应激标记物H202、SOD、ROS表达下降。说明TFP5可通过降低体内炎症反应及氧化应激反应发挥对足细胞的保护作用 。