MDM2介导的有丝分裂灾难- - -糖尿病肾病足细胞损伤的新机制

Podocyte injury and loss are essential events of diabetic nephropathy. However, the mechanism of podocyte damage is not completely understood. It is well known that terminally differentiated cells are restricted in 'post-mitosis' state and cannot enter the cell cycle to divide, otherwise the outcome is catastrophic. The death caused by abnormal mitosis is defined as mitosis catastrophe. Murine double minute 2(MDM2) is a positive regulator of cell cycle and modulates the process of mitosis through several signaling pathways. Our preliminary data showed that mitotic podocytes are detected in patients diagnosed as diabetic nephropathy, simultaneously the expression of MDM2 and its downstream effector telomerase reverse transcriptase (TERT) are both upregulated. Moreover, the abundance of TERT is negative correlated with the number of podocytes. The expression of MDM2 and TERT in podocytes is also elevated significantly upon exposure to high glucose in vitro. Taken together, we hypothesize that in diabetic condition the mitosis catastrophe of podocytes induced by MDM2 is the major factor account for podocyte depletion. On the basis of our previous experiments we are going to investigate the significance of podocyte mitosis catastrophe in the pathogenesis of diabetic nephropathy and the effect of MDM2-TERT signaling pathway in above processes in cultured human podocytes, podocyte conditional MDM2 knockout mice and patients with diabetic nephropathy. By this investigation, we expect to clarify the molecular mechanism of podocyte injury and depletion in the early stage of diabetic nephropathy and shed light on the exploration of its effective therapies.

足细胞损伤和缺失是糖尿病肾病（DN）发病机制的关键环节，然而高糖所致足细胞损伤的机制尚未完全阐明。研究表明，作为终末分化细胞的足细胞一旦重新进入细胞周期发生有丝分裂将致其死亡-即有丝分裂灾难。MDM2是一种E3泛素连接酶，可通过多种信号通路调控细胞周期和有丝分裂。我们的前期研究发现，DN患者肾组织中存在异常有丝分裂的足细胞，并伴有MDM2及其下游分子TERT表达上调，且后者表达强度与足细胞数目呈负相关；体外研究显示，高糖可上调足细胞MDM2和TERT表达，提示MDM2介导的有丝分裂灾难可能是DN足细胞数目减少的重要原因。在前期研究基础上，本项目拟采用体外细胞模型、足细胞特异性MDM2基因敲除的DN小鼠模型和临床DN患者肾活检标本为研究对象，探讨足细胞有丝分裂灾难在DN进程中的病理生理意义及MDM2-TERT信号通路在足细胞有丝分裂灾难发生中的关键作用，以期为DN的早期防治提供新的理论依据。

足细胞损伤是糖尿病肾病（DN）发生发展的关键环节。作为一种终末分化细胞，足细胞可因重新进入细胞周期发生有丝分裂灾难而导致死亡，但足细胞有丝分裂灾难是否参与DN的进程尚不清楚。E3泛素连接酶MDM2是调控细胞周期和有丝分裂的关键分子，然而它在DN中的作用及机制尚未完全阐明。在本项目中，我们分别采用DN患者肾活检标本、体外细胞模型和STZ诱导的DN小鼠模型作为研究对象，探讨了足细胞有丝分裂灾难在DN进程中的病理生理意义及MDM2在足细胞有丝分裂灾难发生中的关键作用和调控机制，还探讨了MDM2在DN系膜细胞增殖及细胞外基质积聚中的作用及分子机制。本项目主要研究结果如下：（1）MDM2在不同病理类型原发性肾小球疾病患者肾组织中的表达均明显上调，且在FSGS患者中MDM2的表达与尿蛋白水平及ACR正相关，提示MDM2可能在足细胞损伤中发挥重要作用；（2）在DN患者、STZ诱导的DN小鼠模型以及体外高糖刺激的足细胞中均可观察到足细胞发生有丝分裂灾难现象，并伴有MDM2表达上调，而敲低MDM2可减轻足细胞有丝分裂灾难及足细胞损伤，表明MDM2可通过介导足细胞有丝分裂灾难参与DN的发生；（3）进一步研究提示，MDM2并非通过经典的p53依赖途径介导高糖导致的足细胞损伤，而是通过激活Notch1通路及下游分子Hes-1介导足细胞有丝分裂灾难；（4）在高糖刺激下系膜细胞MDM2表达明显上调，而敲低MDM2可减轻高糖诱导的系膜细胞增殖及细胞外基质合成, 其具体机制为MDM2通过结合NICD并影响其泛素化修饰从而激活其下游通路分子，最终导致肾小球系膜基质沉积。这些研究结果从细胞周期调节异常的角度，深化了人们对足细胞损伤机制的认识，明确了MDM2-Notch1信号通路在DN足细胞损伤中的关键作用，为DN的早期干预提供了新的理论和实验依据。