Hippo通路在高糖诱导的肾小球系膜细胞增殖中的作用及机制

The proliferation of mesangial cells is essential during the early stage of diabetic nephropathy. The Hippo pathway is a newly found protein kinase cascade governing cell proliferation and apoptosis, but its relationship with the proliferation of mesangial cells has not been reported yet. According to our preliminary data, high glucose could activate the PI3K/Akt pathway and then suppress the Hippo pathway, accelerating the proliferation of mesangial cells, while knockdown of YAP by siRNA could suppress Akt activity. We make hypothesis that the Hippo pathway regulates the proliferation of high glucose-induced mesangial cells, where the YAP transcription coactivator act as a key factor to regulate the Hippo pathway in a positive manner. Accordingly, the current project is designed to investigate, from molecular, cellular to whole-animal studies, the effects of high glucose on the Hippo pathway, the mechanism by which Hippo medicates the proliferation of mesangial cells, and the positive feedback loop between PI3K/Akt and the Hippo pathway. The project will not only explain the molecular mechanism by which the Hippo pathway regulates the proliferation of mesangial cells, but also explore the potential effects of the pathway on the pathogenesis of diabetic nephropathy. The data to be obtained will provide substantial theoretical supporting for research and development of novel drugs against diabetic nephropathy and more fundamental basis for clinical therapy.

肾小球系膜细胞的增殖在糖尿病肾病早期的病理进程中极为重要。Hippo通路是新发现的调节细胞增殖/凋亡的蛋白激酶通路，其与系膜细胞增殖的关系尚无报道。基于前期研究发现高糖可通过激活PI3K/Akt，介导Hippo通路失活，促进系膜细胞增殖，而沉默Hippo通路下游YAP又可抑制Akt激活的现象，申请者提出Hippo通路参与调控了高糖诱导的系膜细胞增殖，且以YAP为关键环节的正反馈调控机制在此过程中发挥了重要作用。据此，本项目拟在分子、细胞以及整体动物水平，研究高糖对Hippo通路的影响、Hippo通路对系膜细胞增殖的调控作用，以及PI3K/Akt与Hippo通路的正反馈调控关系，阐明Hippo通路参与调控肾小球系膜细胞增殖的分子机制，揭示Hippo通路在糖尿病肾病病理进程中的作用，为糖尿病肾病防治药物的研发提供理论依据，为糖尿病肾病的临床治疗策略提供新的思路。

肾脏纤维化是糖尿病肾病重要的病理特征，其中肾小管上皮细胞-间质转分化和肾小球系膜细胞增殖在肾脏纤维化的病理进程中极为重要，然而其具体分子机制尚未完全阐明。Hippo通路是新发现的能够调节细胞增殖、分化、迁移的蛋白激酶通路，然而其与肾脏纤维化的关系缺乏研究。本研究通过细胞实验、动物模型以及肾脏病理等研究，采用基因转染与RNA干扰技术等，证实在体外培养的肾小管上皮细胞和肾小球系膜细胞，以及肾脏组织内，高糖或糖尿病状态可通过激活PI3K/Akt通路，降低Hippo通路核心蛋白激酶MST1、LATS1的磷酸化水平，介导Hippo通路失活。Hippo通路的失活，可以使其主要效应分子YAP蛋白表达水平上升，且迁移进入细胞核，并与转录因子TEAD结合，促进细胞周期蛋白E、细胞凋亡抑制因子等靶基因的表达，调控细胞周期及DNA合成，从而启动其促增殖和抗凋亡活性，诱导细胞的过度增殖；过度增殖的系膜细胞分泌大量的细胞外基质，促进了肾小球的硬化。此外，YAP激活入核，也促进snail、twist等核转录因子的表达，从而抑制E-cadherin转录，使上皮细胞呈现出间质细胞样表型，诱导HK-2细胞发生EMT，促进糖尿病大鼠肾纤维化，加快糖尿病肾病的病理进程。本项目的研究阐明了Hippo通路参与调控肾小管上皮-间质转分化及肾小球系膜细胞增殖的分子机制，揭示Hippo通路在糖尿病肾病，尤其是肾纤维化病理进程中的作用，为糖尿病肾病防治药物的研发提供新的靶标，为糖尿病肾病的临床治疗策略提供新的思路。