apelin 通过改变肾动脉自身调节功能和足细胞结构参与2型糖尿病肾病的发生发展

The earliest lesions of diabetic nephropathy consist of hyperperfusion in kidney and glomeruli and increased microalbumin in urine. Apelin, as a endogenous adpokine, could contract or relax the arterioles through acting on smooth muscle cells or endothelial cells. At the same time, we observed changed permeability of podocytes. Therefore, we hypothesized that apelin, increased with obesity in type 2 diabetes, change the autoregulation of renal artery via acting on smooth muscle cells, which lead to the overflow of kidney. In addition, apelin may promote the functional and morphological change of podocyte after type 2 diabetes. In this study, kk mice are considered to be a type 2 diabetic model. Atuoregulation in renal artery was analysised by vascular perfusion in vitro. Blood flow in renal artery and kidney were detected with ultrasonic measurement to demonstrate the effects of apelin or F13A ( antagonist of apelin) on autoregulation of renal artery.Cultured smooth muscle cells were used to detect effects of apelin or F13A onthe Ca2+ influx and myosin light chanin kinase and myosin light chain phosphatase expression to demonstrate the constractive effects of apelin on smooth muscle cells.Electronic microscopy were used to detect the structure of podocyte in kidney and culture medium with apelin or F13A treatment. cultured podocyte were used to demonstrate the effects of apelin or F13A on permeability of podocyte.Immonchemical staining and specific staining were used to demonstrate the effects of apelin or F13A on glomerular sclerosis, proliferation of mesangial cells in glomerular filtration membarane,capillary collapase and hyaline of aterioles in kidney. Blood urea nitrogen, creatinine in blood and urine,microalbumin and βmicroglobulin in urine are analysised to evaluate the renal functional changes before or after apelin or F13A treatment. After this study, we can conclude that apelin is one of the important factors promoting progression of diabetic nephropathy in type 2 diabetes via changing the autoregulation of renal artery and permeability of podocyte.

糖尿病肾病早期表现主要为肾脏血液灌注量增加、微量白蛋白尿。Apelin作为内源性的脂肪因子可以改变血管收缩舒张功能及内皮细胞通透性。因此，我们认为apelin可以改变肾动脉的其自身调节机制增加肾脏血液灌注量，同时增加肾小球滤过膜足细胞的通透性。本工作以kk小鼠作为2型糖尿病模型， 通过肾动脉离体灌流和肾动脉血流速度检测，证明其对肾动脉自身调节功能及肾脏血流量的影响；通过血管平滑肌细胞培养，观察apelin对细胞内Ca2+离子浓度及肌球蛋白轻链激酶等收缩蛋白的表达影响，论证其对平滑肌细胞功能的影响；通过电镜及足细胞培养检测apelin对其结构及通透性的作用，论证其对肾小球滤过膜通透性的影响；通过组织切片染色，论证apelin对肾小球基质增生、毛细血管塌陷及小动脉血管硬化的影响；通过对小鼠血、尿检测，论证apelin对肾功能的影响。通过以上研究找到apelin促进糖尿病肾病发生发展的作用环节。

糖尿病肾病早期表现主要为肾脏血液灌注量增加、微量白蛋白尿。Apelin 作为内源性的脂肪因子可以改变血管收缩舒张功能及内皮细胞通透性。因此，我们认为apelin可以改变肾动脉的其自身调节机制增加肾脏血液灌注量，同时增加肾小球滤过膜足细胞的通透性。本工作以kk 小鼠作为2 型糖尿病模型， 通过超声检测肾动脉血流量并通过微循环检测装置检测肠系膜动脉的自主调节功能，结果发现：apelin可以减轻小动脉自身调节功能并增加肾脏血流量；通过共聚焦显微镜观察培养血管平滑肌细胞内钙离子浓度改变，结果发现：apelin抑制牵张刺激引起的细胞内Ca2+离子浓度增加以及肌球蛋白轻链激酶等收缩蛋白的磷酸化；通过电镜及免疫荧光染色观察足细胞足突的改变，结果发现：apelin可以促进糖尿病时足细胞足突融合、足细胞凋亡。通过培养足细胞论证apelin对足细胞损伤的机制，结果发现：apelin可以通过抑制蛋白酶体活性导致足细胞内质网应激，通过激活mTOR抑制足细胞自噬，并最终引起足细胞通透性的改变。通过对小鼠血、尿检测及PAS染色和免疫组化染色，发现apelin可以促进糖尿病肾小球滤过膜损伤、尿微量白蛋白增加以及肾小球基底膜增厚肾。通过以上研究认为：apelin 通过改变小动脉收缩舒张功能增加肾脏血流量，并导致足细胞结构和功能改变，促进糖尿病肾病发生发展。