12-脂氧化酶对糖尿病肾小球内AT1受体功能的作用机制研究

It has been shown that AT1 receptor function is not only related to the level of AT1 receptor gene expression,but also related to the level of AT1 receptor in the plasma membrane, AT1 receptor internalization and its existing form.In this study, we select AT1 receptor-associated protein(ATRAP) and AT1 receptor homodimers as an important evaluation criteria of the AT1 receptor internalization and the change of AT1 receptor existing form respectively.In order to clarify the hypothesis that the effect of 12-Lipoxygenase(12-LO) on the ATRAP and plasma membrane AT1 receptor existing form is significantly associated with the function of AT1 receptor in diabetic glomeruli, based on the previous study that 12-LO upregulate AT1 receptor gene expression via p38MAPK pathway that is related to the progression of diabetic nephropathy(DN), we further investigate the relationship between 12-LO and ATRAP,12-LO and AT1 receptor homodimers respectively using the molecular biology technique as a research tool with mesangial cells, podocytes,12-LO gene knockout mice glomeruli and type 1 and type 2 diabetic glomeruli.The purpose of this study is to completely clarify the pathophysiological effect of 12-LO on the AT1 receptor function that play an important role in the progression of DN,and provide new idea and important clue to further clinical trial for usning 12-LO inhibitor to attenuate the progression of DN.

AT1受体功能不仅与AT1受体的表达水平有关，而且与质膜上AT1受体表达量、内化及其存在形式有着密切的关系。本研究以12-脂氧化酶(12-LO)通过p38MAPK上调AT1受体基因表达而促进糖尿病肾病(DN)进展的前期结果为基础，分别选择与AT1受体的内化及其存在形式密切相关的AT1受体相关蛋白(ATRAP)和AT1受体同源二聚体的变化为评价其功能的重要指标，假设糖尿病肾小球内12-LO对ATRAP和AT1受体存在形式的作用是影响质膜上AT1受体功能的关键因素，以系膜细胞、足细胞、12-LO基因敲除小鼠肾小球、1型和2型糖尿病动物肾小球为研究对象，利用先进的分子生物学技术，深入探讨糖尿病肾小球内12-LO与ATRAP、12-LO与AT1受体同源二聚体的关系及其机制，从而完整地阐明DN时12-LO影响AT1受体功能的本质，为通过抑制12-LO途径延缓DN进展提供依据。

血管紧张素Ⅱ（AngⅡ）1型受体（AT1）功能不仅与 AT1的表达水平有关，而且与质膜上AT1表达量、内化及其存在形式有着密切的关系。本研究以12-脂氧化酶(12-LO)通过 p38MAPK上调AT1表达进而促进糖尿病肾病(DN)进展的前期结果为基础，分别选择与AT1的内化及其存在形式密切相关的AT1相关蛋白(ATRAP)和AT1同源二聚体的变化为评价其功能的重要指标，假设糖尿病肾小球内12-LO对ATRAP和AT1存在形式的作用是影响质膜上AT1功能的关键因素，以肾小球细胞、12-LO 基因敲除小鼠肾小球、1 型和2 型糖尿病动物肾小球为研究对象，利用先进的分子生物学技术，深入探讨糖尿病肾小球内12-LO与ATRAP、12-LO与AT1存在形式的关系及其机制。本研究发现12-LO作用产物12(S)-HETE通过p38MAPK信号通路诱导系膜细胞和肾小球内AT1表达增加，而ATRAP表达降低。我们尚未发现12-LO对系膜细胞和糖尿病肾小球内AT1受体二聚体的影响。AT1在2型糖尿病大鼠肾小球内表达增高，而在1型糖尿病大鼠肾小球内无变化，且胰岛素抵抗诱导肾小球细胞内AT1表达上调。12-LO抑制剂治疗对1型糖尿病大鼠肾小球AT1无显著影响。2型糖尿病大鼠肾小球内AT1表达增加与ATRAP表达减少有关。12-LO抑制剂治疗后2型糖尿病大鼠肾小球AT1表达减少，ATRAP表达增加。12-LO和AngⅡ之间存在相互影响和相互协同作用，因此抑制12-LO可以降低AngⅡ水平。通过抑制12-LO降低DN肾小球内12（S）-HETE和AngⅡ，从而增加ATRAP表达，同时降低AT1表达。12-LO与AngⅡ对AT1表达作用相反，因此抑制12-LO时DN肾小球内AT1的变化主要与12（S）-HETE有关，与AngⅡ无关。12-LO通过p38MAPK信号通路调节肾小球内ATRAP水平，影响AT1内化，从而增加DN肾小球内AT1表达。因此，抑制12-LO通过增加DN肾小球内ATRAP表达、降低AT1表达，从而减弱AngⅡ的作用而延缓DN的进展。