ACE2-Ang-(1-7)-Mas轴对脂质肾损伤的影响及其机制研究

Lipid metabolic disorder is an independent risk factor for the development of renal disease. Researching the mechanism of lipid-induced renal injury and exploring its effective prevention and control measures are extraordinary significant to slow the progression of chronic kidney disease. In the hyperlipidemic state, mesangial cells can maintain steady cholesterol levels via negative feedback of SCAP-SREBP-2-LDLr, which alters receptor expression, to avoid excessive lipid accumulation and prevent renal fibrosis caused by the formation of foam cells. The ACE2-Ang(1-7)-Mas axis opposes proliferation, inflammation, oxidative stress，profibrotic actions, and participates in energy metabolism. However, the role of this axis in lipid-induced renal injury is unclear at present. Based on our previous work, this project aims to first show that the expression of the ACE2-Ang(1-7)-Mas axis in high-fat cultured mesangial cells and in a lipid-induced renal injury model in C57BL/6 mice can reduce the expression of LDLr to reduce lipid deposition in the kidney by enhancing SCAP-SREBP-2-LDLr negative feedback, adjust the fibrotic matrix-generated signal transduction pathway TGF-β/Smad3-CTGF, fiber substrate biodegradation signaling pathway TGF-β/Smad3-TIMPs/PAI,furthermore, to play an important role in reducing fibrosis formation in lipid-induced renal injury. Through this project, we can elucidate the mechanism and position of the ACE2-Ang(1-7)-Mas axis on improving both lipid metabolism and lipid-induced renal injury, and provide a new theoretical basis and new ideas for the clinical treatment of lipid-induced renal injury.

脂质代谢紊乱是肾脏疾病进展的独立危险因素，研究脂质肾损伤的发生机制及防治措施对延缓慢性肾脏病进展具有重要意义。高脂血症时肾脏系膜细胞能通过自身SCAP-SREBP-2-LDLr负反馈调节减少肾脏对脂质摄取，从而防止泡沫细胞形成所导致的肾脏纤维化。ACE2-Ang(1-7)-Mas轴参与抗增生、抗炎、抗纤维化以及机体物质能量代谢，但在脂质肾损伤中的研究国内外尚无报道。本项目旨在前期工作基础上首次探讨在高脂培养的系膜细胞及C57BL/6小鼠脂质肾损伤模型中，ACE2-Ang(1-7)-Mas轴通过增强SCAP-SREBP-2-LDLr负反馈调节降低LDLr表达、改善脂质代谢；进而通过抑制纤维生成及降解信号通路TGF-β/Smad3-CTGF、TGF-β/Smad3-TIMPs/PAI，抑制脂质导致的肾脏纤维化，以明确该轴在改善脂质代谢及脂质肾损伤的机制及地位，为临床治疗慢性肾脏病提供理论依据。

背景：血管紧张素转化酶2(ACE2)-血管紧张素1-7[Ang-(1-7)]-Mas受体轴是肾素-血管紧张素系统(RAS)的一个新的分支，其参与了脂质代谢调节的机制尚未明确。我们推测Ang-(1-7)能够调节低密度脂蛋白受体-固醇调节元件结合蛋白2-SREBP分解活性蛋白(LDLr-SREBP2-SCAP)负反馈系统，从而减少肾脏对脂质的摄取，进而改善脂质对肾脏的损伤。研究内容及结论：（1）体外部分：将人肾小球系膜细胞（HMC）分为六组：正常组，LDL组，LDL+Ang-(1-7)组， LDL+Ang-(1-7)+A-779组，Ang-(1-7)组和A-779组。油红O染色法检测人肾小球系膜细胞脂质沉积。化学酶促法检测细胞内胆固醇含量。实时荧光定量PCR和Western blot检测人肾小球系膜细胞中ACE2，LDLr，SREBP2，SCAP，TGF-β1和Smad3的表达。我们发现：首次证实了在HMC中，Ang-(1-7)可以通过调节LDLr-SREBP2-SCAP通路起到调控细胞对脂质吸收的作用，进而抑制LDL介导的TGF-β1/Smad3信号通路的活化（2）体内部分：将雄性C57BL/6小鼠随机分成四组：正常组（普通饲料+生理盐水），高脂组（高脂饲料+生理盐水），高脂+Ang-(1-7)组【高脂饲料+Ang-(1-7)】及Ang-(1-7)组【普通饲料+Ang-(1-7)】。收集小鼠血液检测血清总胆固醇（TC），甘油三酯 (TG)，低密度脂蛋白（LDL），血清尿素氮 (BUN)，血肌酐 (Scr)，C-反应蛋白 (CRP)，TNF-α和 IL-6。收集尿液检测尿微量蛋白。运用HE染色和透射电镜观察肾脏组织形态。通过免疫组化技术，实时荧光定量PCR和Western blot检测小鼠肾脏中LDLr, SCAP, SREBP-2, TNF-α, IL-6 and MCP-1的表达。运用油红O染色法检测小鼠肾脏脂质沉积。化学酶促法检测小鼠肾脏胆固醇含量。研究发现：首次证实Ang-(1-7)能够通过其抗炎作用调节高脂小鼠LDLr-SREBP2-SCAP轴，从而减轻脂质沉积，发挥肾脏保护作用。.意义：我们完成了Ang-(1-7)对改善脂质肾损伤的体内外部分可能机制的研究，从而更好的了解Ang-(1-7)在脂质肾损害中的作用，为研究脂质肾损害提供一种新的思路.