Novel Models of Diabetic Nephropathy

描述(由申请人提供)： 背景：在OVE26 1型糖尿病小鼠，b)饮食诱导肥胖和胰岛素抵抗的C57BI/6小鼠，以及c)2型糖尿病db/db小鼠中，我们发现肾脏转录因子的表达增加，i)固醇调节元件结合蛋白1和2(SREBP-1和SREBP-2)，以及ii)糖反应元件结合蛋白(ChREBP)，导致甘油三酯和胆固醇的合成和积累增加。脂质堆积与强健的肾小球硬化、肾小管间质纤维化和蛋白尿的发展有关。我们还发现法尼类X受体(FXR)在肾脏中高表达，而FXR及其靶酶在糖尿病肾脏中的表达减少。此外，我们已经确定FXR是SREBP-1和ChREBP表达的重要调节因子，以及氧化应激、晚期糖基化终产物(AGEs/RAGE)、促炎细胞因子和纤维化诱导生长因子。 假设：基于这些发现，我们认为FXR在糖尿病肾病的发病机制中起重要作用。我们假设，在1型(OVE26)和2型(db/db)糖尿病小鼠模型中，FXR基因的缺失将显著增强，而FXR的过度表达将减轻糖尿病肾病。 小鼠模型1：a)我们将产生FXR基因敲除小鼠，目前位于C57BI/6遗传背景上，背景：a)OVE26患有1型糖尿病的小鼠，b)饮食诱导肥胖和胰岛素诱导的C57BI/6小鼠FVB，如果需要，BE和DBA/2J遗传背景，2)已被证明具有增加糖尿病肾病易感性的遗传背景。B)然后我们将以FVB为背景的FXR KO小鼠与I)以FVB为背景的OVE26小鼠(1型糖尿病)或II)db/db小鼠(以FVB为背景)杂交，以确定FXR缺失是否会加重和加速糖尿病肾病。C)我们将使用Lox-Cre方法(FXRf/f小鼠与Nphs2 Cre小鼠杂交)在FVB背景下产生肾足细胞特异性FXR基因敲除小鼠。D)然后我们将足细胞FXR KO小鼠与I)OVE26小鼠或II)db/db小鼠杂交。小鼠模型2：A)我们将在FVB背景下产生肾足细胞特异性、条件性和可诱导的FXR转基因小鼠。B)然后我们将足细胞特异性FXR转基因小鼠与OVE26或db/db小鼠杂交，以确定FXR表达增加是否可以减轻或预防糖尿病肾病。 表型：在这些小鼠中，我们将确定a)糖尿病肾病的表现，包括肾小球滤过率、肾小球硬化、肾小管间质纤维化和蛋白尿，以及b)介导糖尿病肾脏损害的细胞和生化机制，包括脂质和碳水化合物代谢、炎症、纤维化、氧化应激和AGEs/RAGE。

DESCRIPTION (provided by applicant): Background: In a) OVE26 mice with type 1 diabetes, b) C57BI/6 mice with diet induced obesity and insulin resistance, and c) db/db mice with type 2 diabetes mellitus, we have found increased renal expression of the transcriptional factors, i) the sterol regulatory element binding proteins 1 and 2 (SREBP-1 and SREBP-2), and ii) the carbohydrate response element binding protein (ChREBP), which result in increased synthesis and accumulation of triglyceride and cholesterol. The lipid accumulation is associated with development of robust glomerulosclerosis, tubulointerstitial fibrosis, and proteinuria. We have also found that the Farnesoid X Receptor (FXR) is highly expressed in the kidney and the expression of FXR and its target enzymes is decreased in the diabetic kidney. Furthermore we have determined that FXR is an important regulator of SREBP-1 and ChREBP expression as well as oxidative stress, advanced glycation end products (AGEs/RAGE), pro-inflammatory cytokines, and fibrosis inducing growth factors. Hypothesis: Based on these finding we propose that FXR plays an important role on the pathogenesis of diabetic nephropathy. We hypothesize that deletion of FXR will markedly enhance and overexpression of FXR will attenuate diabetic nephropathy in mouse models of type 1 (OVE26) and type 2 (db/db) diabetes. Mouse Model 1: A) We will generate FXR knockout mice, currently on the C57BI/6 genetic background, on Background: In a) OVE26 mice with type 1 diabetes, b) C57BI/6 mice with diet induced obesity and insulin the FVB, and if need be and DBA/2J genetic backgrounds, 2 genetic backgrounds that have been documented to have increased susceptibility to diabetic nephropathy. B) We will then cross breed FXR KO mice on FVB background with i) OVE26 mice (type 1 diabetes) on FVB background or ii) db/db mice (type 2 diabetes) on FVB background to determine if FXR deletion accentuates and accelerates diabetic nephropathy. C) We will generate renal podocyte specific FXR knockout mice on FVB background using the Lox-Cre approach (FXRf/f mice crossed with Nphs2 Cre mice). D) We will then crossbreed podocyte FXR KO mice with i) OVE26 mice or ii) db/db mice. Mouse Model 2: A) We will generate renal podocyte specific conditional and inducible FXR transgenic mice in the FVB Background. B) We will then crossbreed the podocyte specific FXR transgenic mice with OVE26 or db/db mice to determine if increased expression of FXR attenuates or prevents diabetic nephropathy. Phenotyping: In these mice we will determine a) the manifestation of diabetic nephropathy, including glomerular filtration rate, glomerulosclerosis, tubulointerstitial fibrosis, and proteinuria and b) the cellular and biochemical mechanisms that mediate diabetic renal injury including lipid and carbohydrate metabolism, inflammation, fibrosis, oxidative stress, and AGEs/RAGE.