The Role of LRG1 in Diabetic Kidney Disease

LRG1 在糖尿病肾病中的作用

• 批准号: 10176474
• 负责人: Kyung Lee
• 金额: $ 33.9万
• 依托单位: ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-08-01 至 2023-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10176474
• 关键词: Affect; Albuminuria; Architecture; Attenuated; Cell Line; Cells; Clinical Research; Clinical Trials; Complement; Cultured Cells; Development; Diabetes Mellitus; Diabetic Nephropathy; Diabetic mouse; Disease; Disease Progression; Early Intervention; End stage renal failure; Endothelial Cells; Endothelium; Event; Foundations; Functional disorder; Future; Genes; Genetic Models; Glucose; Glycoproteins; Human; In Vitro; Inflammatory Response; Injury; Injury to Kidney; Kidney; Knock-out; Knockout Mice; Leucine; LoxP-flanked allele; Mediating; Methods; Microalbuminuria; Modeling; Molecular; Mus; NOS3 gene; Non-Insulin-Dependent Diabetes Mellitus; Pathogenesis; Pathogenicity; Pathway interactions; Patients; Permeability; Process; Proteins; Public Health; Regulation; Role; Sampling; Serum; Signal Transduction; Streptozocin; TGF Beta Signaling Pathway; Testing; Therapeutic; Therapeutic Intervention; Tissues; Transforming Growth Factor beta; Transgenic Mice; Up-Regulation; Urine; angiogenesis; base; cancer cell; cell injury; db/db mouse; diabetes management; diabetic; differential expression; early detection biomarkers; effective therapy; endothelial dysfunction; glomerular endothelium; knock-down; mRNA Expression; malignant retina neoplasm; mouse model; non-diabetic; novel; overexpression; potential biomarker; promoter; single-cell RNA sequencing; transcriptome sequencing; transcriptomics

PROJECT SUMMARY Diabetic kidney disease (DKD) is the most common cause of end-stage renal disease in the US, with limited treatment options. Several recent clinical trials have not succeeded, likely owing to the lack of early intervention. Glomerular endothelial cell (GEC) dysfunction promotes the progression of DKD, and evidence suggests that GEC injury is an early event in diabetic kidneys that may precede the development of microalbuminuria. However, the mechanisms of early GEC injury in DKD remain unclear. Thus, better understanding of the underlying processes of GEC injury is urgently required for development of early therapeutic intervention. To date, the study of GEC injury in DKD has relied on the analysis of whole kidney tissues or isolated glomeruli, which provides indirect and limited information. Recently, we developed and optimized a novel method of effective isolation of GECs from transgenic mice expressing enhanced yellow fluorescent protein (EYFP) under the endothelium-specific Flk1 promoter. Using this approach, we compared the transcriptomic profiles of GECs isolated from streptozotocin (STZ)-induced diabetic endothelial nitric oxide synthase (eNOS)-null mice with GECs from nondiabetic controls. Many of the differentially expressed genes in diabetic GECs were involved in the regulation of angiogenesis and endothelial injury. Among these, a secreted glycoprotein, leucine-rich alpha-2 glycoprotein (LRG1) was found to be highly upregulated in diabetic GECs. LRG1 was recently shown to be the necessary co-activator of TGF-β signaling that is essential in neoangiogenesis of retinal and cancer cells. Thus, we explored whether LRG1 has a regulatory function in angiogenesis in early DKD.We found that 1) mRNA expression of LRG1 is significantly increased in GECs of diabetic mice, as well as in glomeruli of DKD patients; 2) correspondingly, LRG1 expression was found predominantly in GECs of mouse and human kidneys; 3) LRG1 protein is increased in serum and urine samples of DKD patients; 4) high glucose induces LRG1 expression in glomeruli and GECs cultured in vitro; 5) LRG1 knockout reduces albuminuria and glomerular injury, and attenuates angiogenesis in diabetic mice; and 6) LRG1 promotes angiogenesis via induction of the non-canonical TGF-β pathway in GECs, and knockdown of LRG1 in GECs is sufficient to diminish angiogenesis in vitro. Based on these salient findings, we hypothesize that LRG1 is a key pathogenic contributor of disease progression in early DKD by increasing angiogenesis and injury in GECs. Therefore, we propose to 1) determine the role of LRG1 in DKD using type 1 and type 2 DM mouse models; 2) determine the EC-specific role of LRG1 in early and late stages of DKD; and 3) determine the molecular mechanism of LRG1-induced diabetic GEC injury in vitro. Results from the proposed study will build the strong foundation for the future clinical studies to determine whether LRG1 is a potential biomarker of early DKD and whether the systemic blockade of LRG1 would offer therapeutic benefit to intervene against DKD progression.

项目摘要 糖尿病性肾脏疾病（DKD）是美国终末期肾脏疾病的最常见原因，有限 治疗选择。最近的一些临床试验尚未成功，这可能是由于缺乏早期 干涉。肾小球内皮细胞（GEC）功能障碍促进DKD的进展，证据 表明GEC受伤是糖尿病肾脏中的早期事件，可能在发展之前 微量白蛋白尿。但是，DKD早期GEC损伤的机制尚不清楚。那更好 迫切需要了解GEC伤害的基本过程才能提早发展 治疗干预。迄今为止，DKD中GEC损伤的研究已依赖于整个肾脏的分析 组织或孤立的glomerulli，提供间接和有限的信息。最近，我们开发了 优化了一种有效分离GEC的新方法，从表达增强的黄色的转基因小鼠 内皮特异性FLK1启动子下的荧光蛋白（EYFP）。使用这种方法，我们比较了 从链霉菌素（STZ）诱导的糖尿病内皮一氧化氮中分离的GEC的转录组曲线 合成酶（ENOS） - 无糖尿病对照的GEC的null小鼠。许多不同表达的基因 糖尿病GEC参与了血管生成和内皮损伤的调节。其中，一个分泌的 发现糖蛋白，富含亮氨酸的α-2糖蛋白（LRG1）在糖尿病性GEC中已高度更新。 最近，LRG1被证明是TGF-β信号的必要共激活因子，它在 视网膜和癌细胞的新血管生成。那，我们探讨了LRG1是否具有调节功能 DKD早期的血管生成。我们发现1）LRG1的mRNA表达在GEC中显着增加 糖尿病小鼠以及DKD患者的肾小球； 2）相应地，发现LRG1表达 主要在小鼠和人类肾脏的GEC中； 3）血清和尿液中LRG1蛋白增加 DKD患者的样本； 4）高葡萄糖在体外培养的肾小球和GEC中诱导LRG1表达； 5） LRG1敲除可减少蛋白尿和肾小球损伤，并减轻糖尿病小鼠的血管生成；和 6）LRG1通过诱导GEC中的非经典TGF-β途径促进血管生成和敲低 GEC中的LRG1足以减少体外血管生成。根据这些显着的发现，我们 假设LRG1是早期DKD疾病进展的关键致病性贡献者 GEC的血管生成和损伤。因此，我们建议使用类型1确定LRG1在DKD中的作用 和2型DM鼠标模型； 2）确定LRG1在DKD的早期和晚期中的EC特异性作用；和 3）确定体外LRG1诱导的糖尿病GEC损伤的分子机制。结果 拟议的研究将为未来的临床研究奠定坚实的基础，以确定LRG1是否是 DKD早期的潜在生物标志物以及LRG1的全身封锁是否会提供治疗益处 干预DKD进展。