The role of DDR1 in podocyte lipotoxicity and progression of Alport Syndrome

DDR1 在足细胞脂毒性和 Alport 综合征进展中的作用

• 批准号: 9397084
• 负责人: JINJU KIM
• 金额: $ 6万
• 依托单位: UNIVERSITY OF MIAMI SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-07-01 至 2020-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9397084
• 关键词: Actins; Address; Affect; Apoptosis; Blood; Cells; Code; Collagen; Collagen Type I; Collagen Type IV; Data; Deposition; Development; Disease; Dominant-Negative Mutation; End stage renal failure; Exposure to; Extracellular Matrix; Eye; Fatty Acids; Fibrosis; Genes; Genetic; Goals; Hereditary Disease; Hereditary nephritis; Human; Hypertriglyceridemia; In Vitro; Inflammatory; Inherited; Injury; Injury to Kidney; Kidney; Kidney Diseases; Knock-out; Label; Labyrinth; Laminin; Lead; Life; Link; Lipids; Live Birth; Measures; Mediating; Mus; Mutation; Names; Nitrogen; Nonesterified Fatty Acids; Oils; Pathway interactions; Patients; Permeability; Pharmacologic Substance; Play; Production; Proteins; Proteinuria; Receptor Protein-Tyrosine Kinases; Renal function; Renal glomerular disease; Research; Role; Staining method; Stains; Testing; Time; Transfection; Triglycerides; Urine; deafness; discoidin domain receptor 1; glomerular basement membrane; glomerular filtration; improved; in vivo; kidney cell; kidney cortex; mRNA Expression; mouse model; new therapeutic target; novel; novel therapeutics; podocyte; prevent; scavenger receptor; uptake; young adult

Project Summary Podocytes are key components of the glomerular filtration barrier, and their interaction with the glomerular basement membrane (GBM) composition is crucial to maintain the permeability of this barrier. Although the GBM is primarily composed by laminin and Collagen type IV, the de novo production of the α1 chain of collagen type I (Col I) has been observed in mouse models of Alport Syndrome (AS). Discoidin domain receptor 1 (DDR1) is a unique receptor tyrosine kinase that is activated by collagens. Interestingly, deletion of the DDR1 in Col4a3 knockout (Col4a3KO) mice, a mouse model for AS, was shown to improve survival and renal function, suggesting that DDR1 may represent a potential pharmaceutical target for AS. However, if and how DDR1 activation by aberrant collagen production in AS contributes to podocyte injury and the development of proteinuria is poorly understood. Our preliminary data show that DDR1 activity is increased in kidney cortex from Col4a3KO mice. In addition, we show that the degree of DDR1 activation correlates with blood urine nitrogen (BUN). In vitro, we demonstrate that DDR1 is activated by collagen type I (50µg/mL, 18hr) in cultured human podocytes. In other preliminary data, we show for the first time that both collagen I and genetic activation of DDR1 are associated with cellular lipotoxicity, free fatty acid (FFA) uptake and intracellular lipid droplet deposition, suggesting that collagen I-induced/DDR1-mediated lipotoxicity may represent a novel mechanism leading to podocyte injury in AS. We propose to investigate this novel mechanism using a combined in vitro and in vivo approach. In specific aim 1, we will determine if DDR1-induced podocyte injury in AS is due to increased in FFA uptake and triglyceride accumulation. Specific aim 2 will test if DDR1-induced accumulation of triglycerides in podocytes is due to CD36 mediated FFA uptake. Lipid accumulation will be assessed using Oil Red O staining. FFA uptake will be measured by using fluorescent labeled FFAs. In vivo, we will test if genetic deletion of CD36 in Col4a3KO mice will protect from podocyte lipotoxicity and the development of proteinuria when compared to Col4a3KO mice. Findings derived from this application may demonstrate that DDR1 activation in AS induces podocyte triglyceride accumulation and apoptosis mediated through CD36 dependent fatty acid uptake. Results obtained from this research will set the basis for the development of novel drugs for the treatment of AS targeting DDR1 or CD36.

项目摘要 足细胞是肾小球滤过屏障的关键组成部分，它们与肾小球滤过屏障的相互作用， 肾小球基底膜（GBM）组成对于维持该屏障的渗透性至关重要。 虽然GBM主要由层粘连蛋白和IV型胶原组成，但α1-羟色胺的从头产生是不可能的。 在Alport综合征（AS）小鼠模型中观察到I型胶原蛋白链（Col I）。盘状结构域 受体1（DDR 1）是由胶原激活的独特受体酪氨酸激酶。有趣的是，删除 在AS小鼠模型Col 4a 3敲除（Col 4a 3 KO）小鼠中，DDR 1显示出改善存活率， 肾功能，表明DDR 1可能代表AS的潜在药物靶点。然而，如果和 AS中异常胶原蛋白产生导致的DDR 1激活如何导致足细胞损伤以及 对蛋白尿的发生了解甚少。 我们的初步数据显示，在Col 4a 3 KO小鼠的肾皮质中，DDR 1活性增加。在 此外，我们发现DDR 1的激活程度与血尿氮（BUN）相关。在体外，我们 证明在培养的人足细胞中，I型胶原蛋白（50µg/mL，18小时）可激活DDR 1。换句 初步数据，我们首次表明，胶原蛋白I和基因激活的DDR 1是相关的 细胞脂毒性、游离脂肪酸（FFA）摄取和细胞内脂滴沉积，表明 I型胶原诱导的/DDR 1介导的脂毒性可能代表了一种导致足细胞损伤的新机制， 如. 我们建议使用体外和体内相结合的方法来研究这种新的机制。在 具体目标1，我们将确定AS中DDR 1诱导的足细胞损伤是否是由于FFA摄取增加， 甘油三酯蓄积。具体目标2将测试DDR 1诱导的足细胞中甘油三酯的积累是否 这是由于CD 36介导的FFA摄取。将使用油红O染色评估脂质蓄积。FFA摄取 将通过使用荧光标记的FFA来测量。在体内，我们将测试是否有CD 36基因缺失， 当与对照组相比时，Col 4a 3 KO小鼠将保护足细胞脂毒性和蛋白尿的发展。 Col 4a 3 KO小鼠。 从该应用中得到的发现可以证明AS中DDR 1激活诱导足细胞 甘油三酯积累和通过CD 36依赖性脂肪酸摄取介导的细胞凋亡。获得的结果 本研究将为靶向DDR 1的AS治疗新药的开发奠定基础 或CD 36。