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.

项目概要 足细胞是肾小球滤过屏障的关键组成部分，其与肾小球滤过屏障的相互作用 肾小球基底膜（GBM）的组成对于维持这一屏障的通透性至关重要。 尽管 GBM 主要由层粘连蛋白和 IV 型胶原蛋白组成，但 α1 的从头产生 在 Alport 综合征 (AS) 小鼠模型中观察到 I 型胶原蛋白链 (Col I)。 受体 1 (DDR1) 是一种独特的受体酪氨酸激酶，可被胶原蛋白激活。 Col4a3 敲除 (Col4a3KO) 小鼠（一种 AS 小鼠模型）中的 DDR1 被证明可以提高生存率 肾功能，表明 DDR1 可能代表 AS 的潜在药物靶标。 AS 中异常胶原蛋白生成导致的 DDR1 激活如何导致足细胞损伤以及 人们对蛋白尿的发展知之甚少。 我们的初步数据表明 Col4a3KO 小鼠肾皮质中的 DDR1 活性增加。 此外，我们在体外表明 DDR1 激活程度与血尿氮 (BUN) 相关。 证明 DDR1 在培养的人类足细胞中被 I 型胶原蛋白（50μg/mL，18 小时）激活。 初步数据显示，我们首次证明 I 型胶原蛋白与 DDR1 基因激活相关 与细胞脂毒性、游离脂肪酸（FFA）摄取和细胞内脂滴沉积有关，表明 I 型胶原蛋白诱导/DDR1 介导的脂毒性可能代表了导致足细胞损伤的新机制 作为。 我们建议采用体外和体内相结合的方法来研究这种新机制。 具体目标 1，我们将确定 DDR1 诱导的 AS 足细胞损伤是否是由于 FFA 摄取增加和 具体目标 2 将测试 DDR1 诱导的足细胞中甘油三酯的积累是否存在。 由于 CD36 介导的 FFA 吸收，将使用油红 O 染色来评估脂质积累。 将使用荧光标记的 FFA 进行体内测量，我们将测试 CD36 是否存在基因缺失。 与其他小鼠相比，Col4a3KO 小鼠可防止足细胞脂毒性和蛋白尿的发生 Col4a3KO 小鼠。 该应用的结果可能表明 AS 中 DDR1 激活诱导足细胞 通过 CD36 依赖性脂肪酸摄取介导的甘油三酯积累和细胞凋亡。 这项研究将为开发针对 DDR1 的治疗 AS 的新药奠定基础 或CD36。