Mesonephric Model of Podocyte Injury and Regeneration

描述(申请人提供)：本次K99/R00申请的目的是利用我开发的一种新的动物模型系统来研究肾病综合征和足细胞再生的分子机制。这将有助于我的研究生涯向独立调查员职位的过渡。肾病综合征(NS)是一种由于肾小球滤过屏障(GFB)功能障碍而导致的以蛋白尿、低蛋白血症、水肿和高脂血症为特征的肾脏疾病。超过75%的激素抵抗肾病综合征(SRNS)的病因尚不清楚，目前尚无有效的治疗方法，可导致局灶性节段性肾小球硬化(FSGS)和终末期肾病(ESKD)。最近对SRNS遗传病因的发现和对足细胞损伤的啮齿动物模型的研究揭示了足细胞在GFB的正常功能和NS的发病机制中的关键作用。然而，NS的啮齿动物模型不适合高通量筛选该疾病的治疗药物。最近的研究在成人肾脏中发现了一种能够再生包括足细胞在内的肾上皮细胞的肾祖细胞亚群，这提示了一种基于干细胞/祖细胞的治疗肾病综合征的新方法。然而，人们对足细胞的再生机制知之甚少。为解决这些问题，我提议建立斑马鱼中肾细胞作为肾病综合征新的模型系统，方法是：(1)利用我获得的转基因斑马鱼模型筛选改善蛋白尿的化学物质；(2)研究表达wt1b的肾祖细胞在斑马鱼中再生足细胞的潜力以及足细胞再生过程中的基因表达；(3)利用所建立的GFB检测方法(S)筛选温度敏感型肾病斑马鱼突变体，并确定导致NS的新的遗传原因。拟议研究的完成将提供(1)可导致NS治疗的新化合物；(2)对足细胞再生机制的新见解；(3)参与NS发病机制的新基因；以及(4)治疗该疾病的新斑马鱼模型。作为密歇根大学儿科和传染病系的博士后研究员，我致力于将斑马鱼中肾开发成肾病综合征的新动物模型系统，作为我当前的职业目标。我的长期职业目标是在肾脏研究领域确立自己的独立研究员地位，专注于肾脏疾病的动物模型。该奖项的培训(K99)阶段将由弗里德海姆·希尔德布兰特博士指导，他是霍华德·休斯医学研究所的研究员，也是包括肾病综合征在内的儿童肾脏疾病的人类遗传学领域的国际公认领导者。密歇根大学拥有数十个积极合作的研究小组，致力于肾脏发育、肾脏疾病、斑马鱼发育和遗传学方面的工作，这为我的培训和职业发展提供了理想的环境。 公共卫生相关性：在美国，肾小球疾病占终末期肾脏疾病的90%，每年花费200亿美元。肾病综合征是由于肾小球滤过屏障的缺陷所致。最近的研究表明，足细胞和肾祖细胞功能障碍在肾小球的发病机制中起着关键作用。这一概念表明，足细胞和肾祖细胞是治疗这种疾病的新靶点。新的斑马鱼肾病综合征和足细胞损伤模型的建立，可以进行大规模的化学和遗传筛选，将为研究肾小球发病的分子机制和寻找治疗肾小球疾病的新方法奠定基础。

DESCRIPTION (provided by applicant): The objective of this K99/R00 application is to utilize a new animal model system that I have developed for the study of molecular mechanisms of nephrotic syndrome and podocyte regeneration. This will aid the transition of my research career toward an independent investigator position. Nephrotic syndrome (NS) is a kidney disease characterized by proteinuria, hypoalbuminemia, edema, and hyperlipidemia, due to the disruption of renal glomerular filtration barrier (GFB) function. The causes of over 75% of steroid resistant nephrotic syndrome (SRNS) cases are still unknown and no effective therapy is available for the disease, which results in focal segmental glomerular sclerosis (FSGS) and leads to end-stage kidney disease (ESKD). Recent discoveries of genetic causes of SRNS and studies of rodent models of podocyte damage revealed the pivotal role of podocytes in the normal function of GFB and the pathogenesis of NS. However, rodent models for NS are not suitable for high-throughput screening of therapeutical agents for the disease. Recent studies have identified a subset of renal progenitor cells in adult human kidney that are capable of regenerating renal epithelial cells, including podocytes, suggesting a novel stem/progenitor cell-based approach for the treatment of nephrotic syndrome. However, little is known about the regenerative mechanism of podocytes. To address these questions, I propose to establish zebrafish mesonephros as a new model system for nephrotic syndrome by: (1) utilizing the transgenic zebrafish models that I have generated to screen for chemicals ameliorating proteinuria; (2) investigating the potential of wt1b-expressing renal progenitor cell to regenerate podocytes in zebrafish and gene expression during podocyte regeneration; (3) using the established GFB assay(s) to screen for temperature-sensitive nephrotic zebrafish mutants and identify novel genetic causes of NS. Accomplishment of the proposed research will provide (1) new chemical compounds that can lead to therapies for NS; (2) new insights into the mechanism of podocyte regeneration; (3) new genes that are involved in the pathogenesis of NS; and (4) new zebrafish models for the disease. Being a postdoctoral research fellow in the Department of Pediatrics and Communicable Diseases, University of Michigan, I am committed to develop zebrafish mesonephros into a new animal model system for nephrotic syndrome as my immediate career goal. My long-term career goal is to establish myself as an independent investigator in the field of kidney research, focusing on animal models of kidney diseases. The training (K99) phase of this award will be mentored by Dr. Friedhelm Hildebrandt, who is an investigator of Howard Hughes Medical Institute and internationally recognized leader in the fields of human genetics of pediatric kidney diseases, including nephrotic syndrome. University of Michigan has dozens of active and collaborative research groups working on kidney development, kidney diseases, and zebrafish development and genetics, which provides an ideal environment for my training and career development. PUBLIC HEALTH RELEVANCE: Renal glomerular diseases account for 90% of end-stage kidney disease at a cost of $20 billion per year in the US. Nephrotic syndrome is due to the defective glomerular filtration barrier. Recent studies reveal dysfunction of podocytes and renal progenitor cells play critical roles in renal glomerular pathogenesis. This notion suggests podocytes and renal progenitor cells as new therapeutical targets for the disease. Generation of new zebrafish models of nephrotic syndrome and podocyte injury, which are amenable for large-scale chemical and genetic screenings, will allow for studies of molecular mechanisms of glomerular pathogenesis and search for novel therapy for glomerular diseases.