Defining the renal urothelium progenitor and its regulation during development and repair

项目摘要/摘要 先天性尿路梗阻(UTO)是慢性肾脏疾病和终末期肾脏疾病的主要原因 儿童的疾病。目前的管理策略不能防止慢性肾脏疾病的进展。 我最近发表的F32资助的博士后研究数据表明，肾脏尿路上皮重塑是一种 对UTO的保护性适应。先天性和获得性尿路感染共同触发尿路蛋白(UPK)的形成 表达合成膀胱样尿路上皮斑块的尿路上皮细胞(UC)。尿路上皮斑块的消退 患有先天性尿路梗阻的小鼠加速肾实质损失、功能损害和死亡证明 UPK-UC在预防梗阻性肾病中的关键作用。对肾脏尿路上皮的进一步认识 开发和修复可能导致旨在减轻梗阻性肾脏疾病的治疗。基本原理 然而，在我们对发展和UTO期间UPK-UC的形成的理解中存在知识差距。 我最近证明，表达角蛋白5(K5)的UC在发育过程中会分化为UPK-UC 并遵循UTO。这一应用程序直接针对K5-UC是其命运的祖细胞的假设 是由一种内在的分子程序控制的，这种程序会随着时间的推移而变化，并对损伤做出反应。预期中的 本研究的结果是建立了控制K5-UC祖细胞活性的分子机制，并 确定K5-UC祖细胞在发育期间和UTO后的贡献。在《目标1》中，我将 用有机物分析法研究时间限制的K5-UC祖细胞活性是否是细胞固有的； 研究Notch信号在调节K5-UC命运中的作用；并确定调节的分子程序 尿路上皮的发育和修复。在目标2中，我将通过评估 输尿管电切术中K5-UC的耗竭和扩张对尿路上皮发育和修复的影响这些研究 在这一应用中提出的将揭示UPK-UC形成的细胞和分子基础-带有以下假设 K5-UC以凹槽依赖的方式形成UPK-UC。成功完成这些目标将提供 以K5-UC为靶点分化治疗梗阻性肾的基础 输尿管梗阻患者的疾病。为了实现这些目标，我将发展稀有细胞分离、有机化合物的关键技能。 梗阻的发生、信号通路分析、生物信息学、肾脏生理学和病理生理学 肾病。掌握这些技能，并结合有组织的职业发展活动 在我的导师和研究顾问团队的指导下，我将为成功竞争R01资金做好准备 作为一名独立科学家开始我的职业生涯，专注于开发旨在减轻慢性疾病的疗法 尿路梗阻儿童的肾脏疾病进展。

PROJECT SUMMARY/ABSTRACT Congenital urinary tract obstruction (UTO) is a leading cause of chronic kidney disease and end stage renal disease in children. Current management strategies cannot prevent chronic kidney disease progression. Recently published data from my F32-funded postdoctoral studies identify renal urothelial remodeling as a protective adaptation to UTO. Both congenital and acquired UTO trigger the formation of Uroplakin (Upk) expressing urothelial cells (UC) that synthesize a bladder-like urothelial plaque. Depletion of urothelial plaque in mice with congenital UTO accelerates renal parenchymal loss, functional impairment, and death – attesting to key roles for Upk-UCs in preventing obstructive nephropathy. A greater understanding of renal urothelial development and repair may lead to therapies aimed at attenuating obstructive kidney disease. Fundamental knowledge gaps exist, however, in our understanding of the formation of Upk-UCs during development and UTO. I have recently demonstrated that Keratin 5 (K5)-expressing UCs differentiate into Upk-UCs during development and following UTO. This application takes direct aim at the hypothesis that K5-UCs are progenitors whose fate is governed by an intrinsic molecular program that varies over time and in response to injury. The anticipated outcome of this research is to establish molecular mechanisms that govern K5-UC progenitor activity, and to determine the contributions of K5-UC progenitors during development and following UTO. In Aim 1, I will investigate whether temporally restricted K5-UC progenitor activity is cell-intrinsic using organoid assays; examine the role of Notch signaling in the regulation of K5-UC fate; and define molecular programs that regulate urothelial development and repair. In Aim 2, I will investigate the significance of the K5-UCs by evaluating the impact of depletion and expansion of K5-UCs on urothelial development and repair during UTO. The studies proposed in this application will reveal the cellular and molecular basis of Upk-UC formation – with the hypothesis that K5-UCs form Upk-UCs in a Notch dependent manner. Successful completion of these aims will provide the foundation for the development of therapies that target K5-UCs for differentiation to attenuate obstructive kidney disease in patients with UTO. In fulfilling these Aims, I will develop critical skills in rare cell isolation, organoid development, signaling pathway analysis, bioinformatics, renal physiology and pathophysiology of obstructive nephropathy. Mastery of these skills, in combination with structured career development activities under the guidance of my mentors and research advisory team, will prepare me to successfully compete for R01 funding and launch my career as an independent scientist focused on developing therapies aimed at attenuating chronic kidney disease progression in children with UTO.