Project 2: Defining how the TGF- /FGF2 axis alters the fate of renin producing and vascular smooth muscle cells under conditions that threaten homoeostasis in infancy

总结/摘要 患有盐耗性肾病的幼儿患慢性和严重细胞外 液体（ECF）体积收缩，导致生长迟缓和肾灌注不良。慢性疾病 威胁体内平衡，如低血压、低钾血症、盐耗竭和/或长期使用 血管紧张素转换酶抑制剂（ACEi）或血管紧张素受体阻滞剂（ARB）治疗幼儿 诱导产生肾素的细胞的募集，导致肾小动脉肥大。这 然而，在不影响肾灌注的情况下，这种反应不能持续太长时间。然而，目前还不清楚 是参与这一过程的主要血管生长因子，以及它们如何影响产生肾素的细胞的生长。 细胞以前的研究，包括我们自己的初步工作，表明转化生长因子-b（TGF-b） 和成纤维细胞生长因子-2（FGF-2）与血管紧张素II相互作用，在调节血管张力中起关键作用， 肥大，肾小球（JG）和肾血管平滑肌细胞（RVSMC）增殖。 然而，关于TGF-β和FGF-2在调节肾素释放和血管生成中的作用知之甚少。 小婴儿的肾素生成细胞的表型。在这里，我们将测试的假设，在条件下， 这代表了在婴儿期维持ECF容量和/或肾灌注的生理威胁， Ang Ⅱ-TGF-B和FGF-2轴在维持肾素-血管紧张素Ⅱ的正常命运和功能中起着关键作用。 产生细胞和RVSMC。此外，我们假设，当RAS、TGF-β和 FGF-2通路被破坏，为肾素表型编程的细胞以无序的方式整合在细胞内。 肾小动脉，促进血管向心性肥大性病变的发展，导致不良的 肾灌注和肾纤维化。使用时间和细胞特异性条件删除方法，以及单细胞 转录组学分析，我们将在三个目标测试这一假设。在目标1中，我们将测试假设， Ren 1+细胞TGF-β受体1信号通路在维持JG和RVSMC肥大中的作用 对出生后早期饮食中Na+ / K+摄入或RAS抑制的慢性变化的反应，以及 确定肾素和cAMP途径如何与JG和RVSMC中的TGF-β和/或FGF-2相互作用以调节 内分泌和收缩表型。在目标2中，我们将检验FGF信号通路有助于 在慢性炎症条件下，维持JG和RVSMCs中RAS和TGF-β之间的适当平衡， 年轻小鼠的RAS刺激和抑制。在目标3中，我们将定义转录组谱和蛋白质 肾灌注不良的幼鼠肾小动脉和尿脱落细胞发生变化， 通过Na+耗竭和RAS抑制，并在来自幼儿的细胞和组织中验证这些发现 经历类似的情况。这些研究将阐明新的机制，即RAS与 TGF-b和FGF-2通路在婴儿期调节JG和RVSMC的生长，并维持ECF 在婴儿期慢性RAS刺激或抑制条件下的稳态和肾灌注。

SUMMARY/ ABSTRACT Young children with salt wasting renal diseases are at high risk of developing chronic and severe extracellular fluid (ECF) volume contraction, leading to growth retardation and poor renal perfusion. Chronic conditions that threaten homeostasis such as hypotension, hypokalemia, salt depletion, and/or the prolonged used of angiotensin-converting enzyme inhibitors (ACEi) or angiotensin receptor blockers (ARBs) in young children induce the recruitment of renin-producing cells with the resulting hypertrophy of the renal arterioles. This response however, cannot be sustained for too long without affecting renal perfusion. However, it is unclear what are the major vascular growth factors involved in this process, and how they affect the growth of renin producing cells. Previous studies, including our own preliminary work, suggest that Transforming Growth Factor-b (TGF-b) and Fibroblast Growth Factor-2 (FGF-2) interacting with Angiotensin II, play a key role modulating vascular tone, hypertrophy, and proliferation of juxtaglomerular (JG) and renal vascular smooth muscle cells (RVSMC). However, very little is known about the role that TGF-b and FGF-2 play in the regulation of renin release and the phenotype of renin producing cells in young infants. Here, we will test the hypothesis that under conditions that represent a physiological threat to maintain the ECF volume and/or renal perfusion in infancy, the Ang II-TGF-b and FGF-2 axis plays a critical role maintaining the normal fate and function of renin- producing cells and RVSMC. In addition, we hypothesize that when the balance between the RAS, TGF-b and FGF-2 pathways is disrupted, cells programed for the renin phenotype integrate in a disorderly manner inside the renal arterioles, precipitating the development of vascular concentric hypertrophic lesions leading to poor renal perfusion and kidney fibrosis. Using time and cell specific conditional deletion approaches, and single cell transcriptomic analysis, we will test this hypothesis in three aims. In aim 1 we will test hypothesis that a functional TGF-b receptor 1 signaling pathway in Ren1 + cells is necessary to sustain the hypertrophy of JG and RVSMC in response to chronic changes in dietary Na+ / K+ intake or RAS inhibition during early postnatal life, and determine how renin and the cAMP pathway interact with TGF-b and/or FGF-2 in JG and RVSMC to modulate their endocrine and contractile phenotypes. In aim 2, we will test the hypothesis that FGF signaling contributes to maintain the proper balance between the RAS and TGF-b in JG and RVSMCs under conditions of chronic RAS stimulation and suppression in young mice. In aim 3 we will define the transcriptome profile and protein changes that occur in renal arterioles and cells shed in the urine of young rats with poor renal perfusion induced by Na+ depletion and RAS inhibition, and validate these findings in cells and tissues derived from young infants undergoing similar conditions. These studies will elucidate new mechanisms whereby the RAS interacts with the TGF-b and FGF-2 pathways during infancy to modulate the growth of JG and RVSMCs, and maintain the ECF homeostasis and renal perfusion under conditions of chronic RAS stimulation or suppression in infancy.