TGF-beta superfamily in neonatal obstructive nephropathy and recovery

描述（由申请人提供）：先天性梗阻性肾病是婴儿和儿童肾损害的最重要可识别原因。尽管如此，手术干预的适应症是有争议的，肾功能恢复往往令人失望。虽然在成年动物完全单侧输尿管梗阻（UUO）模型中阐明了尿路梗阻导致肾损伤的机制，但发育中的肾脏对梗阻的反应非常不同。转化生长因子-2（TGF-2）超家族在肾脏发育、伤口愈合、细胞存活、表型上皮-间质转化（EMT）和纤维化中起着重要作用。本研究计划将利用新开发的小鼠模型研究TGF-2超家族在新生小鼠中调节损伤和部分UUO恢复的肾细胞反应，其肾脏发育与20周的人类胎儿相当。该模型允许研究梗阻肾的损伤和恢复，以及对侧肾的补偿。TGF-21信号传导的关键组分的突变体和抑制剂的组合以及反调节骨形态发生蛋白-7（BMP- 7）的组合将用于阐明分子机制。细胞外TGF-21将通过给予核心蛋白聚糖而减少; TGF-21受体活性将通过选择性抑制ALK 5来研究;而TGF-21细胞内信号传导将在Smad 3缺失小鼠中研究。细胞外BMP-7将以3种方式进行操作：在Kielin/chordin样蛋白（KCP）缺失小鼠中降低内源性BMP-7活性;在子宫致敏相关基因-1（USAG-1）缺失小鼠中增加内源性BMP-7;以及外源性重组BMP-7。将通过免疫组织化学和定量形态测量学研究肾成熟、增殖、凋亡和EMT以及巨噬细胞、成纤维细胞和胶原沉积的指标。将平行进行体外细胞培养研究，并将获得的信息（细胞内或连接蛋白的拉伸诱导调节）应用于动物研究。手术缓解梗阻与TGF-21的操作相结合，可能会导致新的生物标志物或治疗方法在儿童梗阻性肾病中进行测试。公共卫生相关性：所有肾衰竭儿童中有一半出生时就有泌尿道缺陷，其中最常见的是由于尿液流动受阻。手术干预的时机和指征尚不清楚，恢复情况也无法预测。提出了一种新的动物（小鼠）模型来研究新生儿肾脏损伤的主要细胞机制，以及评估新的治疗方法以促进恢复。

DESCRIPTION (provided by applicant): Congenital obstructive nephropathy is the most important identifiable cause of renal impairment in infants and children. Despite this, indications for surgical intervention are controversial, and renal recovery is often disappointing. While mechanisms responsible for renal injury resulting from urinary tract obstruction are elucidated in models of complete unilateral ureteral obstruction (UUO) in adult animals, the developing kidney responds very differently to obstruction. The transforming growth factor-2 (TGF-2) superfamily plays a central role in renal development, wound healing, cell survival, phenotypic epithelial-mesenchymal transition (EMT), and fibrosis. This research plan will utilize a newly developed murine model to study the renal cellular response of the TGF-2 superfamily in regulating injury and recovery from partial UUO in the neonatal mouse, whose renal development is comparable to that of a 20-week human fetus. The model permits study of injury and recovery by the obstructed kidney, and compensation by the contralateral kidney. A combination of mutants and inhibitors of critical components of TGF-21 signaling, as well as of counter-regulatory bone morphogenetic protein-7 (BMP- 7) will be used to elucidate molecular mechanisms. Extracellular TGF-21 will be decreased by administration of decorin; TGF-21 receptor activity will be investigated by selective inhibition of ALK5; while TGF-21 intracellular signaling will be studied in Smad3 null mice. Extracellular BMP-7 will be manipulated in 3 ways: reduced endogenous BMP-7 activity in the kielin/chordin-like protein (KCP) null mouse; increased endogenous BMP-7 in the uterine sensitization-associated gene-1 (USAG-1) null mouse; and exogenous recombinant BMP-7. Indices of renal maturation, proliferation, apoptosis, and EMT, as well as macrophages, fibroblasts, and collagen deposition will be studied by immunohistochemistry and quantitative morphometrics. In vitro cell culture studies will be performed in parallel, and information gained (stretch-induced modulation of intracellular or junctional proteins) will be applied to the animal studies. The combination of surgical relief of obstruction with manipulation of TGF-21 may lead to new biomarkers or therapies to test in children with obstructive nephropathy. PUBLIC HEALTH RELEVANCE: Half of all children with kidney failure are born with birth defects of the urinary tract, and the most common of these are due to obstruction to urine flow. The timing and indications for surgical intervention are unclear, and recovery is unpredictable. A new animal (mouse) model is proposed to study the major cellular mechanisms for injury to the newborn kidney, as well as to evaluate new therapies to enhance recovery.