FEASIBILITY OF REPAIRING GBM DEFECTS IN VIVO

描述（由申请人提供）：肾小球基底膜（GBM）是一种特化的细胞外基质，是血管系统和泌尿空间之间肾脏肾小球滤过屏障（GFB）的重要组成部分。GBM最初由足细胞和肾小球内皮细胞共同合成，并位于足细胞和肾小球内皮细胞之间，并被每个毛细血管袢底部的系膜细胞结合。GBM的主要成分是层粘连蛋白-521（α 5 β 2 γ 1）、IV型胶原（α 3/4/5网络）、巢蛋白-1和-2以及硫酸肝素蛋白聚糖聚集蛋白。在这9种成分中，影响其中4种的突变会导致人类肾脏疾病：影响胶原IV链的突变会导致Alport综合征（遗传性肾小球肾炎），影响层粘连蛋白β 2（LAMB 2）的突变会导致皮尔逊综合征/先天性肾病综合征。肾小球缺陷的明确性质和良好表征的小鼠模型的存在使得这些疾病对于靶向治疗特别有吸引力。几个研究小组已经探索了基于细胞的疗法的潜力，旨在取代Alport小鼠中缺失的IV型胶原蛋白网络。虽然骨髓和其他细胞移植或输注对肾脏疾病进展的积极影响似乎很有希望，但这些程序对GBM胶原IV网络的报告影响是不同的和有争议的。在这里，我们提出的实验，旨在直接测试的可行性，改变GBM后，它已经成熟的小鼠模型Alport综合征和皮尔逊综合征/先天性肾病综合征，目标是取代异常的网络中存在的突变体与网络通常存在于健康的成熟肾小球。我们将使用新产生的足细胞特异性反向四环素反式激活因子转基因小鼠和现有的IV型胶原蛋白和层粘连蛋白突变体和转基因小鼠，试图诱导GBM成熟后的网络恢复。这些可行性研究的结果不仅对治疗人类肾脏疾病有意义，而且对我们对基底膜生物学，GBM可塑性和细胞/基质相互作用的基本理解也有意义！ 公共卫生相关性：肾小球基底膜（GBM）是肾小球滤过屏障的重要组成部分，可防止血浆蛋白渗漏到尿液中。一些肾病患者GBM组成异常。本提案的总体目标是使用小鼠模型系统来研究异常GBM组成是否可以在患者中正常化，以及这是否会改善和/或延长肾功能。

DESCRIPTION (provided by applicant): The glomerular basement membrane (GBM) is a specialized extracellular matrix and a crucial component of the kidney's glomerular filtration barrier (GFB) between the vasculature and the urinary space. The GBM is initially co-synthesized by, and lies between, the podocytes and the glomerular endothelial cells, and it is bound by mesangial cells at the base of each capillary loop. The GBM's major components are laminin-521 (alpha5beta2gamma1), type IV collagen (the alpha3/4/5 network), nidogens-1 and -2, and the heparin sulfate proteoglycan agrin. Of these 9 components, mutations that affect 4 of them cause human kidney disease: mutations that affect the collagen IV chains cause Alport syndrome (hereditary glomerulonephritis), and those that affect laminin beta2 (LAMB2) cause Pierson syndrome/congenital nephrotic syndrome. The well-defined nature of the glomerular defects and the existence of well- characterized mouse models make these diseases especially attractive for targeted therapy. Several groups have already explored the potential for cell-based therapies aimed at replacing the missing collagen IV network in Alport mice. Although the positive effects of bone marrow and other cell transplants or infusions on progression of kidney disease seem promising, the reported effects of these procedures on the GBM's collagen IV network have been disparate and controversial. Here we propose experiments aimed at directly testing the feasibility of altering the GBM after it has matured in mouse models of Alport syndrome and Pierson syndrome/congenital nephrotic syndrome, with the goal of replacing the abnormal networks present in the mutants with the networks normally present in healthy mature glomeruli. We will use newly generated podocyte-specific reverse tetracycline transactivator transgenic mice and existing collagen IV and laminin mutant and transgenic mice to attempt to induce network restoration after GBM maturation. The results of these feasibility studies will have implications not only for treating human kidney disease, but also for our basic understanding of basement membrane biology, GBM plasticity, and cell/matrix interactions! PUBLIC HEALTH RELEVANCE: The glomerular basement membrane (GBM) is an important component of the glomerular filtration barrier that prevents the leakage of plasma proteins into the urine. Some patients with kidney disease have abnormal GBM composition. The overall goal of this proposal is to use a mouse model system to investigate whether an abnormal GBM composition can be normalized in patients and whether this will improve and/or prolong kidney function.