Genetic Regulation, Tubular Processing and Clinical Relevance of Collecting Duct alpha-Defensins 1-3

项目总结/摘要 膀胱输尿管反流影响1 - 2%的儿童，其中三分之一的患者会出现尿路感染。 感染（UTI）。肾脏，特别是集合管在肾脏的先天防御中起着关键作用， 泌尿系统我们的研究小组已经证明，闰细胞是广泛的先天性免疫缺陷的主要来源。 免疫抗菌肽DEFA1A3（α-防御素1 - 3）及其抗菌蛋白（AMP）产物，人 中性粒细胞蛋白1 - 3（HNP 1 - 3）在人肾脏的先天免疫防御中是重要的。就在今年， 报道a）DEFA1A3 DNA拷贝数的变化与UTI易感性和抗生素相关， 膀胱输尿管反流和尿路感染儿童的反应，B）DEFA 1A 3 mRNA和蛋白产物、HNP 1 - 3表达 不限于白细胞，也存在于肾集合管上皮细胞中; c）尿HNP 1 - 3在治疗期间增加 尿路感染我们的上述发现与HNP 1 - 3作为AMP的已知功能相关， 微生物这些发现是重要的，因为VUR和UTI的管理是复杂的，目前无法 根据个人风险进行个性化治疗。例如，如果所有患者都在复发的假设下接受治疗， 肾盂肾炎的风险，儿童将暴露于不必要的成像相关的辐射和抗生素预防， 预防UTI。相反，如果所有患者均接受保守治疗，则一个子集将发生潜在可预防的肾功能衰竭。 疤痕和慢性肾病。此外，鉴定在肾集合管细胞中表达的DEFA1A3， 肾DEFA1A3表达可能成为治疗靶点。因为DEFA1A3 DNA拷贝数 这种变异与UTI有关，最近发现肾集合管细胞表达HNP 1 - 3， HNP 1 - 3在肾脏中的作用代表了临床相关的研究方向。根据以前的出版物和初步 研究中，我们假设肾脏和嗜中性粒细胞来源的DEFA1A3/HNP 1 - 3在细胞生理学中具有不同的作用。 病理学和DNA拷贝数的变化影响。 为此，我们将确定对肾脏储存、分泌和激活至关重要的独特分子机制- 在人类和鼠UTI中衍生的α防御素（Aim 1）。我们将利用蛋白质组学技术， 骨髓移植和尿中的泌尿道感染的儿童。在目标2中，我们将使用小鼠肾脏模型， 移植α-防御素的人源化小鼠，以检查肾脏在先天防御中的作用， 泌尿病原体在目标3中，我们将利用这种小鼠模型和体外研究来剖析DNA拷贝数的作用。 UTI易感性和抗生素协同作用中DEFA1A3基因座的变异。

PROJECT SUMMARY/ABSTRACT Vesicoureteral reflux affects 1-2% of all children and up to one third of these patients will experience urinary tract infections (UTI). The kidney and specifically the collect duct play a critical role in the innate defense of the kidney and urinary tract. Our group has demonstrated that intercalated cells are the primary source of a wide spectrum of innate immune antimicrobial peptides. DEFA1A3 (α-defensin 1-3) and its antimicrobial protein (AMP) product, human neutrophil protein 1-3 (HNP1-3), is important in the innate immune defense of the human kidney. Just this year we have reported that a) variations in the number of DEFA1A3 DNA copies correlate with UTI susceptibility and antibiotic response in children with vesicoureteral reflux and UTIs, b) DEFA1A3 mRNA and protein product, HNP1-3 expression is not limited to leukocytes, but also present in renal collecting duct epithelial cells and c) urine HNP1-3 increase during UTIs. Our aforementioned findings correlate with the known function of HNP1-3 as an AMP that rapidly destroys microbes. These finding are important because management of VUR and UTIs is complicated by the current inability to personalize treatment according to individual risk. For instance if all patients are treated under the assumption of recurrent pyelonephritis risk, children will be exposed to unnecessary imaging related radiation and antibiotic prophylaxis to prevent UTIs. Conversely if all patients are treated conservatively, a subset will develop potentially preventable renal scarring and chronic kidney disease. Further, identification of DEFA1A3 expressed in renal collecting duct cells, raises the possibility that renal DEFA1A3 expression could be targeted for therapeutics. Because DEFA1A3 DNA copy number variation is associated with UTI and the recent discovery that renal collecting duct cells express HNP1-3, determining HNP1-3’s role in the kidney represents clinically relevant research direction. Based on prior publications and preliminary studies, we hypothesize that renal- and neutrophil-derived DEFA1A3/HNP1-3 have distinct roles in cellular physiology/ pathology and are affected by DNA copy number variations. To this end, we will identify the unique molecular mechanisms critical to storage, secretion and activation of renally- derived alpha defensins in humans and murine UTI (Aim 1). We will utilize proteomic techniques using a mouse model of bone marrow transplantation and urine in children with UTI. In Aim 2, we will use a murine model of kidney transplantation of a humanized mouse for alpha-defensins to examine the kidney’s role in innate defense against uropathogens. In Aim 3, we will utilize this murine model and in vitro studies to dissect the role of DNA copy number variations in the DEFA1A3 locus in UTI susceptibility and antibiotic synergy.