Role of Peroxidasin in Glomerular Matrix Homeostasis

过氧化物酶在肾小球基质稳态中的作用

• 批准号: 8700396
• 负责人: Gautam Bhave
• 金额: $ 14.51万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-07-15 至 2018-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8700396
• 关键词: Academic Medical Centers; Acids; Affect; Afferent Neurons; Apoptosis; Atherosclerosis; Attention; Basement membrane; Binding; Biochemical; Biochemistry; Biological Assay; Biology; Career Choice; Cell Adhesion; Cell Communication; Cell Surface Proteins; Cell physiology; Cells; Chemicals; Clinical; Collagen Type IV; Defect; Development; Diabetes Mellitus; Diabetic Nephropathy; Disease; Doctor of Medicine; Doctor of Philosophy; End stage renal failure; Environment; Enzymes; Epithelial Cells; Exhibits; Extracellular Matrix; Fibrosis; Functional disorder; Funding; Generations; Glucose; Goodpasture antigen; Growth Factor; Head; Homeostasis; Hospitals; Hydrogen Peroxide; Hyperglycemia; Hypochlorous Acid; Immunoglobulin Domain; Immunoglobulins; Inflammation; Injury; Integrins; Intervention; Investigation; Kidney; Kidney Diseases; Knowledge; Laboratories; Lead; Leucine-Rich Repeat; Lipids; Manuscripts; Mass Spectrum Analysis; Mediating; Medicine; Mentors; Modeling; Mus; Nature; Nephrology; Neurons; Neurosciences; Oxidants; Pathogenesis; Pathologic; Patients; Peroxidases; Phenotype; Phloroglucinol; Play; Post-Translational Protein Processing; Production; Protein Binding Domain; Proteins; Publications; Published Comment; Publishing; Pyridoxamine; Reactive Oxygen Species; Renal Hypertension; Renal function; Research; Role; Scientist; Seminal; Signal Transduction; Site; Superoxides; Tertiary Protein Structure; Testing; Time; Tissues; Training; Translating; Tubular formation; Tyrosine; United States National Institutes of Health; Up-Regulation; Wit; Work; base; career development; college; crosslink; db/db mouse; diabetic; experience; glomerular basement membrane; glycation; halogenation; improved; in vitro Model; in vivo; inhibitor/antagonist; innovation; insight; instructor; interstitial; kidney cell; mouse model; mutant; novel; oxidative damage; peroxidasin; podocyte; public health relevance; therapeutic target; von Willebrand Factor

DESCRIPTION (provided by applicant): Diabetic nephropathy (DN) is the leading cause of end-stage renal disease in the U.S. A central derangement in diabetic nephropathy is excessive generation of reactive oxygen species (ROS). Superoxide and its product, hydrogen peroxide (H2O2), are ROS that have garnered greatest attention in DN. We recently discovered that peroxidasin, a protein embedded in extracellular matrix, converts H2O2 to highly reactive hypobromous (HOBr) and hypochlorous (HOCl) acid, collectively denoted as hypohalous acids (HOX). Peroxidasin uses HOX to form novel sulfilimine bonds (S=N) in basement membrane collagen IV, the first identified function for peroxidasin. But, in a diabetic milieu saturated wit H2O2 substrate, peroxidasin may produce excessive HOX. HOX are highly reactive and oxidatively halogenate proteins to adversely affect cell function. In preliminary work using a mouse model of diabetic nephropathy, we have found that glomerular and tubular peroxidasin expression and HOX mediated protein damage are increased. Based on these findings, we hypothesize that peroxidasin plays a pathogenic role in diabetic nephropathy by generating toxic HOX that halogenate proteins to alter cell function and promote extracellular matrix expansion. To test this hypothesis, Aim 1 will determine how peroxidasin functions within basement membranes examining the interaction between collagen IV and peroxidasin. In Aim 2, we will examine the role of peroxidasin and HOX in hyperglycemia induced renal cell dysfunction and matrix accumulation as an in vitro model of diabetic nephropathy. Using innovative mass spectrometry, we will identify specific sites of HOX mediated protein halogenation associated with peroxidasin upregulation. In Aim 3, we will directly test the role of peroxidasin and HOX in a mouse model of diabetic nephropathy. Building upon the mass spectrometry studies in Aim 2, we will identify protein halogenation sites in vivo. Furthermore, we will pharmacologically block peroxidasin and determine whether the intervention ameliorates diabetic nephropathy. The proposed work holds promise to provide insight into the pathogenesis of diabetic nephropathy and establish peroxidasin as a potential therapeutic target. Environment The proposed studies will be conducted at the Vanderbilt University Medical Center within the Division of Nephrology. The division has 14 NIH funded laboratories with over 13 million dollars in annual NIH funding and is one of only nine NIH George O'Brien Kidney Research Centers in the U.S. thus providing Dr. Bhave ample financial, administrative, and scientific support. Dr. Bhave is primarily mentored by Dr. Billy Hudson, a well- established scientist with nearly 40 years of NIH funding. His expertise resides in basement membrane and type IV collagen biochemistry and his seminal work includes the identification of the Goodpasture antigen and the development of pyridoxamine as a glycation inhibitor for the treatment of diabetic nephropathy. Dr. Bhave is also supported by Drs. Raymond Harris and Roy Zent as co-mentors to help develop an independent line of investigation and career path. Dr. Harris is Chief of the Division of Nephrology and his experience in models of diabetic nephropathy is crucial to the proposed work. Dr. Zent heads a well-funded group studying renal development and integrins and will provide expertise in the investigation of peroxidasin and cell-matrix interactions. Taken together, Dr. Bhave's mentors collectively represent over 75 years of experience in career development, mentoring, and scientific investigation. Candidate Dr. Gautam (Jay) Bhave is currently an Instructor in the Division of Nephrology at Vanderbilt with >80% of his time dedicated to bench research. He completed his M.D. and Ph.D. degrees at Baylor College of Medicine with High Honors and clinical training at Johns Hopkins Hospital and Vanderbilt. His graduate work examined sensitization of sensory neurons with tissue injury and inflammation and garnered high-profile, first author publications in Nature Neuroscience, Neuron, and PNAS. After clinical training, Dr. Bhave joined the lab of Dr. Billy Hudson as a research fellow and began investigating how a newly discovered sulfilimine (S=N) cross-link in the collagen IV protein network is formed. The work has reached fruition with the discovery of a catalyzing enzyme, peroxidasin, and a first author manuscript describing this work was recently published in Nature Chemical Biology accompanied by a commentary and cover illustration. Ultimately, Dr. Bhave hopes to translate his seminal biochemical discovery into an independent line of investigation examining the role of peroxidasin in matrix remodeling and expansion in renal disease.

描述（由申请人提供）：糖尿病肾病（DN）是美国终末期肾病的主要原因。糖尿病肾病的中枢紊乱是活性氧（ROS）的过度产生。超氧化物及其产物过氧化氢（H2O2）是DN中最受关注的ROS。我们最近发现，过氧化物酶，一种嵌入细胞外基质的蛋白质，将H2O2转化为高活性的次溴酸（HOBr）和次氯酸（HOCl），统称为次卤酸（HOX）。过氧化物酶使用HOX在基底膜胶原蛋白IV中形成新的硫亚胺键（S=N），这是过氧化物酶的第一个确定的功能。但是，在H2O2底物饱和的糖尿病环境中，过氧化物酶可能产生过量的HOX。HOX是高度反应性和氧化卤化的蛋白质，对细胞功能产生不利影响。在使用糖尿病肾病小鼠模型的初步工作中，我们发现肾小球和肾小管过氧化物酶表达和HOX介导的蛋白质损伤增加。基于这些发现，我们假设过氧化物酶通过产生毒性HOX来改变细胞功能并促进细胞外基质扩张，从而在糖尿病肾病中发挥致病作用。为了验证这一假设，目标1将确定过氧化物酶如何在基底膜内发挥作用，检查IV型胶原和过氧化物酶之间的相互作用。在目的2中，我们将研究过氧化物酶和HOX在高血糖诱导的肾细胞功能障碍和基质积累中的作用，作为糖尿病肾病的体外模型。使用创新的质谱，我们将确定与过氧化物酶上调相关的HOX介导的蛋白质卤化的特定位点。在目标3中，我们将直接测试过氧化物酶和HOX在一个细胞中的作用。 糖尿病肾病小鼠模型。基于目标2中的质谱研究，我们将在体内鉴定蛋白质卤化位点。此外，我们将阻断过氧化物酶，并确定干预是否改善糖尿病肾病。这项工作有望深入了解糖尿病肾病的发病机制，并将过氧化物酶作为潜在的治疗靶点。环境拟定研究将在范德比尔特大学医学中心肾脏科进行。该部门拥有14个NIH资助的实验室，每年NIH资助超过1300万美元，是美国仅有的9个NIH乔治奥布莱恩肾脏研究中心之一，从而为Bhave博士提供了充足的财务，行政和科学支持。巴韦博士主要由比利哈德逊博士指导，他是一位有着近40年NIH资助的知名科学家。他的专长在于基底膜和IV型胶原蛋白生物化学，他的开创性工作包括Goodpasture抗原的鉴定和吡哆胺作为糖基化抑制剂治疗糖尿病肾病的开发。Raymond Harris博士和Roy Zent博士也作为共同导师支持Bhave博士，帮助开发独立的研究路线和职业道路。Harris博士是肾脏科主任，他在糖尿病肾病模型方面的经验对拟议的工作至关重要。Zent博士领导着一个资金充足的研究肾脏发育和整合素的小组，并将在过氧化物酶和细胞基质相互作用的研究方面提供专业知识。总的来说，巴韦博士的导师们在职业发展、指导和科学研究方面拥有超过75年的经验。候选人Gautam（Jay）Bhave博士目前是范德比尔特肾脏科的讲师，他80%以上的时间致力于实验室研究。他完成了医学博士学位和博士她在贝勒医学院获得高级荣誉学位，并在约翰霍普金斯医院和范德比尔特接受临床培训。他的研究生工作研究了组织损伤和炎症对感觉神经元的致敏作用，并在Nature Neuroscience，Neuron和PNAS上获得了备受瞩目的第一作者出版物。经过临床培训后，Bhave博士加入了比利哈德逊博士的实验室，担任研究员，并开始研究IV型胶原蛋白网络中新发现的硫亚胺（S=N）交联如何形成。这项工作随着催化酶过氧化物酶的发现而取得成果，描述这项工作的第一作者手稿最近发表在《自然化学生物学》上，并附有评论和封面插图。最终，Bhave博士希望将他开创性的生物化学发现转化为一个独立的研究路线，研究过氧化物酶在肾脏疾病基质重塑和扩张中的作用。