Glomerular Capillaries-- Normal and Pathologic

肾小球毛细血管——正常和病理

基本信息

批准号：
8792268
负责人：
MARILYN Gist FARQUHAR
金额：
$ 52.4万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN DIEGO
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-06-05 至 2017-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8792268
关键词：
Adriamycin PFS Affect Animals Apoptosis Binding Biochemical Biological Assay Cell Survival Cells Complex Data Development Diabetic Nephropathy Disease Fluorescence Focal Adhesions Focal glomerulosclerosis Glomerular Capillary Glomerulonephritis Goals Grant Image In Situ Injury Instruction Intercellular Junctions Kidney Knock-out Knockout Mice Life Ligation Mediating Molecular Mus Nephrosis Pathologic Permeability Phenotype Play Principal Investigator Process Progress Reports Proteins Proteinuria Proteomics Renal glomerular disease Research Role Signal Pathway Signal Transduction Testing Vascular Endothelial Growth Factor Receptor Vascular Endothelial Growth Factor Receptor-2 Vascular Endothelial Growth Factors Vinculin Work absorption cellular imaging db/db mouse glomerular filtration insight mutant nephrin novel podocyte research study response slit diaphragm

项目摘要

During the last 39 years under this grant our research has been directed toward understanding the cellular and molecular mechanisms of glornerular permeability and protein absorption as well as the derangements in these processes that occur in glomerular diseases. The experiments proposed in this application represent a direct continuation of our ongoing work on characterization of slit diaphragms and the role of GIV in podocyte function and survival after podocyte injury. We will focus on three specific aims: Specific Aim # 1 : To characterize and determine the functions of novel, putative slit diaphragm components identified by MS and quantitative organellar proteomics in slit diaphragm fractions. We will verify their junctional localization and determine their binding partners and functions in podocytes. Specific Aim #2: To investigate the role of GIV in mediating Akt signaling and cell survival in passive Heymann nephritis, focal glomerulosclerosis, and diabetic nephropathy. In PAN nephrosis GIV is upregulated, assembles a VEGFR2/GIV/Gal3 complex, and functions to maintain podocyte survival. We will determine if GIV plays a similar role in other diseases associated with podocyte injury. Specific Aim #3: To further investigate the mechanisms and spatial-temporal aspects of GIV's effects on podocytes. Our preliminary data indicate that GIV might function at focal adhesions (FA) and/or at slit diaphragms as it colocalizes with vinculin and VEGFR2 at FA and it binds CD2AP. We will investigate the dynamics ofthe interactions between GIV and VEGFR2 and CD2AP by live cell imaging, use proximity ligation assays and superresolution IF imaging (STORM) to determine where interaction occurs, and test whether GIV depletion or expression of GIV mutants affect FA or junction formation. These studies can be expected to provide novel insights into understanding ofthe cellular and molecular mechanisms of glomerular filtration and glomerular injury and their alterations in glomerular diseases associated with proteinuria. RELEVANCE (See instructions): Podocyte injury is the initiating cause of many glomerular diseases. The studies planned will provide key insights into the signaling neworks that regulate podocyte organization, functions and survival in response to glomerular injury in diseases associated with proteinuria.

在过去 39 年里，我们的研究一直致力于了解肾小球通透性和蛋白质吸收的细胞和分子机制以及肾小球疾病中发生的这些过程的紊乱。本文提出的实验应用代表了我们正在进行的狭缝隔膜表征工作的直接延续 GIV 在足细胞功能和足细胞损伤后存活中的作用。我们将重点关注三个具体目标：具体目标#1：表征和确定新颖的、假定的狭缝光阑的功能通过 MS 和定量细胞器蛋白质组学鉴定狭缝隔膜组分中的成分。我们将验证它们的连接定位并确定它们在足细胞中的结合伙伴和功能。具体目标 #2：研究 GIV 在介导 Akt 信号传导和被动细胞存活中的作用海曼肾炎、局灶性肾小球硬化和糖尿病肾病。在 PAN 肾病中，GIV 是上调，组装 VEGFR2/GIV/Gal3 复合物，并发挥维持足细胞存活的作用。我们将确定 GIV 是否在与足细胞损伤相关的其他疾病中发挥类似作用。具体目标#3：进一步研究 GIV 影响的机制和时空方面足细胞。我们的初步数据表明 GIV 可能在粘着斑 (FA) 和/或狭缝处起作用隔膜，因为它与纽蛋白和 VEGFR2 在 FA 共定位并结合 CD2AP。我们将调查通过活细胞成像观察 GIV 与 VEGFR2 和 CD2AP 之间相互作用的动态，使用接近度连接测定和超分辨率 IF 成像 (STORM) 以确定相互作用发生的位置并进行测试 GIV 缺失或 GIV 突变体的表达是否影响 FA 或连接形成。这些研究有望为理解细胞和分子生物学提供新的见解。肾小球滤过和肾小球损伤的机制及其在肾小球疾病中的改变与蛋白尿有关。相关性（参见说明）：足细胞损伤是许多肾小球疾病的始发原因。计划的研究将提供关键深入了解调节足细胞组织、功能和生存的信号网络与蛋白尿相关的疾病中的肾小球损伤。