Mechanisms of glomerular disesase progression

肾小球疾病进展的机制

基本信息

批准号：
8071219
负责人：
JEFFREY R SCHELLING
金额：
$ 32.7万
依托单位：
CASE WESTERN RESERVE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-07-01 至 2013-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8071219
关键词：
Adhesions Albuminuria Antibodies Azotemia Binding Biological Assay Blood Urea Nitrogen Brain Breeding Cell Adhesion Cell Culture Techniques Cell Differentiation process Cell Line Cell membrane Cell physiology Cells Characteristics Chimeric Proteins Chinese Hamster Ovary Cell Chronic Kidney Failure Cicatrix Coculture Techniques Collagen Collagen Type IV Complex Confocal Microscopy Cytoplasmic Tail Cytosol Data Defect Disease Docking Dominant-Negative Mutation ECM receptor Embryo Exhibits Extracellular Matrix Family Focal Adhesions GTP-Binding Proteins Genetic Glomerular Capillary Glomerular Mesangial Cell Goals Guanine Nucleotide Dissociation Inhibitors Health Histology Human In Vitro Incubated Injury Integrins Kidney Kidney Diseases Knockout Mice Lead Ligands Ligation Lysine MMP14 gene Maps Metalloproteases Modeling Morphology Mus Myofibroblast Nephrectomy Pathologic Pathway interactions Peptides Phenotype Physiological Placenta Plastics Precipitation Rattus Recruitment Activity Renal glomerular disease Reporter Role Signal Pathway Signal Transduction Smooth Muscle Actin Staining Method Technology Testing Tissues Transmission Electron Microscopy Vitronectin angiogenesis base cell transformation disease phenotype glomerular basement membrane glomerular capillary endothelium glomerulosclerosis in vivo interstitial laminin-10 loss of function mesangial cell migration mouse model mutant nidogen-1 podocyte premature prevent protein activation receptor research study rho working group

项目摘要

DESCRIPTION (provided by applicant): Integrins are extracellular matrix receptors, which regulate vital cell functions. The ¿8 subunit partners with alpha-v, and expression is restricted to kidney, brain, and placenta. ¿8 expression is localized to mesangial cells (MCs) in vivo and in vitro. Itg¿8-/- mice develop mild renal disease, but the phenotype is incompletely defined due to premature lethality. Kidney ¿8 expression is decreased in mouse models of glomerular disease, and MCs derived from Itg¿8-/- mice develop myofibroblast features. Of known ¿8 ligands, only latency-associated peptide (LAP), which combines with TGF¿ to form the small latent complex (SLC), to maintain TGF¿ in an inactive (latent) state, is detected in kidney. SLC binding to ¿8 can lead to TGF¿ activation through MT1-MMP cleavage of LAP, though MT1-MMP is not expressed in glomeruli, but can be induced in disease states. We show that LAP is expressed in MCs, and stimulates ¿8-dependent lamellopodia and migration, consistent with Rac1 activation. As a screen for MC ¿8 ligands, we show that matrix secreted by podocytes is superior to mesangial matrix for ¿8-dependent cell adhesion - the first demonstration of any ¿8 ligand supporting stable adhesions. We have shown that ¿8 interacts with Rho GDI. The major function of GDI is to sequester Rho family G-proteins in cytosol, which prevents G-protein activation or association with plasma membrane effectors. We show that ¿8 clustering enhances ¿8-GDI interaction and Rac1 activation in MCs, whereas Itg¿8-/- and Gdi-/- MCs exhibit myofibroblast features, such as alpha-smooth muscle actin (SMA) assembly and RhoA activation, as well as Rac1 suppression. We hypothesize that latent TGF¿ ligation with MC ¿8 recruits GDI-bound Rac1 to the ¿8 cytoplasmic tail. The ¿8-GDI interaction dissociates Rac1 from GDI, which facilitates Rac1 activation to suppress MC myofibroblast differentiation. In pathologic states, MC ¿8 expression is decreased, resulting in loss of LAP stimulation, cytoplasmic retention of Rac1 by GDI, and impaired MC binding to GBM. The net effect is that MCs transform into myofibroblasts with altered surveillance of the glomerular capillary endothelium. Specific aims: (1) In vivo characterization of ¿8 function using mice with MC-targeted ¿8 deletion and a rat model of acquired glomerulosclerosis. (2) Characterize the MC ¿8 ligand. (3) Test whether ¿8 functions as a RacGDI displacement factor to suppress MC myofibroblast differentiation. PUBLIC HEALTH RELEVANCE Over 20 million people in the U.S. suffer from chronic kidney disease, which usually starts with injury to the glomerulus, the filtering unit of the kidney. Our proposal aims to define the role of a specific receptor, the beta-8 integrin, which is expressed on the glomerular mesangial cell, in regulating kidney disease. This integrin is found in the brain, kidney and placenta and has been well-characterized only in brain. We are currently the only group working on kidney beta-8 integrin, and we provide preliminary data that beta-8 loss of function leads to kidney disease. To achieve the goals of this proposal, we are using mouse models, in which the beta-8 integrin has been selectively deleted from mesangial cells, to avoid confounding effects from brain. In addition, we are employing mesangial cell culture models, which can be more easily manipulated, upon completion of this project; we hope to provide new information about the kidney beta-8 integrin, which could lead to new therapies for chronic kidney diseases.

描述（由适用提供）：整联蛋白是细胞外基质受体，可调节重要细胞功能。 „与α-V的8个亚基合作伙伴，表达仅限于肾脏，大脑和斑点。 8表达位于体内和体外的弥赛亚细胞（MCS）。 ITG？8 - / - 小鼠会出现轻度的肾脏疾病，但由于过早的致死性，表型未完全定义。在肾小球疾病的小鼠模型中，肾脏8的表达降低，源自ITG¿8 - / - 小鼠的MC发展了肌纤维细胞特征。在肾脏中检测到已知的8种配体，仅与TGF与TGF组合形成小型潜在复合物（SLC），以在肾脏中检测到不活跃（潜伏）状态的小型潜伏复合物（SLC），以将TGF固定在肾脏中。 SLC与8的结合可以通过MT1-MMP裂解导致TGF lap的激活，尽管MT1-MMP在Glomerulli中未表达，但可以在疾病状态下诱导。我们表明，膝盖是在MC中表达的，并刺激了与Rac1激活一致的8依赖性薄膜植物和迁移。作为8种配体的屏幕，我们表明，足细胞分泌的基质优于Messianial矩阵，用于8依赖性细胞粘附 - 第一个证明了任何8种支持稳定辅助的配体。我们已经证明„ 8与Rho GDI相互作用。 GDI的主要功能是隔离细胞质中的Rho家族G蛋白，从而防止G蛋白激活或与质膜效应相关。我们表明„ 8聚类增强功能`8-GDI相互作用和RAC1激活在MC中，而ITG？8 - / - 和GDI - / - MCS暴露了肌纤维细胞功能，例如Alpha-Smooth Muscle Muscle Muscle肌动蛋白（SMA）组件（SMA）组件和Rhoa激活以及RAC1抑制。我们假设与MC的潜在TGF研观8招募了GDI结合的Rac1至8个细胞质尾巴。 8-GDI相互作用将Rac1与GDI分离，Rac1激活了Rac1激活以抑制MC肌纤维细胞的分化。在病理状态下，MC»8表达降低，导致膝上刺激的损失，GDI对Rac1的细胞质保留以及MC与GBM的结合受损。净效应是，MCS随着肾小球毛细管内皮的监测改变为肌纤维细胞。具体目的：（1）使用具有MC靶向8缺失的小鼠和获得的肾小球硬化的大鼠模型的小鼠对8功能的体内表征。（2）表征MC€8配体。（3）测试„ 8是否充当RACGDI位移因子来抑制MC肌纤维细胞分化。在美国，超过2000万人患有公共卫生相关性，患有慢性肾脏疾病，通常始于肾脏的过滤单位Glomerulus受伤。我们的建议旨在定义特定接收器的beta-8整联蛋白的作用，该整合素在肾小球肾小球细胞上表达，在控制肾脏疾病中。该整联蛋白在大脑，肾脏和plapeta中发现，并且仅在大脑中得到很好的特征。目前，我们是唯一从事肾脏β-8整联蛋白的小组，我们提供了β-8功能丧失导致肾脏疾病的初步数据。为了实现该提案的目标，我们使用的是鼠标模型，其中β-8整合素已选择性地从弥赛亚细胞中删除，以避免大脑的混淆效果。此外，我们正在采用弥赛亚细胞培养模型，该模型可以在完成该项目后更容易操纵。我们希望提供有关肾脏β-8整合素蛋白的新信息，这可能会导致慢性肾脏疾病的新疗法。