Molecular Determinants of Kidney Podocyte Architecture in Health, Injury, and Recovery

健康、损伤和恢复中肾足细胞结构的分子决定因素

基本信息

批准号：
10522754
负责人：
Hani Suleiman
金额：
$ 34.65万
依托单位：
WASHINGTON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-07-05 至 2026-04-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10522754
关键词：
3-Dimensional Actin-Binding Protein Actins Actomyosin Affect Appearance Architecture Area Biology Cell Culture Techniques Cell Shape Cells Culture Techniques Cytoskeleton Data Diabetic Nephropathy Disease Electron Microscopy Embryo Epithelial Cells Equilibrium Evolution Extracellular Matrix Focal Segmental Glomerulosclerosis Foot Process Genes Genetic Models Glomerular Capillary Goals Guanosine Triphosphate Phosphohydrolases Health Homeostasis Hydrogels Image Imaging Techniques Imaging technology Injury Isoactin Kidney Kidney Diseases Kidney Failure Kidney Glomerulus Knockout Mice Labyrinth Lead Light Link Machine Learning Mechanics Membrane Methods Microfilaments Microscopy Modeling Molecular Monomeric GTP-Binding Proteins Morphology Mus Nature Nonmuscle Myosin Type IIA Optics Pathway interactions Physical condensation Play Process Protein Isoforms Recovery Regulation Renal function Renal glomerular disease Resolution Role Sarcomeres Spatial Distribution Specific qualifier value Structure System Techniques Testing Thick Three-Dimensional Imaging Tropomyosin Western Blotting Work base beta Actin biological systems biophysical properties blood filter blood filtration cell type design gamma Actin genetic variant glomerular basement membrane glomerular filtration imaging approach injured injury recovery mouse genetics mouse model novel novel therapeutic intervention podocyte reconstruction response to injury rho rho GTP-Binding Proteins slit diaphragm transcriptome sequencing

项目摘要

Modified Project Summary/Abstract Section The podocytopathies are a group of glomerular diseases that affect the kidney’s ability to filter the blood and often lead to kidney failure. Healthy podocytes cover the glomerular capillaries with thousands of extensions called foot processes that interdigitate with one another and maintain their elaborate cell shape by tightly regulating their actin cytoskeleton. Podocytes respond to insults in a typical fashion by undergoing foot process effacement, a dramatic shift in podocyte morphology and the disappearance of the intricate foot processes, which often associates with the “actin mat”, an actin condensation at the bottom of the effaced areas. We recently used super-resolution imaging to study the podocyte actin cytoskeleton in 3D in both healthy and diseased conditions. We showed that healthy podocyte foot processes contain non-contractile actin cables, while contractile cables are maintained high in the cell bodies. In contrast, injured podocytes appear to have contractile actin cables in effacement areas juxtaposed to the glomerular basement membrane (GBM), indicating a shift in the spatial distribution of actin cables after injury. The overall goal of this proposal is to define the molecular mechanisms that regulate the various types of actin cables in podocytes and the nature of the changes that cause the contractile actin cables in the cell body to shift towards the effaced areas adjacent to the GBM after injury. In Aim 1, we will investigate the roles of the two isoactins, beta and gamma actin, in podocyte pathobiology. Podocytes express high levels of these almost-identical evolutionally-conserved isoactins. While beta actin in non-muscle cells is considered the main isoactin, as evident from the embryonic lethality when inactivated, the role of gamma actin is still elusive. we will use various kidney disease mouse models, including the gamma-actin knockout mouse, to answer some fundamental questions about the role of the two isoactins in podocyte biology. Furthermore, we will utilize a novel technique to study primary podocytes as they spread out of isolated kidney glomeruli onto a substrate-micropatterned hydrogel. This approach will allow us to study the dynamic changes in the actin cytoskeleton in injured podocytes and will shed more light on the fate of the actin mats in effaced podocytes. It will help us identifying the role of Rho small GTPases and its downstream effectors, formins, in the actin mat formation. In Aim 2, we will study the tropomyosin isoform composition in podocytes and their roles in specifying the spatial distribution of different types of actin cables in the kidney podocytes. We hypothesize that changes in tropomyosin composition in injured podocytes causes the ectopic appearance of contractile actin cables in the effaced areas, and this, in turn, is regulated by different formins. Understanding how tropomyosins regulate the various types of actin cables could provide the missing link to podocyte foot process effacement. Our goal is to expand our understanding of the molecular mechanisms that regulate the composition and dynamics of the actin cytoskeleton, a step that will help us in designing novel therapeutic approaches to directly impact podocyte foot process architecture and help cure kidney glomerular diseases.

修改后的项目摘要/摘要部分足细胞病是一组肾小球疾病，影响肾脏过滤血液的能力，并经常导致肾衰竭。健康的足细胞覆盖肾小球毛细血管，有数千个称为足突的延伸，这些足突相互交错并紧密地维持其复杂的细胞形状。调节足细胞的肌动蛋白细胞骨架，以典型的方式通过足突消失、足细胞形态的巨大变化和复杂足突的消失来响应损伤，它通常与“肌动蛋白垫”相关，即被擦除区域底部的肌动蛋白凝结物。我们最近使用超分辨率成像来研究健康和患病条件下的足细胞肌动蛋白细胞骨架。含有非收缩性肌动蛋白线，而收缩性线在细胞体中保持较高水平，相反，受损的足细胞似乎在消失区域具有收缩性肌动蛋白线。与肾小球基底膜（GBM）并列，表明损伤后肌动蛋白线的空间分布发生变化。该提案的总体目标是定义调节足细胞中各种类型肌动蛋白线的分子机制以及足细胞的性质。在目标 1 中，我们将研究两种同肌动蛋白 β 和 γ 的作用。肌动蛋白，在足细胞病理学中，足细胞表达高水平的这些几乎相同的进化保守的同肌动蛋白，虽然非肌肉细胞中的β肌动蛋白被认为是主要的同肌动蛋白，从失活时的胚胎致死性可以明显看出，但γ肌动蛋白的作用仍然是。我们将使用各种肾脏疾病小鼠模型，包括γ-肌动蛋白敲除小鼠，来回答有关两种同肌动蛋白在足细胞中的作用的一些基本问题。此外，我们将利用一种新技术来研究原代足细胞从分离的肾小球扩散到基质微图案水凝胶上时的情况，这种方法将使我们能够研究受损足细胞中肌动蛋白细胞骨架的动态变化，并会脱落更多。阐明消失的足细胞中肌动蛋白垫的命运将有助于我们确定 Rho 小 GTP 酶及其下游效应子福尔明在肌动蛋白垫形成中的作用。 2，我们将研究足细胞中的原肌球蛋白亚型组成及其在指定肾足细胞中不同类型肌动蛋白电缆的空间分布中的作用，而这反过来又受到不同福明的调节，了解原肌球蛋白如何调节。各种类型的肌动蛋白电缆可以提供我们的目标是扩大我们对调节肌动蛋白细胞骨架的组成和动力学的分子机制的理解，这将有助于我们设计直接影响足细胞足突结构的新治疗方法。治疗肾脏肾小球疾病。