Role of Integrins in Bladder Urothelium

整合素在膀胱尿路上皮中的作用

基本信息

批准号：
8037784
负责人：
Warren G. Hill
金额：
$ 37.04万
依托单位：
BETH ISRAEL DEACONESS MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-03-15 至 2015-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8037784
关键词：
Abbreviations Ablation Actins Adaptor Signaling Protein Adenosine Affect Alleles Animals Apical Apoptosis Ataxia Attenuated Basement membrane Behavior Binding Biochemical Biological Assay Biotinylation Bladder Bladder Control Bladder Diseases Bladder Urothelium Cations Cell Adhesion Molecules Cell membrane Cell surface Cell-Cell Adhesion Cells Chemicals Complement Complex Confocal Microscopy Connective Tissue Cystitis Cytoskeleton Data Defect Diabetes Mellitus Electric Capacitance Electron Microscopy Enzymes Epithelial Epithelium Exhibits Exocytosis Extracellular Matrix Family Fill-It Focal Adhesion Kinase 1 Focal Adhesions Frequencies GAG Gene Glycosaminoglycans Hemidesmosomes Heterozygote Human Hydrostatic Pressure Hyperreflexia Immunoprecipitation Incontinence Integrin Binding Integrin Signaling Pathway Integrins Interstitial Cystitis Ion Channel Ions Israel Kinetics Knock-out Knockout Mice Knowledge Lamina Propria Ligands Link Measures Mechanics Mediating Mediator of activation protein Medical center Membrane Membrane Protein Traffic Membrane Proteins Molecular Monitor Mouse Strains Mus Nerve Fibers Nomenclature Output Overactive Bladder PTK2 gene Pain Participant Pathway interactions Phenotype Phosphate Buffer Phosphotransferases Play Prostaglandins Prostatic hypertrophy Proteins Protocols documentation Publishing Purinoceptor Quality of life Receptor Protein-Tyrosine Kinases Reflex action Regulation Relaxation Resistance Role Saline Second Messenger Systems Sensory Signal Pathway Signal Transduction Signaling Molecule Sodium Channel Spastic Sphincter Spinal cord injury Stimulus Stress Stretching Surface Symptoms Technology Testing Tight Junctions Transmission Electron Microscopy UPK2 gene Urethral sphincter Urine Urodynamics Urothelial Cell Urothelium Viscosity Work adhesion receptor afferent nerve apical membrane autocrine cell motility cell type epithelial Na+ channel extracellular in vivo inhibitor/antagonist insight integrin-linked kinase knockout animal novel therapeutics paracrine public health relevance receptor recombinase response second messenger sensor

项目摘要

DESCRIPTION: Integrins are a broad family of cell surface adhesion and signaling molecules which link the extracellular matrix (ECM) to the cytoskeleton of the cell. Integrins bind directly to ECM ligands on the extracellular surface and are linked to the actin cytoskeleton through multiple adaptor proteins and receptor tyrosine kinases on the plasma membrane. They play critical roles in all cell types and regulate cell migration, differentiation, proliferation and apoptosis. They are also force sensors and transduce mechanical stimuli into biochemical signals. This project explores the role of integrins and integrin-signaling pathways within the epithelium lining the bladder - the urothelium - where almost nothing is known of their function. Our primary hypothesis is that integrins represent the initial upstream mechanical sensor in the bladder and detect bladder filling through membrane stretch. In order to investigate this hypothesis we have created a conditional knockout of 21-integrin within the urothelium (21-cKO). Loss of 21-integrin knocks out all integrins within the umbrella and intermediate cells making the superficial layers of the epithelium integrin-null. Preliminary data indicates that the bladders of these mice release significantly less ATP into the lumen upon stretch, hyperactivate ion conductances upon stretch and exhibit detrusor hyperreflexia and detrusor-sphincter dyssynergia when measured by cystometry. These findings are strongly indicative of an obstructed bladder phenotype possibly due to inadequate urethral sphincter relaxation. Taken together these symptoms indicate a mechanical signaling deficit originating in the urothelium which leads to an obstructed bladder phenotype. An understanding of the signaling mechanisms employed by the bladder as it fills is essential in understanding many different bladder disorders, from overactive bladder to interstitial cystitis. Specific aim 1 of our study will examine the repertoire of integrin binding partners in umbrella cells in vivo and will explore the mechanisms by which integrins regulate ATP secretion. ATP has been show to be an extremely important autocrine and paracrine mediator in the bladder and signals through purinergic receptors on both the luminal surface and in suburothelial layers e.g. on afferent nerve fibers. Specific aim 2 will investigate how integrins interact with and regulate mechanosensory ion channels within the urothelium using selective inhibitors and ion substitution protocols to determine which ion channels regulated by integrins. The role of ion fluxes in regulating membrane trafficking and ATP release will also be studied using (i) capacitance changes in response to stretch to monitor membrane trafficking; and (ii) short circuit currents on bladders mounted in Ussing chambers. Specific aim 3 will use cystometry in the presence of various pharmacological modulators of ion channels and receptors to investigate at the molecular level how integrins regulate filling and voiding behavior. PUBLIC HEALTH RELEVANCE: There are large numbers of people who for many different reasons - spinal cord injury, diabetes, cystitis - suffer from disorders of the bladder which can broadly be characterized as overactive bladder. They suffer greatly from embarrassing and debilitating symptoms of incontinence, urgency, pain and frequency which profoundly affect quality of life. This project will provide new knowledge of the mechanisms by which the bladder senses filling and responds and in so doing may suggest new therapeutic options for bladder disease.

描述：整合素是一个广泛的细胞表面粘附和信号分子家族，它将细胞外基质 (ECM) 连接到细胞的细胞骨架。整合素直接与细胞外表面的 ECM 配体结合，并通过质膜上的多个接头蛋白和受体酪氨酸激酶与肌动蛋白细胞骨架连接。它们在所有细胞类型中发挥关键作用并调节细胞迁移、分化、增殖和凋亡。它们也是力传感器，并将机械刺激转换成生化信号。该项目探讨了膀胱内衬上皮（尿路上皮）内整合素和整合素信号传导通路的作用，但人们对它们的功能几乎一无所知。我们的主要假设是整合素代表膀胱中最初的上游机械传感器，并通过膜拉伸检测膀胱充盈。为了研究这一假设，我们在尿路上皮内创建了 21-整合素的条件敲除 (21-cKO)。 21-整合素的丢失会敲除伞状细胞和中间细胞内的所有整合素，从而使上皮表层整合素无效。初步数据表明，这些小鼠的膀胱在拉伸时释放到管腔中的 ATP 显着减少，拉伸时离子电导过度活跃，并且通过膀胱测压测量时表现出逼尿肌反射亢进和逼尿肌-括约肌协同失调。这些发现强烈表明膀胱梗阻表型可能是由于尿道括约肌松弛不足所致。总的来说，这些症状表明源自尿路上皮的机械信号传导缺陷，导致膀胱梗阻表型。了解膀胱充盈时所采用的信号传导机制对于了解从膀胱过度活动症到间质性膀胱炎等许多不同的膀胱疾病至关重要。我们研究的具体目标 1 将检查体内伞细胞中整合素结合伴侣的全部功能，并探索整合素调节 ATP 分泌的机制。 ATP 已被证明是膀胱中极其重要的自分泌和旁分泌介质，并通过管腔表面和尿路上皮下层（例如膀胱）上的嘌呤能受体发出信号。在传入神经纤维上。具体目标 2 将研究整合素如何与尿路上皮内的机械感觉离子通道相互作用并对其进行调节，使用选择性抑制剂和离子替代方案来确定哪些离子通道受整合素调节。离子通量在调节膜运输和 ATP 释放中的作用也将通过以下方式进行研究：(i) 响应于拉伸的电容变化以监测膜运输； (ii) 安装在使用室中的囊上的短路电流。具体目标 3 将在离子通道和受体的各种药理学调节剂存在的情况下使用膀胱测压法，在分子水平上研究整合素如何调节充盈和排尿行为。公众健康相关性：有很多人由于多种不同的原因（脊髓损伤、糖尿病、膀胱炎）而患有膀胱疾病，这些疾病大致可称为膀胱过度活动症。他们深受尿失禁、尿急、疼痛和尿频等令人尴尬和衰弱的症状的困扰，这些症状严重影响了生活质量。该项目将提供有关膀胱感知充盈和反应机制的新知识，从而可能为膀胱疾病提出新的治疗选择。