Influence of extracellular matrix fibronectin on platelet derived growth factor signaling

细胞外基质纤连蛋白对血小板衍生生长因子信号传导的影响

• 批准号: 9121935
• 负责人: Christopher Farrar
• 金额: $ 2.91万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-01 至 2017-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9121935
• 关键词: Animal Tarsus; Atherosclerosis; Attenuated; Binding; Biological; Calcium; Cells; Chimeric Proteins; Clinical; Controlled Study; Data; Deposition; Development; Diabetic ulcer; Disease; Down-Regulation; EGF gene; Embryo; Engineering; Extracellular Matrix; Extracellular Matrix Degradation; Extracellular Matrix Proteins; Fibroblast Growth Factor; Fibroblasts; Fibronectins; Fibrosis; Fluo 4; Functional disorder; Goals; Granulation Tissue; Healed; Homeostasis; Human; Intracellular Signaling Proteins; Knockout Mice; Laboratories; Libraries; Lung; Malignant Neoplasms; Measures; Mediating; Mesenchymal; Methods; Mitogens; Mus; Myofibroblast; PRKCA gene; Pathway interactions; Peptides; Phenotype; Physiological; Platelet-Derived Growth Factor; Platelet-Derived Growth Factor beta Receptor; Process; Proliferating; Proteolysis; Proteolytic Processing; Pulmonary Fibrosis; Regulation; Role; Signal Pathway; Signal Transduction; Staging; System; Testing; Time; Tissues; Tyrosine; Up-Regulation; Wound Healing; attenuation; cancer type; cell growth regulation; cell motility; chronic wound; extracellular; healing; insight; novel; public health relevance; response; scaffold; success; tool; wound

 DESCRIPTION (provided by applicant): Platelet derived growth factor (PDGF) is the major mitogen for mesenchymal cells and is used clinically to promote diabetic ulcer healing. Clinical successes using PDGF to promote wound healing have been underwhelming and inconsistent. Fibronectin is a critical ECM protein that is secreted by fibroblasts in a protomeric form and assembled into a fibrillar matrix by a cell-mediated process termed fibronectin matrix assembly. Preliminary data presented in this proposal show that cell-assembled ECM fibronectin fibrils attenuate PDGF-induced intracellular calcium release. Intracellular calcium release was measured in Fluo-4- loaded fibronectin-null (FN-/-) mouse embryonic fibroblasts (MEFs), allowing for precise control over the amount of fibronectin to which cells were exposed and the initiation of fibronectin matrix assembly. Fibronectin- mediated attenuation of PDGF-induced calcium release represents a previously undescribed mechanism by which ECM fibronectin may downregulate PDGF signaling, and thus spatially and/or temporally control PDGF activity in healing wounds. Alternatively, several diseases including pulmonary fibrosis, atherosclerosis and some types of malignancies are characterized, in part, by an upregulation of PDGF activity. Thus, a fibronectin fragment that inhibits cell responses to PDGF may be beneficial in treating these conditions. The goal of this study is to determine the mechanism and consequences of ECM fibronectin-mediated regulation of cellular responses to PDGF in fibroblasts. In order to achieve the goals of this project, the following specific aims have been developed: (1) Localize the effect of ECM fibronectin on cell responsiveness to PDGF and develop small fibronectin peptides regulate cell responsiveness to PDGF, (2) determine the mechanism by which ECM fibronectin regulates fibroblast responses PDGF and (3) determine the effect of regulation of responses to PDGF by ECM fibronectin on cell phenotype. Important tools for these studies will include calcium analysis in our Fluo-4-loaded FN-/- MEF system, as well as our laboratory's extensive library of tools used to study fibronectin matrix assembly and expertise in developing novel fibronectin-derived fusion proteins. Experimental results will provide insight into how cell responses to PDGF are regulated during normal wound healing processes, how they may be dysregulated during pathological wound healing responses, and how specific fibronectin fragments may be used therapeutically to control endogenous PDGF activity.

 描述（由申请人提供）：血小板衍生生长因子 (PDGF) 是间充质细胞的主要有丝分裂原，在临床上用于促进糖尿病溃疡愈合，但使用 PDGF 促进伤口愈合的临床成功效果并不理想，而且纤维连接蛋白是一种关键的 ECM。由成纤维细胞以原体形式分泌的蛋白质，并通过称为纤连蛋白基质组装的细胞介导过程组装成纤维基质。该提案中提供的初步数据表明，细胞组装的 ECM 纤连蛋白原纤维可减弱 PDGF 诱导的细胞内钙释放。在加载 Fluo-4 的纤连蛋白缺失 (FN-/-) 小鼠胚胎成纤维细胞 (MEF) 中测量细胞内钙释放，从而允许。用于精确控制细胞暴露的纤连蛋白的量以及纤连蛋白基质组装的启动。 PDGF诱导的钙释放的机制代表了一种先前未描述的机制，通过该机制，ECM纤连蛋白可以下调PDGF信号传导，从而在空间和/或时间上控制伤口愈合中的PDGF活性。或者，包括肺纤维化、动脉粥样硬化和某些类型的恶性肿瘤在内的多种疾病的特征。因此，抑制细胞对 PDGF 反应的纤连蛋白片段可能有助于治疗这些病症。本研究的目的是确定其机制。 ECM 纤连蛋白介导的成纤维细胞对 PDGF 反应的调节的后果 为了实现该项目的目标，已经制定了以下具体目标：（1）定位 ECM 纤连蛋白对细胞对 PDGF 反应的影响并开发。小纤连蛋白肽调节细胞对 PDGF 的反应，(2) 确定 ECM 纤连蛋白调节成纤维细胞反应 PDGF 的机制，以及 (3) 确定调节对 PDGF 反应的作用PDGF by ECM 纤连蛋白对细胞表型的影响这些研究的重要工具将包括我们装载 Fluo-4 的 FN-/- MEF 系统中的钙分析，以及我们实验室用于研究纤连蛋白基质组装和开发专业知识的广泛工具库。实验结果将深入了解正常伤口愈合过程中细胞对 PDGF 的反应如何调节、病理性伤口愈合反应过程中细胞反应如何失调以及特定的纤连蛋白如何调节。片段可在治疗上用于控制内源性PDGF活性。