Fibroblast TGF-beta/Smad Signaling in Scleroderma

硬皮病中的成纤维细胞 TGF-β/Smad 信号转导

• 批准号: 6660301
• 负责人: John Varga
• 金额: $ 34.02万
• 依托单位: UNIVERSITY OF ILLINOIS AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-09-01 至 2007-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6660301
• 关键词: CD40 molecule; RNA; biological signal transduction; biopsy; cell line; cell proliferation; clinical research; collagen; fibroblasts; fibrosis; gene expression; gene targeting; genetically modified animals; human tissue; inflammation; laboratory mouse; microarray technology; northern blottings; polymerase chain reaction; protein structure function; scleroderma; tissue /cell culture; transcription factor; transfection /expression vector; transforming growth factors; western blottings

DESCRIPTION (provided by applicant): In systemic sclerosis (SSc), excessive accumulation of collagen leads to fibrosis and subsequent dysfunction of skin and internal organs. Transforming growth factor-beta (TGF-beta) plays a pivotal role in initiating and propagating fibrosis. Because of its key role in SSc, understanding how TGF-beta regulates fibroblast functions would clarify the pathogenesis of SSc and identify novel targets for therapeutic intervention. The recent discovery of the Smad family of intracellular TGF-beta signal transducers and the recognition of their major roles in multiple diseases, provides novel approaches for investigating the involvement of TGF-beta in pathogenesis of SSc. The proposed studies utilize a combination of in vitro and in vivo approaches in order to answer four important questions fundamental to understanding the role of the TGF-beta/Smad axis in SSc: What are fibrosis-related genes whose expression in human fibroblasts is modulated by Smad3? How is the Smad pathway integrated into the cellular signaling network of normal and SSc fibroblasts during inflammation and repair? Are intrinsic alterations in Smad signaling implicated in the SSc fibroblast phenotype? What is the functional role of Smad3 in the development of scleroderma in mice? Our hypothesis is that fibrosis in SSc is due to sustained activation of fibroblast gene expression, and that Smad3 plays a pivotal role in this process. The hypothesis will be evaluated in four interrelated aims. In Specific Aim 1, we will use DNA microarrays to profile genes whose expression is modulated by inducible expression of Smad3 in immortalized human fibroblasts; In Specific Aim 2, we will examine modulation of fibroblast TGF-beta/Smad signaling by inflammatory cells through CD4O. In Specific Aim 3, we will characterize the Smad pathway in SSc fibroblasts and tissues in order to identify intrinsic alterations in the regulation and/or function of the TGF-beta/Smad axis in SSc that may contribute to fibroblast activation. Observations derived from in vitro experiments cannot always accurately predict protein function in vivo. We hypothesize that Smad3 is an essential mediator of TGF-beta signaling, and mice lacking Smad3 have reduced susceptibility to pathological tissue fibrosis. To test this hypothesis directly, in Specific Aim 4 we will induce cutaneous fibrosis in Smad3-null mice in order to determine the role of Smad3 in this process. The results will clarify the role of Smad signal pathway in mediating the development of fibrosis in SSc, and reveal the feasibility of therapeutically targeting.

描述（由申请人提供）：在系统性硬化症（SSC）中，胶原蛋白的过度积累会导致纤维化和随后的皮肤和内脏功能障碍。转化生长因子β（TGF-β）在启动和传播纤维化中起关键作用。 由于其在SSC中的关键作用，因此了解TGF-beta如何调节成纤维细胞功能将阐明SSC的发病机理并确定治疗干预的新颖靶标。最近发现细胞内TGF-β信号传感器的SMAD家族及其在多种疾病中的主要作用的识别，提供了研究TGF-beta参与SSC发病机理的新方法。拟议的研究利用体外和体内方法的结合，以回答四个重要的问题，以了解TGF-beta/smad轴在SSC中的作用至关重要：纤维化相关的基因在人类成纤维细胞中的表达是什么？ 在炎症和修复过程中，如何将SMAD途径整合到正常和SSC成纤维细胞的细胞信号网络中？ SSC成纤维细胞表型中的SMAD信号传导的固有变化是否存在？ SMAD3在小鼠的硬皮病发展中的功能作用是什么？我们的假设是SSC中的纤维化是由于成纤维细胞基因表达的持续激活所致，并且Smad3在此过程中起关键作用。该假设将以四个相互关联的目的进行评估。在特定的目标1中，我们将使用DNA微阵列来介绍其表达的基因，其表达是由永生的人成纤维细胞中Smad3诱导的表达调节的。在特定目标2中，我们将检查通过CD4O通过炎症细胞对成纤维细胞TGF-β/SMAD信号的调节。在特定的目标3中，我们将表征SSC成纤维细胞和组织中的SMAD途径，以确定SSC中TGF-Beta/Smad轴调节和/或功能的固有变化，这可能会导致成纤维细胞激活。从体外实验得出的观察结果不能总是准确地预测体内蛋白质功能。我们假设SMAD3是TGF-β信号传导的重要介质，而缺乏SMAD3的小鼠对病理组织纤维化的敏感性降低。为了直接检验该假设，在特定的目标4中，我们将诱导SMAD3-NULL小鼠的皮肤纤维化，以确定SMAD3在此过程中的作用。结果将阐明SMAD信号途径在介导SSC纤维化发展中的作用，并揭示治疗靶向的可行性。