ACTIVATED MACROPHAGE/MONOCYTE TGFB1-INDUCED UPREGULATION OF COLLAGEN SYNTHESIS

活化巨噬细胞/单核细胞 TGFB1 诱导胶原合成上调

• 批准号: 5206125
• 负责人: ANITA C GILLIAM
• 金额: --
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/5206125
• 关键词: antibody specificity; autoantibody; autoimmune disorder; bone marrow transplantation; collagen; disease /disorder model; fibroblasts; fibrosis; gene expression; graft versus host disease; histopathology; immunoprecipitation; in situ hybridization; laboratory mouse; macrophage; monocyte; northern blottings; phenotype; protein biosynthesis; scleroderma; sclerosis; skin; transforming growth factors; ultrasonography; western blottings

Systemic sclerosis/scleroderma is a chronic autoimmune disease of unknown etiology, characterized by the excessive deposition of collagen in viscerae and skin, altered cell-mediated immunity, and the production of autoantibodies. Existing murine models for scleroderma are limited in their usefulness for study of the complex immunologic abnormalities, and human disease is difficult to study because the early changes are subtle and diagnosis is often delayed. Evidence from in vitro and in vivo work on pulmonary lesions in human scleroderma suggest that TGFbeta1 produced by infiltrating monocytes is a potent stimulus for collagen gene upregulation by fibroblasts leading to fibrosis. A murine sclerodermatous graft-versus- host disease (GVHD) model (C57BL/6J to LP/J) in which animals develop GVHD, skin thickening and autoantibodies after bone marrow transplantation across minor histocompatibility loci (H-2b) provides the ideal opportunity to study these events in an intact organism. Control mice receiving the reciprocal bone marrow transplantation LP/6 to C57BL/6J develop GVHD and dermal mononuclear cell infiltrates, but do not develop the skin thickening. Aim I of this proposal will characterize fully the sclerodermatous GVHD mice and confirm their usefulness as a model for human scleroderma. Disease progression will be correlated with histologic, biochemical and immunologic parameters by measuring dermal thickness using ultrasonography and physical measurements of intact skin and histologic sections, assaying autoantibody production by antinuclear antibody tests, immunophenotyping of the dermal mononuclear infIltrating cells, and quantifying dermal collagen gene expression by northern blot analysis of total RNA prepared from dermis at time points after transplantation in sclerodermatous and control animals. Aim II tests the central role of TGFbeta1-producing monocytes in causing collagen gene upregulation leading to skin fibrosis. Immunophenotyping and in situ hybridization using TGFbeta1 and pro-alpha(I)collagen probes to demonstrate co-localization of TGFbeta1-producing monocytes and collagen-producing fibroblasts in early skin lesions is the goal of this proposed work. Finally, the model provides a system in which variables can be manipulated to test the hypothesis that monocyte TGFbeta1 production is critical to initiation and progression of fibrosis, a logical extension of the work proposed here. Developing the murine model, confirming its validity for scleroderma, and identifying major early immunologic events in scleroderma will provide a means to test innovative immunotherapies in vivo.

系统性硬化/硬皮病是一种未知的慢性自身免疫性疾病 病因，以胶原过度沉积为特征 内脏和皮肤，改变细胞介导的免疫和产生 自身抗体。现有的硬皮病小鼠模型仅限于 它们在研究复杂的免疫异常方面的作用，以及 人类疾病很难研究，因为早期的变化很微妙 而且诊断往往会被推迟。来自体外和体内研究的证据 硬皮病患者的肺部病变提示TGFbeta1是由 渗透的单核细胞是胶原基因上调的有力刺激因素 通过成纤维细胞导致纤维化。一种小鼠硬皮病移植物抗- 动物发育的宿主病(GVHD)模型(C57BL/6J至Lp/J) 骨髓移植后移植物抗宿主病、皮肤增厚和自身抗体 跨次要组织相容性基因座(H-2b)提供了理想的机会 在一个完整的生物体中研究这些事件。对照组小鼠接受了 Lp/6至C57BL/6J双向骨髓移植发展为GVHD和 真皮单个核细胞会渗入，但不会发展成皮肤 变厚了。这项提案的目标一将充分说明 硬皮病型移植物抗宿主病小鼠的实验研究 人类硬皮病。疾病的进展将与组织学相关， 通过测量真皮厚度获得生化和免疫学参数 完整皮肤和组织学的超声检查和物理测量 切片，通过抗核抗体试验检测自身抗体的产生， 真皮单个核细胞的免疫表型，以及 Northern印迹分析法定量检测真皮胶原基因的表达 从真皮移植后不同时间点提取总RNA 硬皮病和对照动物。AIM II测试了 转化生长因子β1产生的单核细胞在诱导胶原基因上调中的作用 到皮肤纤维化。免疫表型和原位杂交法 TGFbeta1和前α(I)胶原蛋白探针共定位研究 早期分泌转化生长因子β1的单核细胞和胶原生成的成纤维细胞 皮肤损伤是这项拟议工作的目标。最后，该模型 提供了一个系统，在该系统中可以操作变量以测试 假设单核细胞产生TGFbeta1对启动和 纤维化的进展，这是这里提出的工作的合理延伸。 发展小鼠模型，证实其对硬皮病的有效性，并 确定硬皮病的主要早期免疫学事件将提供 在体内测试创新免疫疗法的手段。