Immune mechanisms that lead to irreversible scleroderma

描述（由申请人提供）： 硬皮病是一种慢性自身免疫性疾病，其特征在于由于成纤维细胞上调胶原蛋白的合成而导致器官和皮肤纤维化。迄今为止，对于硬皮病没有有效的治疗方法。我们正在使用小鼠硬皮病移植物抗宿主病（Scl GVHD）模型早期硬皮病，这可能是更适合治疗。在该模型中，我们可以在患有Scl GVHD的小鼠中在骨髓移植后21 - 28天内产生可测量的（厚达40%）皮肤增厚。我们和其他人已经表明，在硬皮病和早期炎性Scl GVHD中，趋化因子巨噬细胞趋化蛋白-1（MCP-1）、浸润性单核细胞/巨噬细胞细胞群体和细胞因子转化生长因子-β TGF-β是主要参与者。我们还通过早期体内施用TGF-β的特异性抑制剂潜伏相关肽（LTP）来预防小鼠Scl GVHD。在后期纤维化疾病中，炎症消退，并且成纤维细胞被认为具有永久改变的不受调节的胶原蛋白合成表型。从早期可逆性炎性疾病到后来的非炎性纤维化疾病的转变的触发因素尚不清楚。我们推测，在早期炎症性纤维化的独特的皮肤环境涉及免疫细胞和成纤维细胞之间的串扰。关键的成纤维细胞信号是免疫细胞归巢到皮肤并被激活所必需的，并且关键的免疫信号是产生不可逆的成纤维细胞表型所必需的。我们计划在以下研究中检查免疫细胞和成纤维细胞之间的串扰，这些研究集中在皮肤树突状细胞作为免疫应答的启动者、单核细胞/巨噬细胞和MCP-1。目的1：免疫细胞研究。a.什么时候皮肤纤维化不再能预防或逆转？这将在临床上建立"可逆"与"不可逆"疾病。B。刺激（Fit3配体）或抑制（CTLA41g）树突状细胞的干预措施对启动免疫应答至关重要，其作用是什么？C.单核细胞/巨噬细胞干预的影响是什么？骨髓移植后，我们是否还能用去除巨噬细胞的骨髓或体内去除巨噬细胞来产生Scl GVHD？我们能在MCP-1敲除小鼠中产生Scl GVHD吗？Aim II.成纤维细胞研究。a.在早期炎症性Scl GVHD中，皮肤成纤维细胞是否分泌免疫调节分子（特别是MCP-1）？我们能否在体外用干扰RNA（RNAi）抑制成纤维细胞MCP-1并阻断免疫细胞的激活和成纤维细胞表型的改变？B。什么样的免疫触发因素与成纤维细胞过度和持续分泌胶原蛋白有关？SMAD失调是一个严重事件吗？抗凋亡细胞克隆能解释不可逆的成纤维细胞表型吗？Scl GVHD中产生的肌成纤维细胞数量增加是否标志着不可逆纤维化的发生？ 我们是为数不多的使用这种有价值的硬皮病模型的实验室之一。我们在皮肤免疫生物学、树突状细胞和单核细胞/巨噬细胞生物学方面拥有专业知识。我们非常适合开展这个项目，以研究免疫细胞和纤维化疾病中成纤维细胞之间的相互作用，这是硬皮病研究中一个全新的令人兴奋的研究领域。为早期硬皮病开发更有效的诊断工具和免疫调节疗法是本研究的最终目标。

DESCRIPTION (provided by applicant): Scleroderma is a chronic autoimmune disease characterized by fibrosis of organs and skin, due to upregulated synthesis of collagen by fibroblasts. There is no effective treatment for scleroderma to date. We are using murine sclerodermatous graft versus host disease (Scl GVHD) to model early scleroderma, which may be more amenable to therapy. In this model, we can generate measurable (up to 40% thicker) skin thickening within 21-28 days post bone marrow transplantation in mice with Scl GVHD. We and others have shown that in scleroderma and in early inflammatory Scl GVHD, the chemokine macrophage chemotactic protein-1 (MCP-1), an infiltrating monocyte/macrophage cell population and the cytokine transforming growth factor-beta TGF-beta are major players. We have also prevented murine Scl GVHD with early administration of a specific inhibitor of TGF-beta, latency-associated peptide (LAP) in vivo. In later fibrosing disease, inflammation subsides, and the fibroblasts are thought to have a permanently altered phenotype of unregulated collagen synthesis. The triggers for the switch from early reversible inflammatory disease and later noninflammatory fibrotic disease are not known. We hypothesize that the unique cutaneous environment in early inflammatory fibrosis involves cross talk between immune cells and fibroblasts. Critical fibroblast signals are required for immune cells to home to skin and become activated, and critical immune signals are required to produce an irreversible fibroblast phenotype. We plan to examine the cross talk between immune cells and fibroblasts in the following studies that are focused on cutaneous dendritic cells as initiators of the immune response, monocyte/macrophages and MCP-1. Aim 1: Immune cell studies. A. When can LAP no longer prevent or reverse skin fibrosis? This will establish "reversible" versus " irreversible" disease clinically. B. What are the effects of interventions stimulating (Fit3 ligand) or inhibiting (CTLA41g) dendritic cells, which are essential to initiate an immune response? C. What are the effects of monocyte/ macrophage interventions? Can we still generate Scl GVHD with macrophage-depleted bone marrow or by depleting macrophages in vivo after BMT? Can we generate Scl GVHD in MCP-1 knockout mice? Aim II. Fibroblast studies. A. Do cutaneous fibroblasts secrete immunomodulatory molecules (particularly MCP-1) in early inflammatory Scl GVHD? Can we inhibit fibroblast MCP-1 with interference RNA (RNAi) and block activation of immune cells and an altered fibroblast phenotype in vitro? B. What immunologic triggers are related to the excessive and persistent secretion of collagen by fibroblasts? Is SMAD dysregulation a critical event? Do clones of cells resistant to apoptosis explain the irreversible fibroblast phenotype? Are increased numbers of myofibroblasts generated in Scl GVHD that signal the onset of irreversible fibrosis? We are one of the few laboratories using this valuable model for scleroderma. We have the expertise in cutaneous immunobiology, and in dendritic cell and monocyte/macrophage biology. We are ideally suited to carry out this project to examine the cross talk between immune cells and fibroblasts in fibrosing disease, an entirely new and exciting area of research in scleroderma research. Developing more effective diagnostic tools and immunomodulatory therapies for early scleroderma is the ultimate goal of this research.