ENDOTHELIAL CELL AND MONOCYTE ACTIVATION IN A MURINE GVHD SCLERODERMA MODEL

系统性硬化症/硬皮病的早期事件，一种慢性纤维化 病因不明的自身免疫性疾病，知之甚少， 很难在患者身上进行研究。该疾病的特征在于 潜伏的临床进展，改变的细胞介导的免疫， 自身抗体的产生和胶原蛋白的过度沉积， 内脏和皮肤成纤维细胞。皮肤纤维化似乎发生在 首先是真皮血管周围，这表明触发事件局限于 人类硬皮病的血管周围假设：激活 真皮血管周围区室中募集的单核细胞是一种关键的 可以在小鼠中建模的硬皮病中的早期事件 硬皮病性移植物抗宿主病（GVHD）。当骨髓和 将来自B10.D2（H-2k）小鼠的脾细胞移植到致死的 照射的Balb/c小鼠（H-2k），动物发生轻度GVHD， 自身免疫性疾病和皮肤增厚，类似于人类 硬皮病接受同基因骨髓和脾脏的对照动物 细胞（Balb/c移植到Balb/c）没有显示这种变化。具体 目的一：确定单核细胞早期关键事件的发生顺序 在动物的皮肤微血管区室中的激活， 硬皮病GVHD，以便更好地了解病理生理学 硬皮病内皮细胞（ICAM-1）免疫组化染色 1、VCAM-1、E-选择素和血管性血友病因子）和单核细胞（Mac-1）。 1、Ia、VLA-4、CD16和iNOS）活化标志物的表达。 可检测的胶原蛋白mRNA上调，皮肤将识别这些 早期事件使用图像分析来定量密度， 浸润细胞的表型。具体目标二：使用流程 流式细胞术，关联来自Aim I的免疫染色结果以鉴定和 量化浸润动物皮肤的细胞群的表型 硬皮病性GVHD的患者。 浆内 TGF β 1和表面Mac-1的染色将解决这个问题： 产生TGF β的单核细胞浸润先于真皮纤维化？ 将硬皮病的这些早期事件联系起来， 到测试新型单核细胞定向免疫疗法的干预措施， 体内，然后转化为人类硬皮病的临床试验。

The early events in systemic sclerosis/scleroderma, a chronic fibrosing autoimmune disease of unknown etiology, are poorly understood and have been difficult to study in patients. The disease is characterized by insidious clinical progression, altered cell-mediated immunity, production of autoantibodies, and excessive deposition of collagen by fibroblasts in viscerae and skin. Cutaneous fibrosis appears to occur first around dermal vessels, suggesting triggering events localized to the perivasculature in human scleroderma. Hypothesis: activation of recruited monocytes in dermal perivascular compartments is a critical early event in scleroderma which can be modeled in murine sclerodermatous graft versus host disease (GVHD). When bone marrow and spleen cells from B10.D2 (H-2k) mice are transplanted into lethally irradiated Balb/c mice (H-2k), the animals develop mild GVHD, autoimmune disease, and skin thickening resembling that of human scleroderma. Control animals receiving syngeneic bone marrow and spleen cells (Balb/c transplanted to Balb/c) show no such changes. Specific Aim I: To determine the sequence of early critical events in monocyte activation in the cutaneous microvascular compartment of animals with sclerodermatous GVHD in order to better understand the pathophysiology of sclerodermatous disease. Immunostaining for endothelial cell (ICAM- 1, VCAM-1, E-selectin, and von Willebrand factor) and monocyte (Mac- 1,Ia, VLA-4, CD16, and iNOS) activation markers at time points before detectable collagen mRNA upregulation and skin will identify these early events. Image analysis is used to quantitate density and phenotypes of infiltrating cells. Specific Aim II: Using flow cytometry, correlate immunostaining results from Aim I to identify and quantify phenotypes of cell populations infiltrating skin of animals with sclerodermatous GVHD at early time points. Intracytoplasmic staining for TGFbeta1 and surface Mac-1 will address the question: does infiltration by TGFbeta-producing monocytes precede dermal fibrosis? Correlating these early events in sclerodermatous disease will point to interventions testing novel monocyte-directed immunotherapies in vivo, then translated into clinical trials for human scleroderma.