Role of TGF-beta and CTGF Signaling Transgenic Mouse Models of Scleroderma

硬皮病或系统性硬化症(SSC)是一种影响各种器官的结缔组织紊乱。 系统。这种疾病很复杂，其特点是胶原蛋白过度堆积和其他 皮肤和内脏中的细胞外基质成分。因为转化生长因子-β信号的增加 我们最近建立了一种新的小鼠模型，在该模型中，转化生长因子-β受体1 在出生后的成纤维细胞中被结构性激活(TBR1CA；Col1a2-Creer)。这些老鼠重述了 人类SSC的主要特征，表现为真皮明显而广泛的纤维化，较薄的表皮， 毛囊脱落，肺、肾小血管壁纤维性增厚。初级皮肤 这些小鼠的成纤维细胞显示下游转化生长因子-β靶标的表达增加，复制了 移植的SSc真皮成纤维细胞的生化表型特征。尤其是有一个明显的 结缔组织生长因子(CTGF)表达增加。由于CTGF的表达增加 被认为在疾病过程中发挥了关键作用，我们培育出了过度表达的转基因小鼠 成纤维细胞特异性启动子/增强子介导的结缔组织生长因子在成纤维细胞中的表达(L) 胶原蛋白基因。这些动物的毛发严重脱落。皮肤活检的初步组织学检查 表现为真皮明显的全身性纤维化，真皮增厚和炎性浸润物。 皮肤纤维化区。小鼠胚胎成纤维细胞来源的初步分析 转基因小鼠表现出I型胶原和TIMP-3的高表达。 我们建议对Col1a2-CTGF小鼠及其移植的皮肤成纤维细胞进行鉴定。至 了解成纤维细胞中CTGF表达增加导致纤维化疾病的机制， 我们还将对这些小鼠皮肤成纤维细胞的分子表型进行比较 TBR1CA；Col1a2-Creer小鼠和特定的人类硬皮病患者的那些。为了进一步 研究这些机制，我们建议减轻这两种皮肤成纤维细胞的纤维化表型 通过特定信号通路或药物抑制剂建立硬皮病转基因小鼠模型 SiRNA。我们还将尝试通过杂交在体内抑制这些转基因小鼠的纤维化表型 在这些小鼠中，Smads或整合素D6为零突变。最后，我们将研究 使TBR1CA；Col1a2-Creer小鼠对博莱霉素性肺纤维化的敏感性增加 Col1a2-CTGF小鼠是否也有类似的敏感性。这些研究应该会提供关于 转化生长因子-β和结缔组织生长因子在纤维化疾病中的相对重要性。

Scleroderma or systemic sclerosis (SSc) is a disorder of the connective tissues affecting various organ systems. The disease is complex and is characterized by excessive accumulation of collagen and other extracellular matrix components in the skin and internal organs. Because increased signaling by TGF-beta has been implicated in this disease we recently established a novel mouse model in which the TGF-beta Receptor1 is constitutively activated in fibroblasts post-natally (TBR1CA; Col1a2-CreER). These mice recapitulated the major features of human SSc, showing pronounced and generalized fibrosis of the dermis, thinner epidermis, loss of hair follicles, and fibrotic thickening of small blood vessel walls in lung and kidney. Primary skin fibroblasts of these mice showed elevated expression of downstream TGF-beta targets, reproducing the hallmark biochemical phenotype of explanted SSc dermal fibroblasts. In particular there was a marked increase in connective tissue growth factor (CTGF) expression. Since increased expression of CTGF has been implicated to play a key role in the disease process, we generated transgenic mice that over-express CTGF in fibroblasts (Col1a2-CTGF) by using a fibroblast-specific promoter/enhancer from the pro-D2(l) collagen gene. The animals exhibit a severe loss of hair. Initial histological examination of skin biopsies showed pronounced and generalized fibrosis of the dermis, thicker epidermis and inflammatory infiltrates in the area of the skin fibrosis. Preliminary analysis of mouse embryonic fibroblasts derived from these transgenic mice showed elevated expression of collagen type I and Timp-3. We propose to characterize the Col1a2-CTGF mice as well as their explanted skin fibroblasts. To understand the mechanisms by which increased expression of CTGF in fibroblasts causes a fibrotic disease, we will also perform a comparison of the molecular phenotypes of the skin fibroblasts of these mice with those of TBR1CA; Col1a2-CreER mice and with those of specific human scleroderma patients. To further examine these mechanisms we propose to attenuate the fibrotic phenotypes of the skin fibroblasts of the two transgenic mouse models of scleroderma by pharmacological inhibitors of specific signaling pathways or siRNAs. We also will attempt to inhibit the fibrotic phenotypes of these transgenic mice in vivo by crossing null mutations in either SmadS or integrin D6 in these mice. Finally, we will examine the mechanisms that cause increased sensitivity to bleomycin-induced lung fibrosis in TBR1CA; Col1a2-CreER mice and test whether a similar sensitivity occurs in Col1a2-CTGF mice. These studies should give information about the relative importance of TGF-beta and CTGF in causing fibrotic diseases.