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

TGF-β 和 CTGF 信号传导转基因小鼠硬皮病模型的作用

• 批准号: 7486189
• 负责人: BENOIT DE CROMBRUGGHE
• 金额: $ 38.8万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-09-01 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7486189
• 关键词: Affect; Animals; Area; Attenuated; Biochemical; Biopsy; Bleomycin; Blood Vessels; Collagen; Collagen Gene; Collagen Type I; Complex; Connective Tissue Diseases; Dermal; Dermis; Disease; Embryo; Enhancers; Epidermis; Exhibits; Extracellular Matrix; Fibroblasts; Fibrosis; Hair; Hair follicle structure; Human; Inflammatory Infiltrate; Integrins; Kidney; Lung; Molecular; Mus; Organ; Patients; Phenotype; Play; Process; Relative (related person); Role; Scleroderma; Signal Pathway; Signal Transduction; Skin; Systemic Scleroderma; Testing; Thick; Transforming Growth Factor beta; Transgenic Mice; Transgenic Organisms; body system; connective tissue growth factor; in vivo; inhibitor/antagonist; mouse hypertrophic chondrocyte-specific protein 24; mouse model; novel; null mutation; promoter

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.

硬皮病或系统性硬化症 (SSc) 是一种影响多种器官的结缔组织疾病 系统。这种疾病很复杂，其特点是胶原蛋白和其他物质过度积累。 皮肤和内脏器官中的细胞外基质成分。因为 TGF-β 信号传导增强 由于与这种疾病有关，我们最近建立了一种新型小鼠模型，其中 TGF-β 受体1 出生后在成纤维细胞中组成型激活（TBR1CA；Col1a2-CreER）。这些小鼠重现了 人类SSc的主要特征，显示真皮明显且广泛的纤维化，表皮变薄， 毛囊脱落，肺和肾小血管壁纤维化增厚。原发性皮肤 这些小鼠的成纤维细胞显示下游 TGF-β 靶标的表达升高，再现了 外植的 SSc 真皮成纤维细胞的标志性生化表型。特别是有一个明显的 结缔组织生长因子（CTGF）表达增加。由于 CTGF 表达增加 被认为在疾病过程中发挥关键作用，我们培育了过度表达的转基因小鼠 使用来自 pro-D2(l) 的成纤维细胞特异性启动子/增强子在成纤维细胞 (Col1a2-CTGF) 中产生 CTGF 胶原蛋白基因。这些动物表现出严重的毛发脱落。皮肤活检的初步组织学检查 显示出明显且广泛的真皮纤维化、表皮增厚和炎症浸润 皮肤纤维化的区域。对源自这些的小鼠胚胎成纤维细胞的初步分析 转基因小鼠表现出 I 型胶原蛋白和 Timp-3 表达升高。 我们建议对 Col1a2-CTGF 小鼠及其外植的皮肤成纤维细胞进行表征。到 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的皮肤成纤维细胞的分子表型与 TBR1CA 的那些； Col1a2-CreER 小鼠和特定人类硬皮病患者的小鼠。为了进一步 检查这些机制，我们建议减轻两种皮肤成纤维细胞的纤维化表型 通过特定信号通路的药物抑制剂建立硬皮病转基因小鼠模型或 siRNA。我们还将尝试通过杂交来抑制这些转基因小鼠体内的纤维化表型 这些小鼠中 SmadS 或整合素 D6 均无突变。最后，我们将研究以下机制： 导致 TBR1CA 对博来霉素诱导的肺纤维化的敏感性增加； Col1a2-CreER 小鼠和测试 Col1a2-CTGF 小鼠中是否出现类似的敏感性。这些研究应该提供有关 TGF-β 和 CTGF 在引起纤维化疾病中的相对重要性。