Targeting the Apoptosis-Resistant Pulmonary Myofibroblast

描述(申请人提供)：特发性肺纤维化(IPF)是一种无法治愈的致命疾病，发病率和死亡率都在增加。尽管最近协调努力将已在体外和临床前模型中得到验证的发现迅速转化为生物学上有希望的干预措施，但在治疗方面尚未取得重大突破。以纤维化为特征的破坏性重塑的最后一个共同途径是抗凋亡的肌成纤维细胞，它既创造并响应改变的介质和基质微环境，从而使纤维形成永久化。阻止肌成纤维细胞表型的发展，或一旦建立就逆转它，为成功治疗IPF提供了最大的希望。细胞表面糖蛋白Thy-1是一种已知的肿瘤抑制因子，它作为细胞表型的上下文依赖调节因子，在成纤维细胞灶内的肌成纤维细胞中沉默，这是IPF的标志性组织病理学损害，其存在预示着预后不良。在培养的肺成纤维细胞中，Thy-1缺失可促进细胞增殖、细胞因子和生长因子的表达以及反应性、迁移、肌成纤维细胞分化和抗凋亡，所有这些都在Thy-1重新表达后可逆。THY-1与αv整合素和Syndecan 4在细胞表面相互作用，调节细胞-细胞和细胞-基质相互作用，并 机械耦合，抑制转化生长因子-β的激活和肌成纤维细胞的分化。Thy-1对肌成纤维细胞表型的影响是广泛的，包括：减少一些肌肉特异性蛋白和肌源性转录因子的表达，抑制收缩， 和促进细胞凋亡。此外，Thy-1的一些功能可以通过注射可溶性Thy-1来概括，初步数据表明，在小鼠模型中，可溶性Thy-1可以逆转已建立的纤维化，这表明Thy-1本身可以用于治疗或修改。综上所述，这些知识引导我们得出一个总体假设，即Thy-1介导的信号广泛抑制肌成纤维细胞的转化，促进细胞凋亡，恢复肺成纤维细胞的体内平衡功能。对相关分子机制的更好理解将发现逆转IPF和其他纤维化疾病的病理性肌成纤维细胞表型的新策略。基于Thy-1异质性和靶向性干扰的体外和体内模型为确定肌成纤维细胞靶向策略提供了极好的机会，具体目的如下：目的1：确定Thy-1抑制肺成纤维细胞肌成纤维细胞分化的机制；目的2：确定Thy-1促进肌成纤维细胞凋亡的机制；以及目标3：利用Thy-1调节的信号逆转体内抗凋亡的肌成纤维细胞表型。意义：由于Thy-1调节多个对纤维化形成至关重要的信号通路，因此利用其功能比靶向 单一途径，这在IPF中已被证明是无效的方法。因此，拟议的研究将确定Thy-1抑制肺成纤维细胞纤维化前分化的机制，并将这一知识转化为纤维肺疾病的创新治疗干预措施。

DESCRIPTION (provided by applicant): Idiopathic pulmonary fibrosis (IPF) is an incurable fatal disease with increasing incidence and mortality. Despite recent coordinated attempts to rapidly translate findings which have been validated in vitro and in preclinical models into biologically promising interventions, there have been no major therapeutic breakthroughs. A final common pathway for the destructive remodeling which characterizes fibrosis is the apoptosis-resistant myofibroblast, which both creates and responds to an altered mediator and matrix microenvironment, thus perpetuating fibrogenesis. Halting the development of the myofibroblast phenotype, or reversing it once established, offers the best hope of successfully treating IPF. The cell surface glycoprotein Thy-1, a known tumor suppressor which acts as a context-dependent regulator of cell phenotype, is silenced in the myofibroblasts within fibroblastic foci, the signature histopathologic lesions in IPF, the presence of which portends a poor prognosis. Absence of Thy-1 in cultured lung fibroblasts promotes proliferation, cytokine and growth factor expression and responsiveness, migration, myofibroblastic differentiation, and resistance to apoptosis, all reversible upon re-expression of Thy-1. Thy-1 interacts with alpha v integrins and syndecan 4 at the cell surface, modulating cell-cell and cell-matrix interactions and mechanical coupling, to inhibit TGF-beta activation and myofibroblastic differentiation. The effects of Thy-1 on the myofibroblast phenotype are broad, including: decreased expression of a number of muscle-specific proteins and myogenic transcription factors, inhibition of contractility, and promotion of apoptosis. Furthermore, some of the functions of Thy-1 can be recapitulated by the administration of soluble Thy-1, and preliminary data demonstrate that soluble Thy-1 reverses established fibrosis in a mouse model, indicating that Thy-1 itself can be used or modified for therapeutic benefit. Taken together, this knowledge leads us to the overall hypothesis that Thy-1-mediated signaling broadly suppresses myofibroblastic transformation and promotes apoptosis, restoring homeostatic function in lung fibroblasts. Improved understanding of the molecular mechanisms involved will uncover novel strategies for reversing the pathogenic myofibroblast phenotype in IPF and other fibrotic disorders. In vitro and in vivo models based on heterogeneity and targeted disruption of Thy-1 offer excellent opportunities for defining myofibroblast-targeting strategies, as outlined in the following specific aims: Aim 1: To define the mechanisms by which Thy-1- inhibits myofibroblastic differentiation of lung fibroblasts; Aim 2: To define the mechanisms by which Thy-1 promotes myofibroblast apoptosis; and Aim 3: To harness Thy-1-modulated signaling to reverse the apoptosis-resistant myofibroblast phenotype in vivo. Significance: Because Thy-1 modulates multiple signaling pathways critical to fibrogenesis, capitalizing on its function is more attractive than targeting a single pathway, which has been shown to be an ineffective approach in IPF. The proposed studies will thus define mechanisms by which Thy-1 suppresses profibrotic differentiation of lung fibroblasts, and translate that knowledge into innovative therapeutic interventions for fibrotic lung disease.