Epithelial-Mesenchymal Interactions in Fibrosis Resolution

项目摘要 肺纤维化(PF)仍然是一个主要的和日益增长的医疗负担，治疗方案并不令人满意 这并不能逆转既定的疾病。我们先前的工作已经证实，成纤维细胞中YAP/TAZ的激活是一种 传播PF进展的病理性反馈环的中心特征。在前一个供资周期中， 我们发现多巴胺D1受体(D1R)激动剂是选择性灭活肺中YAP/TAZ的一种策略。 成纤维细胞，部分通过转换导致小鼠实验性肺纤维化的加速缓解 肺成纤维细胞从基质沉积到基质降解状态。此外，我们还证明了人类的肺 PF患者表现出DOPA脱羧酶(DDC)的表达缺陷，DDC是催化 L-多巴转化为生物活性多巴胺。这些研究使我们建议恢复 肺内内源性局部多巴胺水平是基质降解和纤维化的重要触发因素 清除对于成功修复肺是必不可少的，但在肺纤维化时会受到损害。我们的初步数据显示 博莱霉素损伤后幼鼠肺组织中DDC转录本的一过性抑制和升高 在纤维化消退期间，老年小鼠的DDC持续减少，与之平行 纤维化症。此外，小分子抑制DDC酶活性或D1R从第21天到42天。 博莱霉素在幼鼠体内消融肺纤维化的自发消退，显示了其重要作用 用于肝纤维化消退中的多巴胺信号。此外，我们发现在脑脊液上清液中可以检测到多巴胺。 精确切割肺切片，并在培养的纤维化肺切片中减少，证实局部合成 肺内的多巴胺。基于这些发现，我们建议检验上皮细胞 多巴胺合成对纤维化消退和肺纤维化肺组织恢复正常多巴胺水平至关重要 组织可以促进肺组织对胶原的吸收和修复。我们将在三个目标中检验这一假设 跨越未解决的小鼠肺纤维化模型以及小鼠和人的体外模型 肺组织。为了定义多巴胺信号的功能作用，我们将利用细胞特定的条件性 遗传模型以及表征良好的小分子抑制剂和多巴胺激动剂在这些 系统。我们的研究将共同确定控制多巴胺的细胞来源和调控系统 在正常肺修复过程中的生物利用度，并将描绘该修复系统如何在人类肺功能衰竭中失效。这些 研究可能会揭示促进纤维化消退和肺修复的新治疗方法。

Project Summary Pulmonary fibrosis (PF) remains a major and growing medical burden with unsatisfactory therapeutic options that fail to reverse established disease. Our prior work has identified that YAP/TAZ activation in fibroblasts is a central feature of the pathological feedback loop that propagates progression of PF. In the prior funding cycle, we identified Dopamine D1 receptor (D1R) agonism as a strategy to inactivate YAP/TAZ selectively in lung fibroblasts, leading to accelerated resolution of experimental pulmonary fibrosis in mice in part by switching lung fibroblasts from matrix depositing to matrix degrading state. Moreover, we demonstrated that the lungs of individuals with PF exhibit a deficit in expression of DOPA decarboxylase (DDC), the enzyme that catalyzes conversion of L-DOPA into bioactive dopamine. These studies lead us to propose that restoration of endogenous local dopamine levels in the lung is an essential trigger for the matrix degradation and fibrosis clearance that is essential to successful repair of the lung, but is impaired in PF. Our preliminary data show that Ddc transcripts are transiently depressed in lung tissue of young mice following bleomycin injury and rise during fibrosis resolution, whereas aged mice exhibited sustained reductions in Ddc that parallel persistent fibrosis. Moreover, small molecule inhibition of Ddc enzymatic activity or the D1R from day 21 to 42 post- bleomycin in young mice ablates the spontaneous resolution of lung fibrosis, demonstrating the essential role for dopamine signaling in fibrosis resolution. In addition, we find that dopamine is detectable in supernatants of precision cut lung slices and is diminished in slices cultured from fibrotic lungs, confirming the local synthesis of dopamine within the lung. Based on these findings, we propose to test the central hypothesis that epithelial dopamine synthesis is essential to fibrosis resolution and that restoration of normal dopamine levels in PF lung tissue can promote collagen resorption and repair of the lung. We will test this hypothesis in three aims spanning non-resolving mouse models of pulmonary fibrosis as well as ex vivo models of mouse and human lung tissues. To define the functional roles of dopamine signaling we will leverage both cell-specific conditional genetic models as well as well-characterized small molecule inhibitors and dopamine agonists in these systems. Together our studies will define the cellular sources and regulatory systems that control dopamine bioavailability during normal lung repair, and will delineate how this repair system fails in human PF. These studies may reveal new therapeutic approaches to promote fibrosis resolution and lung repair.