Sirtuins in Lung Aging and Fibrosis

人类纤维化病症影响许多器官系统，包括心脏、血管、肾、肝和肺， 在美国，由于过度的疾病负担，包括我们的退伍军人管理局（VA）患者， 人口最常见的纤维化肺病，特发性肺纤维化（IPF），其特征在于： 过度瘢痕组织形成和肺实质的不可逆破坏，导致气体交换 最终导致呼吸衰竭。老龄化是IPF最强的风险因素;然而， 与年龄相关的纤维化疾病易感性的细胞/分子机制仅 开始被探索。提高对衰老在这种疾病中作用的认识将有助于 开发具有更高临床疗效的新疗法。 为了支持VA优异奖的更新申请，我们最近为我们的 了解肺损伤和修复机制中的衰老机制。IPF人类受试者表达 在肌纤维母细胞灶和从IPF分离的离体成纤维细胞中，线粒体sirtuin，SIRT 3水平低 肺SIRT 3的这种下调在小鼠非消退性纤维化的衰老模型中复制; 年轻小鼠中的纤维化在博来霉素后2-4个月基本上消退，老年小鼠表现出持续的纤维化。 我们的数据表明，年轻小鼠的这种纤维化消退能力与SIRT 3的恢复有关。 在肺损伤的后期修复阶段，这一点在老年小鼠中不存在。然而， SIRT 3（通过肺靶向非病毒cDNA质粒递送）恢复老年人纤维化消退的能力 小鼠这种保护作用与FoxO 3a的体内激活有关，这一点可以通过较高水平的FoxO 3a来证明。 核FoxO 3a。细胞衰老和促纤维化细胞因子转化生长因子-β1（TGF-β 1） β1）诱导SIRT 3和FoxO 3a下调，导致线粒体功能障碍， 肺成纤维细胞的衰老和凋亡抵抗。我们的研究也支持这样一种可能性， 肺泡巨噬细胞中SIRT 3的调节通过旁分泌调节FoxO 3a的成纤维细胞活化 机制此外，组蛋白乙酰转移酶，p300和环AMP反应元件结合， CREB结合蛋白（CBP）参与TGF-β1对SIRT 3的表观遗传下调 和细胞衰老。SIRT 3-FoxO 3a信号传导轴的活化受损促进衰老， 抗纤维化的成纤维细胞表型，导致持续性/非消退性纤维化。 在这项拨款申请中要检验的中心假设是，持续性/非消退性纤维化 与衰老相关的是由成纤维细胞中Foxo 3a的活化受损介导的， 成纤维细胞本身或巨噬细胞衍生的旁分泌机制，有助于线粒体 肺成纤维细胞的功能障碍、衰老和凋亡抵抗。 我们的具体目标是：（1）阐明肺组织中SIRT 3的p300/CBP依赖的表观遗传调控 成纤维细胞和肺泡巨噬细胞介导的前分辨率表型;（2）研究的作用， SIRT 3-FoxO 3a信号轴在恢复线粒体生物能量学和细胞凋亡易感性中的作用 衰老/纤维化的肺成纤维细胞;和（3）确定是否由SIRT 3重建， 遗传/药理学方法通过激活FoxO 3a促进老年小鼠的纤维化消退， 肺成纤维细胞完成本提案中的目标将：（a）揭示表观遗传机制， 控制SIRT 3表达与细胞衰老/老化;（B）提供对SIRT 3的作用的机制见解， SIRT 3-FoxO 3a信号传导在线粒体生物能量学和细胞可塑性/命运调节中的作用;和（c） 阐明促消退巨噬细胞的巨噬细胞-成纤维细胞串扰和SIRT 3调节;和（d）提供 SIRT 3-FoxO 3a信号通路的激活促进纤维化消退， 治疗进展性肺病的新方法。

Human fibrotic disorders affect many organ systems including heart, blood vessels, kidney, liver and lung, accounting for excessive disease burden in the U.S., including among our Veterans Administration (VA) patient population. The most common fibrotic lung disease, idiopathic pulmonary fibrosis (IPF), is characterized by excessive scar tissue formation and irreversible destruction of the lung parenchyma, resulting in gas-exchange abnormalities and ultimately respiratory failure. Aging is strongest risk factor for IPF; however, the cellular/molecular mechanisms that account for the age-associated predilection to fibrotic disease are only beginning to be explored. Improved understanding of the role of aging in this disease will aid in the development of novel therapies with greater clinical efficacy. In support of this VA Merit Award renewal application, we have made several recent contributions to our understanding of aging mechanisms in lung injury and repair mechanisms. Human subjects with IPF express low levels of the mitochondrial sirtuin, SIRT3, in myofibroblastic foci and in ex-vivo fibroblasts isolated from IPF lungs. This down-regulation of SIRT3 is replicated in an aging model of non-resolving fibrosis in mice; while fibrosis in young mice largely resolves over 2-4 months post-bleomycin, aged mice manifest persistent fibrosis. Our data indicate that this capacity for fibrosis resolution in young mice is associated with recovery of SIRT3 levels in the late reparative phase of lung injury, while this is absent in aged mice. However, re-constitution of SIRT3 (via lung-targeted non-viral cDNA plasmid delivery) restores the capacity for fibrosis resolution in aged mice. This protective effect is associated with in-vivo activation of FoxO3a, evidenced by higher levels of nuclear FoxO3a. Both cellular senescence and the pro-fibrotic cytokine, transforming growth factor-β1 (TGF- β1) induce a down-regulation of SIRT3 and FoxO3a, events that lead to mitochondrial dysfunction, senescence and apoptosis resistance of lung fibroblasts. Our studies also support the possibility that modulation of SIRT3 in alveolar macrophages regulates fibroblast activation of FoxO3a by a paracrine mechanism. Additionally, the histone acetyltransferases, p300 and cyclic AMP-response element binding protein (CREB)-binding protein (CBP), are implicated in the epigenetic down-regulation of SIRT3 by TGF-β1 and cellular senescence. Impaired activation of the SIRT3-FoxO3a signaling axis promotes a senescent and apoptosis-resistant fibroblast phenotype, which drives persistent/non-resolving fibrosis. The central hypothesis to be tested in this grant proposal is that persistent/non-resolving fibrosis associated with aging is mediated by impaired activation of Foxo3a in fibroblasts, either by SIRT3 deficiency in fibroblasts themselves or by macrophage-derived paracrine mechanisms, contributes to mitochondrial dysfunction, senescence and apoptosis resistance of lung fibroblasts. Our specific aims are to: (1) elucidate p300/CBP-dependent epigenetic regulation of SIRT3 in lung fibroblasts and alveolar macrophages that mediate pro-resolution phenotypes; (2) investigate the role of the SIRT3-FoxO3a signaling axis in restoring mitochondrial bioenergetics and apoptosis susceptibility to senescent/fibrotic lung fibroblasts; and (3) determine whether reconstitution of SIRT3 by genetic/pharmacologic approaches promotes fibrosis resolution in aged mice via the activation of FoxO3a in lung fibroblasts. The completion of the Aims in this proposal will: (a) uncover epigenetic mechanisms that control SIRT3 expression with cellular senescence/aging; (b) provide mechanistic insights into the role of SIRT3-FoxO3a signaling in the regulation of mitochondrial bioenergetics and cellular plasticity/fate; and (c) elucidate macrophage-fibroblast crosstalk and SIRT3 regulation of pro-resolving macrophages; and (d) provide proof-of-concept that activation of the SIRT3-FoxO3a signaling pathway promotes fibrosis resolution, leading to new therapeutic approaches for progressive lung diseases.