Metabolic control of DNA repair in pulmonary fibrosis

摘要：高龄是发生肺纤维化的重要危险因素，但潜在的 导致这种联系的机制还不清楚。在本应用程序中，我们描述了一个基本的 衰老通过损害代谢反应促进肺纤维化发展的机制 在肺泡上皮细胞中。在年轻的小鼠身上，我们发现博莱霉素激活了一种代谢程序 哪些肺泡上皮II型细胞(AEC2)迅速减少细胞质中乙酰-辅酶A的利用 同时将乙酰辅酶A的生产机制转移到细胞核，以增强核心组蛋白 乙酰化和增强DNA修复。我们发现，这些代谢变化的中心是激活 AMPK对于减少乙酰辅酶A的细胞质利用和动员乙酰辅酶A都是至关重要的 向细胞核产生三磷酸腺苷-柠檬酸裂解酶(ACL)。重要的是，我们证明了这种自适应的相互作用 老年小鼠肺中胞质和胞核之间的联系受损，这在很大程度上是由于AMPK减少 活动。此外，我们还证明，通过增强AMPK的激活或增加代谢的可用性 中间体我们可以增强核心组蛋白乙酰化，促进DNA修复和减弱纤维化反应 在老年小鼠未受损伤的全肺组织中和在博莱霉素作用下的培养的AEC中。加在一起， 这些发现导致我们提出了以下中心假设：我们假设与年龄相关的下降 AMPK的激活有助于增强肺对博莱霉素的敏感性 在恢复AMPK激活或提高核内代谢中间产物的可用性方面将 增强核心组蛋白乙酰化，促进DNA修复，减轻肺纤维化反应。为了测试 这些假设我们提出如下：在具体目标1，我们将确定AMPK的重要性 激活在调节细胞代谢和控制核心组蛋白乙酰化/DNA修复中的作用 我们将确定激活这一通路是否会减少肺组织的纤维化反应 在特定的目标2中，我们将确定前交叉韧带动员对细胞核的关键作用 在博莱霉素进行核心组蛋白乙酰化和DNA修复后，我们将测定ATP-柠檬酸裂解酶 与年龄匹配的对照组相比，IPF肺组织中的水平降低；最后，在特定目标3中，我们将 通过确定是否供应不同的药物来确定旁路AMPK活性不足的治疗有效性 代谢底物恢复核心组蛋白乙酰化，增强DNA修复并减弱纤维化反应 在幼年和老年小鼠的肺中。总而言之，这项提案将建立老龄化的机制 增加博莱霉素中毒后肺纤维化的易感性。此外，我们预计这些调查结果 研究将为未来的研究奠定基础，测试新的药理学方法 本申请中详述的靶向代谢途径可以减轻纤维化的发病和/或严重程度 在肺和其他肺外组织中的反应。

ABSTRACT: Advanced age is an important risk factor for developing pulmonary fibrosis but the underlying mechanisms leading to this association are not understood. In this application, we describe a fundamental mechanism by which aging promotes the development of pulmonary fibrosis by impairing metabolic responses in the alveolar epithelium of the lung. In young mice, we show that bleomycin activates a metabolic program in which alveolar epithelial type II cells (AEC2) rapidly reduce utilization of acetyl-CoA in the cytoplasm and simultaneously divert the machinery for acetyl-CoA production to the nucleus in order to enhance core histone acetylation and augment DNA repair. We found that central to these metabolic changes is the activation of AMPK, which is critical for both reducing cytoplasmic utilization of acetyl CoA and mobilizing the acetyl-CoA producing enzyme ATP-citrate lyase (ACL) to the nucleus. Importantly, we show that this adaptive interplay between the cytoplasm and nucleus is impaired in the lungs of older mice due, in large part, to reduced AMPK activity. Further, we demonstrate that by enhancing AMPK activation or increasing the availability of metabolic intermediates we can augment core histone acetylation, increase DNA repair and attenuate fibrotic responses in uninjured whole lung tissues of older mice and in cultured AECs exposed to bleomycin. Taken together, these findings led us to propose the following central hypothesis: We hypothesize that age-related decreases in AMPK activation contribute to the enhanced susceptibility of the lung to bleomycin and that strategies aimed at restoring AMPK activation or enhancing the availability of metabolic intermediates in the nucleus will enhance core histone acetylation, increase DNA repair and attenuate fibrotic responses in the lung. To test these hypotheses we propose the following: In Specific Aim 1, we will establish the importance of AMPK activation in the regulating cellular metabolism and controlling core histone acetylation/DNA repair in the alveolar epithelium and we will determine whether activating this pathway reduces fibrotic responses in lungs of young and old mice; In Specific Aim 2, we will establish the critical role of ACL mobilization to the nucleus after bleomycin for core histone acetylation and DNA repair and we will determine whether ATP-citrate lyase levels are reduced in IPF lung tissue compared to age-matched controls; and lastly, in Specific Aim 3, we will establish the therapeutic utility of bypassing deficient AMPK activity by determining whether supplying different metabolic substrates restores core histone acetylation, augments DNA repair and attenuates fibrotic responses in lungs of young and old mice. In summary, this proposal will establish the mechanisms by which aging enhances susceptibility to lung fibrosis after bleomycin insult. Further, we anticipate that findings from these studies will lay the foundation for future investigations testing whether novel pharmacological approaches targeting metabolic pathways detailed in this application can attenuate the onset and/or severity of fibrotic responses in lung, and in other extrapulmonary tissues.