AMPK/SIRT1/PGC-1α, a critical pathway in dry AMD

摘要 老年性黄斑变性(AMD)是美国55岁以上人群失明的主要原因 发达国家。AMD的两种形式之一是“干性”形式，目前还没有有效的方法 治疗。因此，AMD的新疗法的开发有一个未得到满足的医学需求。一个 包括AMD在内的许多视网膜疾病都与线粒体功能障碍有关。功能失调 线粒体导致活性氧簇(ROS)水平升高和代谢活性缺陷。 自噬缺失也会导致线粒体功能障碍，并被认为增加了氧化的易感性 压力和AMD。我们最近发现了功能障碍的自噬、ROS增加和功能障碍 AMD供眼RPE中的线粒体。然而，导致这些的潜在机制 导致AMD的代谢同素酶缺陷仍不清楚。过氧化物酶体增殖物激活 受体-γ共激活物-1α(pGC-1)在线粒体生物发生和发展中起重要作用 氧化代谢。它还调节自噬和有丝分裂吞噬。PGC-1活性受两个主要因素的刺激 因子：AMP活化蛋白激酶(AMPK)和NAD依赖的脱乙酰酶，SIRT1。初步证据 我们实验室的研究表明AMPK/SIRT1/PGC-1在AMD RPE中表达下调。基于我们的 初步数据，我们假设RPE中抑制的AMPK/SIRT1/PGC-1通路诱导 线粒体、自噬功能障碍和ROS产生增加，从而导致代谢异常 活动，脂肪和糖原的积累，以及玻璃体的形成，导致了AMD的病理生理。为了测试 我们的假设是，我们已经建立了一个取之不尽的AMD体外疾病模型，方法是从 AMD供者的眼睛，随后是IPSC的产生和随后分化为RPE(AMD RPE- IPSC-RPE)。我们还从年龄匹配的正常供者的RPE(Normal RPE-IPSC-RPE)中产生了IPSC。 作为控制点。我们证实AMD RPE-IPSC-RPE模拟了他们的疾病表型 父母供体，主要的AMD RPE，这验证了我们的模型。此外，我们还建立了一个动物模型 检测pGC-1抑制对视网膜色素上皮和视网膜健康的影响，并观察视网膜色素上皮和光感受器 退化。我们提出了两个目标：AIM1将测试AMPK/SIRT-1/PGC-1通路在干燥症中的抑制作用 使用我们建立的AMD供者和AMD患者的体外模型。AIM2将调查蜂窝 以及pGC-1在小鼠模型中对视网膜色素上皮和视网膜健康的作用的分子机制。最终，这些 研究将为干性AMD的分子机制提供洞察力，并可能促进新的治疗方法 干预措施。

ABSTRACT Age-related macular degeneration (AMD) is the major cause of blindness in people over age 55 in the U.S. and the developed world. One of the two forms of AMD is the “dry” form, for which currently there are no effective treatments. Consequently, there is an unmet medical need for the development of new therapies for AMD. A number of retinal diseases, including AMD, are associated with mitochondrial dysfunction. Dysfunctional mitochondria induce increased levels of reactive oxygen species (ROS) and defective metabolic activity. Autophagy loss also results in mitochondrial dysfunction and is suggested to increase susceptibility to oxidative stress and AMD. We have recently shown dysfunctional autophagy, increased ROS, and dysfunctional mitochondria in RPE derived from AMD donor eyes. However, the underlying mechanisms inducing these defective metabolic homeostases leading to AMD remain unknown. The Peroxisome proliferator-activated receptor-gamma coactivator (PGC)-1alpha (PGC-1) plays a major role in mitochondrial biogenesis and oxidative metabolism. It also regulates autophagy and mitophagy. PGC-1 activity is stimulated by two main factors: AMP-activated protein kinase (AMPK) and NAD+ - dependent deacetylase, SIRT1. Preliminary evidence from our laboratory suggests that the AMPK/SIRT1/PGC-1 is downregulated in AMD RPE. Based on our preliminary data, we hypothesize that the repressed AMPK/SIRT1/PGC-1 pathway in RPE induces mitochondrial, autophagic dysfunction, and increased ROS production, which results in abnormal metabolic activity, lipid and glycogen accumulation, and drusen formation, leading to the AMD pathophysiology. To test our hypothesis, we have developed an inexhaustible AMD in vitro disease model by isolating native RPE from AMD donors’ eyes, followed by the generation of iPSC and their subsequent differentiation into RPE (AMD RPE- iPSC-RPE). We have also generated iPSC from RPE of age-matched normal donors (Normal RPE-iPSC-RPE) that serve as the control. We confirmed that the AMD RPE-iPSC-RPE mimic the disease phenotypes of their parental donors, the primary AMD RPE, which validates our model. Additionally, we established an animal model to test the role of PGC-1 repression on RPE and retinal health and observed RPE and photoreceptor degeneration. We propose two aims: Aim1 will test the role of AMPK/SIRT-1/PGC-1 pathway inhibition in dry AMD using our established in vitro model from AMD donors and AMD patients. Aim2 will investigate the cellular and molecular mechanisms of PGC-1 actions on RPE and retinal health in a mouse model. Ultimately, these studies will provide insight into the molecular mechanisms of dry AMD and may facilitate new therapeutic interventions.