The Warburg Effect and Diabetic Retinopathy

瓦尔堡效应和糖尿病视网膜病变

• 批准号: 10635331
• 负责人: Ahmed S Ibrahim
• 金额: $ 38.5万
• 依托单位: WAYNE STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-06-01 至 2028-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10635331
• 关键词: 5' -AMP-activated protein kinase; Acceleration; Aging; Angiogenic Switch; Animals; Benchmarking; Bioenergetics; Bioinformatics; Blindness; Blood Vessels; CRISPR/Cas technology; Cell Proliferation; Cells; Cellular Metabolic Process; Complication; Data; Development; Diabetes Mellitus; Diabetic Retinopathy; Diabetic mouse; Disease; Dose; Endoplasmic Reticulum; Endothelial Cells; Endothelium; Enzymes; Extravasation; Fright; Functional disorder; Gene Expression; Glucose; Glycolysis; Goals; Human; Hyperglycemia; Hypoxia; Hypoxia Inducible Factor; Inositol; Knockout Mice; Knowledge; Lasers; Lipids; Mediating; Mediator; Metabolic; Metabolism; Mission; Mitochondria; Modeling; National Eye Institute; Neuroglia; Neurons; Oxidative Phosphorylation; Oxygen; Pathology; Pathway Analysis; Pathway interactions; Patients; Phenotype; Phosphotransferases; Production; Protein Biosynthesis; Proteins; Research Personnel; Retina; Retinal Neovascularization; Ribonucleases; Risk Factors; Role; Sampling; Signal Transduction; Streptozocin; Testing; Tissues; Tube; Visual Acuity; Warburg Effect; angiogenesis; bevacizumab; cancer cell; conditional knockout; diabetic; endoplasmic reticulum stress; in vitro Model; in vivo; inhibitor; insight; mitochondrial dysfunction; neovascularization; neurovascular; neurovascular unit; novel; nucleotide metabolism; pharmacologic; prevent; proliferative diabetic retinopathy; response; sensor; success

Project Summary/Abstract Retinal neovascularization (RNV) is a debilitating complication of advanced diabetic retinopathy, which despite the use of anti-VEGF and laser treatments continues to cause blindness. Less is known as to why RNV develops only after patients have had diabetes for decades. Although endothelial cell (EC) angiogenic activation is a hallmark of this transition, how ECs adapt their metabolism to sustain such activation remains a significant gap in our knowledge. Our long-term goal is to determine the bioenergetic mechanisms of RNV. The Warburg effect is a metabolic shift from mitochondrial oxidative phosphorylation (OxPhos) to hyperglycolysis that was first found in cancer cells. This metabolic shift not only produces ATP faster than OxPhos, albeit less efficiently, but also provides precursors required for lipid, protein, and nucleotide synthesis during cell proliferation. Recently the Warburg effect was rediscovered as a key contributor in various endothelial-related diseases; however, its role in diabetic retinopathy is not well-defined. In this application, the overall objective(s) are to define the role of the Warburg effect in diabetic retinopathy and to identify its underlying mechanisms. Here, we propose that multiple hits are needed to cooperatively alter EC metabolism to fulfill biosynthetic demands of transforming a quiescent EC into an angiogenic cell. Tissue hypoxia is the most common risk factor associated with advanced diabetic retinopathy. Thus, our central hypothesis is that diabetes primes quiescent ECs to be angiogenic (first hit) and that hypoxia (second hit) is necessary for angiogenic switch via the metabolic adaptation of the Warburg effect. Aim1 will test the hypothesis that persistent activation of the energy sensor, AMP-activated protein kinase (AMPK) sustains the Warburg effect to mediate EC angiogenic activation. Our approach is to use a two-hit model of diabetes and hypoxia in AMPKα1 endothelial-specific conditional knockout (AMPKα1End-/-) mice and in AMPKα1 silenced human retinal ECs (HRECs) to achieve this aim. We will also use vitreous samples from patients with proliferative diabetic retinopathy to test the correlation between the development of RNV and the Warburg effect- associated metabolites. Aim2 will investigate the role of endoplasmic reticulum (ER) stress in mediating the Warburg effect-induced EC angiogenic activation. We hypothesize that activation of Inositol-requiring enzyme (IRE)1, a unique ER-stress sensor protein with kinase and RNase activities, is a key mediator for the Warburg effect-induced EC angiogenic activation. We will use CRISPR/Cas9 to test the effect of inhibiting downstream signaling of IRE1 kinase and RNase activities on the Warburg effect-induced EC angiogenic activation. We will also test our hypothesis in vivo using a two-hit model of hypoxia and diabetes and selective pharmacological inhibitors. Overall, this new-investigator initiated R01 capitalizes on the interdisciplinary expertise of a biochemist, a mitochondrial biologist, an ER biologist, and a clinician to use a novel two-hit model of advanced diabetic retinopathy to gain mechanistic insights into the bioenergetic basis of RNV. Understanding the role of the Warburg effect will reveal novel targets in the treatment of diabetic retinopathy.

项目摘要/摘要 视网膜新生血管(RNV)是晚期糖尿病视网膜病变的一种衰弱并发症，尽管 抗血管内皮生长因子抗体和激光治疗的使用继续导致失明。关于RNV为什么会发展，人们知之甚少。 只有在患者患了几十年的糖尿病之后。尽管内皮细胞(EC)血管生成激活是一种 这一转变的标志是，内皮细胞如何调整其新陈代谢以维持这种激活仍然是一个显著的差距 据我们所知。我们的长期目标是确定RNV的生物能量机制。华宝效应 是首次发现的从线粒体氧化磷酸化(OxPhos)到高糖酵解的代谢转变 在癌细胞中。这种代谢变化不仅产生ATP的速度比OxPhos快，尽管效率较低，而且 提供细胞增殖过程中脂肪、蛋白质和核苷酸合成所需的前体。最近一段时间 Warburg效应被重新发现为各种内皮相关疾病的关键因素；然而，它的作用 在糖尿病视网膜病变中的定义并不明确。在本申请中，总体目标(S)是定义 糖尿病视网膜病变中的Warburg效应及其潜在机制。在这里，我们建议 需要多个Hit来协同改变EC的新陈代谢，以满足转化为 静止的EC转化为血管生成细胞。组织缺氧是与晚期癌症相关的最常见的危险因素 糖尿病视网膜病变。因此，我们的中心假设是糖尿病使静止的内皮细胞具有血管生成作用(第一 Hit)和缺氧(第二次Hit)是通过Warburg代谢适应进行血管生成开关所必需 效果。Aim1将检验能量感受器AMP激活蛋白激酶持续激活的假设 (AMPK)维持Warburg效应以介导EC血管生成激活。我们的方法是使用两次成功的模型 AMPKα1内皮特异性条件性基因敲除(AMPKα1End-/-)小鼠和AMPKα1中糖尿病和缺氧的研究 为了达到这一目的，沉默的人视网膜内皮细胞(HRECs)。我们还将使用来自慢性玻璃体疾病患者的玻璃体样本 以测试RNV的发展与Warburg效应之间的相关性- 相关的代谢物。AIM2将研究内质网(ER)应激在介导 Warburg效应诱导内皮细胞血管生成激活。我们假设肌醇需求酶的激活 (IRE)1是一种独特的内质网应激感受器蛋白，具有激酶和核糖核酸酶活性，是Warburg信号转导的关键调节因子 作用诱导内皮细胞血管生成激活。我们将使用CRISPR/Cas9来测试抑制下游的效果 IRE1激酶和核糖核酸酶活性在Warburg效应诱导的EC血管生成激活中的信号转导。我们会 我还在体内使用缺氧和糖尿病的两次打击模型和选择性药物来测试我们的假设 抑制剂。总体而言，这一由新研究员发起的R01利用了 生物化学家、线粒体生物学家、内质网生物学家和临床医生使用先进的 糖尿病视网膜病变，以获得对RNV的生物能量学基础的机械见解。了解企业的角色 Warburg效应将为糖尿病视网膜病变的治疗提供新的靶点。