Ferroptosis as a Death Mechanism in Lung Injury - Project 2

铁死亡作为肺损伤的死亡机制 - 项目 2

• 批准号: 10631057
• 负责人: Valerian E Kagan
• 金额: $ 36.75万
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-01-03 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10631057
• 关键词: Acute; Acute Lung Injury; Acute Respiratory Distress Syndrome; Alveolar Macrophages; Animals; Apoptosis; Apoptotic; Arachidonate 15-Lipoxygenase; Bacterial Pneumonia; Biological Markers; Brain Diseases; Cardiolipins; Cell Death; Cell Death Signaling Process; Cells; Cessation of life; Chronic; Clinical; Complex; Cytoprotective Agent; Development; Environment; Epithelial Cells; Epithelium; Event; Evolution; Exposure to; Human; Hydrogen Peroxide; Immune; Immune response; Immunocompromised Host; Immunosuppression; Impairment; Individual; Inflammation; Innate Immune Response; Kidney Diseases; Lead; Lipid Peroxidation; Lipids; Lipoxygenase Inhibitors; Liver diseases; Lung; Macrophage; Membrane; Mitochondria; Modeling; Molecular; Mus; Oxidation-Reduction; Oxidative Stress; Pathogenesis; Pathogenicity; Pathway interactions; Patients; Phosphatidylethanolamine; Phospholipids; Play; Property; Protein Isoforms; Pseudomonas aeruginosa; Pseudomonas aeruginosa infection; Reaction; Reactive Oxygen Species; Reporting; Respiratory Tract Infections; Role; Sampling; Scaffolding Protein; Signal Transduction; Signaling Molecule; Testing; Virulence Factors; Virulent; Work; cell injury; design; epithelial injury; glutathione peroxidase; in vivo; inhibitor; lipidomics; lung injury; mortality; novel; novel therapeutic intervention; novel therapeutics; oxidation; pathogen; pathogenic bacteria; peroxidation; phospholipid inhibitor; predictive marker; prevent; programs; selenoenzyme; small molecule

In ARDS, bacterial pathogens damage host cells, activate innate immune responses, and create a pro-oxidant environment leading to cell death via one of the death programs. This Project will focus on a recently described death program, ferroptosis, realized via Fe-dependent activation of lipid peroxidation under conditions of deficiency of glutathione peroxidase 4 (GPX4), a seleno-enzyme uniquely capable of reducing phospholipid hydroperoxides. We identified 15-hydroperoxy-arachidonoyl-phosphatidylethanolamines (15-HOO-AA-PE) as specific lipid biomarkers of ferroptosis. We also discovered that complexes of 15-lipoxygenase (15LOX) with a scaffold protein, PEBP1, play the major role in generating 15-HOO-AA-PE signals. We divulged ferroptosis as a death program of the human pulmonary epithelium. Ferroptosis occurs in alternatively activated macrophages with low levels of NO•/iNOS, thus causing immuno-suppression. Unexpectedly, we discovered that a common Gram-negative pathogen, P. aeruginosa – that does not contain polyunsaturated phospholipid oxidation substrates– expresses 15LOX (pLoxA) which oxidizes host polyunsaturated PE, generates 15-HOO-AA-PE and causes ferroptosis in epithelial cells and macrophages independently of the endogenous host 15-LOX. Ferroptosis-inducing pLoxA was detected in clinical P. aeruginosa isolates from ARDS patients. 15-HOO-AA- PE were identified in the lung samples from severely immuno-compromised patients with ARDS. Thus, we postulate the existence of a vicious cycle whereby inflammation/oxidative stress driven ferroptosis supported by endogenous 15-LOX acts as the major contributor to immunosuppression that sets the stage for the secondary P. aeruginosa infection of immune-impaired lung and further enhancement of ferroptosis by exogenous bacterial pLoxA. We propose to design and use selective small molecule pLoxA inhibitors, which will act as anti-ferroptotic agents thus representing new classes of pulmonary protectors. Aim 1 will reveal and decipher pathogenic mechanisms through which reactions of phospholipid peroxidation catalyzed by isoforms of endogenous mammalian 15-LOX or exogenous bacterial pLoxA – in conditions of GPX4/GSH deficiency – lead to accumulation of hydroperoxy-phospholipids in murine lung epithelial cells (MLE) and alveolar macrophages and establish molecular identity and ferroptotic properties of these products. By using redox lipidomics we will identify and quantify 15-HOO-AA-PE biomarkers of ferroptosis in vivo using a two-hit model of immunosuppression. We will also employ iNOS KO animals exposed to P. aeruginosa to reveal the role of NO• as a regulator of pLoxA-driven AA-PE oxidation and ferroptotic death in the mouse lung vivo. In Aim 2, we will design and develop selective inhibitors of pLoxA regulating P. aeruginosa-driven ferroptosis in epithelia and macrophages as a new class of small-molecule cytoprotective agents preventing breach of the barrier and immunosuppression.

在 ARDS 中，细菌病原体会损害宿主细胞，激活先天免疫反应，并产生促氧化剂 通过其中一个死亡程序导致细胞死亡的环境该项目将重点关注最近描述的一种。 死亡程序，铁死亡，通过在以下条件下依赖铁的脂质过氧化激活来实现 谷胱甘肽过氧化物酶 4 (GPX4) 缺乏，这是一种独特的能够还原磷脂的硒酶 我们将 15-氢过氧-花生四烯酰-磷脂酰乙醇胺 (15-HOO-AA-PE) 鉴定为 我们还发现 15-脂氧合酶 (15LOX) 与铁死亡的特异性脂质生物标志物。 蛋白质支架 PEBP1 在产生 15-HOO-AA-PE 信号中起主要作用，我们将铁死亡称为铁死亡。 人类肺上皮的死亡程序发生在交替激活的巨噬细胞中。 出乎意料的是，我们发现了一个常见的现象。 革兰氏阴性病原体，铜绿假单胞菌 – 不含多不饱和磷脂氧化 底物 - 表达 15LOX (pLoxA)，氧化宿主多不饱和 PE，生成 15-HOO-AA-PE 和 独立于内源性宿主 15-LOX 导致上皮细胞和巨噬细胞铁死亡。 在 ARDS 患者的临床铜绿假单胞菌中检测到诱导铁死亡的 pLoxA。 在严重免疫功能低下的 ARDS 患者的肺部样本中发现了 PE。 假设存在恶性循环，从而支持炎症/氧化应激驱动的铁死亡 内源性 15-LOX 是免疫抑制的主要贡献者，为免疫抑制奠定了基础 免疫受损肺的继发性铜绿假单胞菌感染和铁死亡的进一步增强 我们建议设计和使用选择性小分子 pLoxA 抑制剂。 将作为抗铁焦亡剂，从而代表新类别的肺保护剂，目标 1 将揭示和 破译异构体催化磷脂过氧化反应的致病机制 内源性哺乳动物 15-LOX 或外源性细菌 pLoxA – 在 GPX4/GSH 缺乏的条件下 – 导致鼠肺上皮细胞（MLE）和肺泡中氢过氧磷脂的积累 巨噬细胞并通过使用氧化还原建立这些产品的分子特性和铁死亡特性。 在脂质组学中，我们将使用两次命中模型来识别和量化体内铁死亡的 15-HOO-AA-PE 生物标志物 我们还将使用暴露于铜绿假单胞菌的 iNOS KO 动物来揭示免疫抑制的作用。 NO• 作为 pLoxA 驱动的 AA-PE 氧化和小鼠肺体内铁死亡的调节剂 在目标 2 中，我们。 将设计和开发 pLoxA 的选择性抑制剂，调节上皮细胞中铜绿假单胞菌驱动的铁死亡 巨噬细胞作为一类新型小分子细胞保护剂，可防止突破屏障 免疫抑制。