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，激活先天免疫回报并创建促氧化剂 通过其中一项死亡计划导致细胞死亡的环境。该项目将重点放在最近描述的 死亡程序，铁吞作用，通过在条件下脂质过氧化的Fe依赖性激活实现 谷胱甘肽过氧化物酶4（GPX4）的缺乏，一种硒酶独特的酶，能够还原磷脂 氢过氧化物。我们确定了15-氢氧基 - 芳基烯酰基磷脂酰乙醇胺（15-Hoo-aa-pe） 铁凋亡的特异性脂质生物标志物。我们还发现15-脂氧合酶（15lox）的复合物与A 脚手架蛋白PEBP1在产生15-HOA-AA-PE信号方面起着主要作用。我们泄露了铁铁作用 人类肺上皮的死亡计划。铁凋亡发生在替代激活的巨噬细胞中 •/iNOS的水平较低，从而导致免疫抑制。出乎意料的是，我们发现这是一个普遍的 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），氧化物宿主多不饱和PE，产生15-Hoo-aa-pe，并且 与内源性宿主15-Lox无关，引起上皮细胞和巨噬细胞的铁凋亡。 在来自ARDS患者的临床铜绿假单胞菌分离株中检测到螺旋病诱导的ploxa。 15-hoo-aa- 在严重免疫受损的ARDS患者的肺样本中鉴定出PE。那，我们 假设存在恶性循环，从而支持了受支持的感染/氧化应激驱动的铁毒性 内源性15-lox充当免疫抑制的主要因素，为 免疫受损肺的继发性铜绿假单胞菌感染，并通过 外源细菌ploxa。我们建议设计和使用选择性的小分子ploxa抑制剂，这些抑制剂 将充当抗肿瘤剂，从而代表新类别的肺部保护剂。 AIM 1将揭示和 解密的致病机制，通过这些机制，通过同工型催化磷脂过氧化的反应 内源性哺乳动物15-lox或外源细菌ploxa - 在GPX4/GSH缺乏症的情况下 导致鼠肺上皮细胞（MLE）和肺泡中的水过氧磷脂积累 巨噬细胞并建立这些产物的分子身份和铁凋亡特性。通过使用氧化还原 脂质组学我们将使用两击模型在体内识别并量化15-hoo-aa-aa-pe生物标志物 免疫抑制。我们还将采用暴露于铜绿假单胞菌的Inos KO动物来揭示 no•作为小鼠肺Vivo中植物驱动的AA-PE氧化和铁毒性死亡的调节剂。在AIM 2中，我们 将设计和开发在上皮和铜绿假单胞菌驱动的浮雕的ploxa的选择性抑制剂中 巨噬细胞是一类新的小分子细胞保护剂，以防止障碍物和 免疫抑制。