Evaluation of Mitochondrial Cardiolipin Modification in Neonatal Hypoxia/Ischemia Encephalopathy

线粒体心磷脂修饰在新生儿缺氧/缺血性脑病中的评价

• 批准号: 10679306
• 负责人: Katlynn Joy Emaus
• 金额: $ 4.08万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 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 2026-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10679306
• 关键词: Anabolism; Animal Model; Antioxidants; Apoptosis; Asphyxia Neonatorum; Automobile Driving; Biological Assay; Blood flow; Brain; Brain Hypoxia-Ischemia; Brain Injuries; Brain region; Cardiolipins; Cell Death; Cell Survival; Charge; Clinical Research; Cre-LoxP; Critical Thinking; Development; Disease model; Environment; Enzymes; Evaluation; Exposure to; Fostering; Future; Gases; Genes; Genetic; Glucose; Homeostasis; Hypoxia; Hypoxic Brain Damage; In Vitro; Infant; Injury; Ischemia; Knock-out; Lipids; Lipolysis; LoxP-flanked allele; Machine Learning; Mass Spectrum Analysis; Mentorship; Methodology; Methods; Mitochondria; Modeling; Modification; Molecular; Molecular Analysis; Morphology; Mus; Neonatal; Neurons; Outcome; Oxidative Stress; Oxygen; Perinatal; Periodicity; Phospholipase; Phospholipids; Play; Production; Protein Isoforms; Quality Control; Reactive Oxygen Species; Regulation; Reperfusion Injury; Reperfusion Therapy; Reporter; Reproducibility; Research; Research Proposals; Role; Sampling; Scientist; Specificity; Spectrometry; Structure; System; Techniques; Technology; Therapeutic; Tissues; Training; Transgenic Mice; Transgenic Organisms; Translational Research; cardiolipin synthase; clinically relevant; conditional knockout; cytochrome c; defined contribution; deprivation; experimental study; gain of function; hypoxia neonatorum; in vivo; interest; ion mobility; lipidomics; loss of function; machine learning pipeline; mitochondrial dysfunction; monolysocardiolipin; mortality; mouse model; neonatal brain; neonatal hypoxic-ischemic brain injury; neuron loss; novel; overexpression; oxidative damage; particle; peroxidation; phospholipase inhibitor; porcine model; prevent; pup; therapeutic target

Project Summary Neonatal hypoxia/ischemia can result in severe damage to the infant brain. Reestablishing blood flow and oxygen delivery (reperfusion) is crucial for survival. However, reperfusion induces an accumulation of reactive oxygen species (ROS) produced by the mitochondria, culminating in oxidative stress and irreversible tissue damage. Current studies suggest that cardiolipin (CL) and its remodeling via oxidative injury to monolysocardiolipin (MLCL) directly participate in activation of mitophagy and lipidic pore formation to regulate cytochrome c release and facilitate induction of programmed cell death. The objectives of this research proposal are to investigate of the roles of CL and CL modification as molecular mechanisms in hypoxic brain injury. Aim 1 will focus on establishing the clinical relevance of CL modification and investigate the role of CL modification in brain injury following HIE. Utilizing the cutting-edge technology of cyclic ion mobility spectrometry mass spectrometry (cIMS-MS) CL and its isoforms will be quantitatively analyzed following hypoxia/ischemia in a neonatal piglet model of HIE. Unlike traditional mass spectrometry, cIMS-MS separates molecules based on structure as well as mass to charge ratio (m/z) allowing detailed analysis of CL species and to directly investigate its remodeling in I/R injury. Several novel transgenic mouse-lines will also be used to manipulate CL biosynthesis and remodeling in vivo. Mouse pups will be exposed to hypoxia/ischemia, then brains will be analyzed for injury to evaluate the contribution of CL modification on outcomes following HIE. Aim 2 will further explore the mechanisms regulated by CL modification following hypoxia/ischemia, including disruption to the mitochondrial network through mitophagy and mitochondrial dynamics. Mitophagic flux will be characterized in primary culture from mice possessing the mitochondrial quality control (mitoQC) reporter and disruptions to mitochondrial morphology will be assessed with our cutting-edge machine learning based quantification methodology. Finally, the mitochondrial specific antioxidant mitoTEMPO and phospholipase inhibitors will be used in combination with our genetic mouse models to define the contribution of oxidative modification and lipolysis to CL modification following hypoxia/ischemia. Together, these experiments will establish the mechanism of post-hypoxic CL modification, interrogated the casual role of CL and MLCL in the progression of brain injury , and define potential therapeutic targets. With exceptional scientific mentorship and rigorous academic study, this research will develop a deeper understanding of the role lipidomics play in HIE. This project will allow extensive training and allow me to gain expertise in a diverse range of animal models and molecular analysis of mechanisms of brain damage. The proposed project creates a unique environment to foster the development and critical thinking of a young scientist.

项目概要 新生儿缺氧/缺血会对婴儿大脑造成严重损害。重建血流和 氧气输送（再灌注）对于生存至关重要。然而，再灌注会导致反应性物质的积累。 线粒体产生的氧簇 (ROS)，最终导致氧化应激和不可逆组织 损害。目前的研究表明心磷脂（CL）及其通过氧化损伤的重塑 单溶心磷脂（MLCL）直接参与线粒体自噬的激活和脂质孔的形成以调节 细胞色素 c 释放并促进程序性细胞死亡的诱导。本研究计划的目标 旨在研究 CL 和 CL 修饰作为缺氧性脑损伤分子机制的作用。目的 1将重点建立CL修饰的临床相关性并研究CL修饰的作用 HIE 后的脑损伤。利用循环离子淌度质谱质量的尖端技术 光谱法 (cIMS-MS) 将在缺氧/缺血后对 CL 及其亚型进行定量分析 HIE的新生仔猪模型。与传统质谱分析不同，cIMS-MS 基于以下因素分离分子： 结构以及质荷比 (m/z)，可对 CL 物质进行详细分析并直接研究 它在 I/R 损伤中的重塑。几种新型转基因小鼠品系也将用于操纵 CL 生物合成 和体内重塑。幼鼠将暴露在缺氧/缺血的环境中，然后对大脑进行损伤分析 评估 CL 修改对 HIE 后结果的贡献。目标2将进一步探索 缺氧/缺血后 CL 修饰调节的机制，包括线粒体破坏 通过线粒体自噬和线粒体动力学网络。线粒体自噬通量将在原代培养物中进行表征 来自拥有线粒体质量控制 (mitoQC) 报告基因和线粒体破坏的小鼠 将使用我们基于机器学习的尖端量化方法来评估形态。最后， 线粒体特异性抗氧化剂 mitoTEMPO 和磷脂酶抑制剂将联合使用 我们的基因小鼠模型定义了氧化修饰和脂肪分解对 CL 修饰的贡献 缺氧/缺血后。这些实验将共同建立缺氧后 CL 的机制 修改，询问 CL 和 MLCL 在脑损伤进展中的偶然作用，并定义潜在的 治疗目标。凭借卓越的科学指导和严谨的学术研究，这项研究将 更深入地了解脂质组学在 HIE 中的作用。该项目将提供广泛的培训和 让我获得各种动物模型和大脑机制分子分析的专业知识 损害。拟议的项目创造了一个独特的环境，以促进发展和批判性思维 年轻的科学家。