Release of Apoptogenic Proteins from Brain Mitochondria

脑线粒体释放凋亡蛋白

• 批准号: 6850275
• 负责人: Nickolay Brustovetsky
• 金额: $ 27.36万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-12-01 至 2009-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6850275
• 关键词: BCL2 gene /protein; Bax gene /protein; apoptosis; brain; calcium flux; cysteine endopeptidases; enzyme activity; exocytosis; free radical oxygen; inhibitor /antagonist; laboratory rat; lipid peroxides; membrane permeability; mitochondria; neural degeneration; neurons; neuropathology; nucleotidyltransferase; peroxidation; phospholipase A2; potassium channel; protein biosynthesis; secretion; tissue /cell culture; tissue /cell preparation

DESCRIPTION (provided by applicant): There is a fundamental gap in our understanding of the mechanisms of the release of mitochondrial apoptogenic factors induced by elevated Ca2+ and by pro-apoptotic proteins. Our long-term goal is to establish the role of mitochondria in neuronal apoptosis. The objective of this study is to delineate the mechanisms of release of apoptogenic proteins from brain mitochondria initiated by elevated Ca2+ or by pro-apoptotic proteins. The central hypothesis of the proposed research is that an increased generation of reactive oxygen species, augmentation of lipid peroxidation, activation of phospholipase A2, and K+ influx in brain mitochondria are the major processes leading to the release of apoptogenic proteins induced by elevated Ca2+ or pro-apoptotic proteins tBID and BAX. In Specific Aim 1 we will establish K+-dependent mechanisms of the Ca2+-induced swelling of brain mitochondria and release of apoptogenic proteins. Inhibitors of mitochondrial K+ channels and the adenine nucleotide translocase will be applied to isolated brain mitochondria or to cultured neurons to establish their role in the Ca2+-induced swelling, and release of apoptogenic proteins. In Specific Aim 2 we will determine the extent to which an activation of mitochondrial K+ channels and the permeability transition contributes to the release of apoptogenic factors induced by pro-apoptotic proteins tBID and BAX. Inhibitors of the permeability transition and blockers of K+ channels will be used to identify their role in the release of the apoptogenic proteins. In Specific Aim 3 we will establish the role of reactive oxygen species, lipid peroxidation and phospholipase A2 in the release of apoptogenic proteins induced by tBID and BAX. Various antioxidants and inhibitors of phospholipase A2 will be used to inhibit the release of apoptogenic proteins. In Specific Aim 4 we will determine the role of caspases in the release of apoptogenic proteins from brain mitochondria exposed to tBID and BAX. Isolated brain mitochondria exposed to tBID and BAX and treated with recombinant caspases will be used to test this hypothesis. The proposed research lays the foundation for a better understanding of the molecular mechanisms of the permeabilization of the outer mitochondrial membrane induced by elevated Ca 2+ or pro-apoptotic proteins tBID and BAX and contributes to filling in a gap in our knowledge of these phenomena.

描述（由申请人提供）：我们对由升高的Ca2+和促凋亡蛋白诱导的线粒体促凋亡因子释放机制的理解存在根本性差距。我们的长期目标是确定线粒体在神经元凋亡中的作用。本研究的目的是描绘从脑线粒体中释放促凋亡蛋白的机制，由升高的Ca 2+或促凋亡蛋白启动。该研究的中心假设是，活性氧的产生增加，脂质过氧化作用增强，磷脂酶A2的活化，以及脑线粒体中的K+内流是导致由升高的Ca 2+或促凋亡蛋白tBID和BAX诱导的促凋亡蛋白释放的主要过程。在具体目标1中，我们将建立Ca2+诱导的脑线粒体肿胀和促凋亡蛋白释放的K+依赖性机制。线粒体K+通道和腺嘌呤核苷酸移位酶的抑制剂将应用于分离的脑线粒体或培养的神经元，以确定它们在Ca2+诱导的肿胀和促凋亡蛋白的释放中的作用。在具体目标2中，我们将确定线粒体K+通道的激活和渗透性转换在多大程度上有助于促凋亡蛋白tBID和BAX诱导的促凋亡因子的释放。将使用渗透性转换抑制剂和K+通道阻滞剂来确定它们在促炎蛋白释放中的作用。在具体目标3中，我们将确定活性氧、脂质过氧化和磷脂酶A2在tBID和BAX诱导的致凋亡蛋白释放中的作用。将使用各种抗氧化剂和磷脂酶A2抑制剂来抑制致炎蛋白的释放。在特定目标4中，我们将确定半胱天冬酶在暴露于tBID和BAX的脑线粒体中释放促凋亡蛋白中的作用。 将使用暴露于tBID和BAX并用重组半胱天冬酶处理的分离脑线粒体来检验该假设。该研究为更好地理解由升高的Ca 2+或促凋亡蛋白tBID和BAX诱导的线粒体外膜透化的分子机制奠定了基础，并有助于填补我们对这些现象的知识空白。