ROLE OF PROHIBITIN IN ISCHEMIC BRAIN INJURY

描述（由申请人提供）：prohibition （PHB）是一种线粒体内膜蛋白，可通过稳定复合物I（电子进入呼吸链的点）的功能和减少线粒体活性氧的产生来保持细胞完整性。在一项蛋白质组学研究中，我们试图确定在小鼠缺血耐受模型中表达的潜在神经保护蛋白，我们发现PHB在神经元线粒体中的表达增加。这些观察结果提出了一种可能性，即预处理模型中PHB的上调促进了神经元存活，而缺血模型中PHB的下调促进了神经元死亡。因此，这项应用的长期目标是阐明PHB在脑缺血引起的脑损伤中的作用，并评估其神经保护潜力。特别是，我们将验证PHB的中心假设，即通过影响线粒体对损伤的抵抗力，PHB是缺血性脑中神经元命运的关键决定因素。本实验将采用体外（氧-葡萄糖剥夺）和体内（短暂性前脑缺血）脑缺血损伤模型。病毒基因转移和小干扰RNA （siRNA）将用于增加或减少PHB在神经元培养物或小鼠海马中的表达。将在神经元培养物或分离线粒体中评估线粒体功能，以探索PHB作用的机制。以下假设将被验证：(a)缺氧缺血下调PHB，这种降低降低了内源性防御机制，并可能增加大脑对损伤的易感性；(b) PHB在神经元培养物中的表达具有神经保护作用，而其下调则增加了对损伤的易感性；(c)小鼠海马中PHB的表达可保护易损神经元免受短暂性前脑缺血的损伤；(d) PHB的神经保护作用机制包括复合物I的稳定和线粒体活性氧的减少。

DESCRIPTION (provided by applicant): Prohibitin (PHB) is a mitochondrial inner membrane protein that may preserve cellular integrity by stabilizing the function of complex I, the electrons entry point into the respiratory chain, and reducing production of mitochondrial reactive oxygen species. In a proteomic study seeking to identify potential neuroprotective proteins expressed in murine models of ischemic tolerance, we found that PHB expression is increased in neuronal mitochondria. These observations raise the possibility that PHB upregulation in preconditioning models promotes neuronal survival, while its downregulation in ischemia facilitates neuronal death. Thus, the long-term objectives of this application are to elucidate the roles of PHB in the brain damage produced by cerebral ischemia and to assess its neuroprotective potential. In particular, we will test the central hypothesis that PHB, by influencing the mitochondrial resistance to injury, is a key determinant of neuronal fate in the ischemic brain. The proposed experiments will use in vitro (oxygen-glucose deprivation), and in vivo (transient forebrain ischemia) models of cerebral ischemic injury. Viral gene transfer and small interfering RNA (siRNA) will be used to increase or decrease PHB expression in neuronal cultures or in the mouse hippocampus. Mitochondrial function will be assessed in neuronal cultures or in isolated mitochondria to explore the mechanisms of the effect of PHB. The following hypotheses will be tested: (a) Hypoxia-ischemia downregulates PHB, a reduction that decreases endogenous defense mechanisms and may increase the susceptibility of the brain to injury; (b) Expression of PHB in neuronal cultures is neuroprotective, while its downregulation increases vulnerability to injury; (c) Expression of PHB in the mouse hippocampus protects vulnerable neurons from the damage produced by transient forebrain ischemia; (d) The mechanisms of the neuroprotective effect of PHB involve complex I stabilization and reduced production of mitochondrial reactive oxygen species. PUBLIC HEALTH RELEVANCE: The proposed studies will investigate a novel aspect of the pathobiology of PHB, related to its role in the death and survival of ischemic neurons. The findings will advance our understanding of the fundamental processes regulating ischemic neuronal death, and have the potential of identifying new treatment strategies for ischemic stroke.