Stress Proteins in Brain Cell Injury

脑细胞损伤中的应激蛋白

• 批准号: 9270635
• 负责人: Rona G Giffard
• 金额: $ 34.44万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-01 至 2018-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9270635
• 关键词: Aftercare; Animals; Apoptosis; Apoptotic; Astrocytes; BCL-2 Protein; BCL2 gene; Behavioral; Brain; Brain Injuries; Calcium; Cause of Death; Cell Death; Cell Survival; Cells; Cerebral Ischemia; Clinical; Clinical Trials; Complex; Endoplasmic Reticulum; Explosion; Family; Family member; Female; Follow-Up Studies; GRP75; GRP78 gene; Goals; Grant; Heat shock proteins; Heat-Shock Proteins 70; Homeostasis; Infusion procedures; Injection of therapeutic agent; Injury; Inner mitochondrial membrane; Ischemia; Ischemic Brain Injury; MCL1 gene; Measures; Membrane; Membrane Potentials; Messenger RNA; MicroRNAs; Mitochondria; Molecular Chaperones; Mus; Necrosis; Neuroglia; Neurologic; Neurons; Outcome; Oxygen Consumption; Pathway interactions; Patients; Protein Family; Proteins; RNA Interference; Recovery; Regulation; Reporting; Research; Role; Site; Stress; Stroke; Testing; Translating; United States; Work; behavioral outcome; brain cell; cell injury; cytochrome c; disability; effective therapy; human disease; improved; in vivo; inhibitor/antagonist; male; mitochondrial membrane; neuronal survival; new therapeutic target; novel; overexpression; public health relevance; stress protein; thrombolysis; uptake

DESCRIPTION (provided by applicant): Stroke is one of the leading causes of death worldwide and a major cause of long-term disability. Although many clinical trials have targeted stroke patients, only thrombolysis has so far emerged as an effective treatment. Nonetheless, new therapeutic targets have emerged. Calcium dysregulation has long been implicated in brain injury in the setting of stroke. Our prior work on this grant, as well as work from others, has shown that the heat shock protein 70 kDa (HSP70) family of chaperones or stress proteins, even if expressed after ischemia, can protect the brain. The regulation of apoptosis by the BCL2 family of proteins is another important determinant of ischemic outcome. These pathways converge to control cellular calcium homeostasis and both apoptotic and necrotic cell death. The endoplasmic reticulum (ER) and mitochondria interact at specific sites called the mitochondrial associated ER membrane (MAM) to regulate cellular calcium homeostasis and cell death. MAM is a critical site of stress/chaperone protein and BCL2 family interaction, and is central to determining the outcome from cerebral ischemia. We recently found that overexpressing GRP78, an ER chaperone and HSP70 family member, preserves mitochondrial function, reduces mitochondrial Ca2+ overload, and improves brain cell survival after stress. We also found that GRP78 translocates to the mitochondrial inner membrane after stress in astrocytes, CNS glial cells that are known to be integral to neuronal survival. In Aim 1 of this proposal we will investigate the role of translocated GRP78 in mitochondrial function by comparing wild type and mitochondrially targeted GRP78. The discovery of posttranscriptional gene silencing by miRNAs has led to an explosion of new hypotheses in human disease. Studies of miRNAs in cerebral ischemia are recent, and most have focused on profiling changes in miRNAs. We recently reported that reducing or blocking miR-181, a brain-enriched miRNA, protects the brain from stroke in the initial post-injury period. Aim 2 is a translational aim and ill follow up these studies to determine the effects of altering miR-181 on long term behavioral outcome from stroke, test post-treatment, and determine whether miR-181 is also effective in female animals. We recently demonstrated that miR-181 can target both GRP78 and anti-apoptotic BCL2 family members BCL2 and MCL1. Therefore, the role of miR-181 in ER-mitochondrial calcium transfer in MAM will be studied. Using computational miRNA target prediction we identified miR-200 as potentially targeting GRP75, a mitochondrial chaperone, and BCL2. Aim 3 of this proposal will focus on the mechanism of protection by reducing miR-181 levels investigating in detail effects on ER and mitochondrial Ca2+, and mitochondrial function. Overall this proposal exams a novel hypothesis: that miRNAs act as master regulators of chaperones and BCL2 family members influencing Ca2+ homeostasis, mitochondria-ER crosstalk, and outcome after cerebral ischemia.

描述（由申请人提供）：中风是全球死亡的主要原因之一，也是长期残疾的主要原因之一。虽然许多临床试验针对中风患者，但迄今为止只有溶栓作为一种有效的治疗方法出现。尽管如此，新的治疗靶点已经出现。长期以来，钙失调一直被认为与中风的脑损伤有关。我们之前的工作以及其他人的工作表明，热休克蛋白70 kDa （HSP70）家族的伴侣蛋白或应激蛋白，即使在缺血后表达，也可以保护大脑。BCL2家族蛋白对细胞凋亡的调控是缺血性预后的另一个重要决定因素。这些途径汇聚控制细胞钙稳态和凋亡和坏死细胞死亡。内质网（ER）和线粒体在称为线粒体相关ER膜（MAM）的特定部位相互作用，调节细胞钙稳态和细胞死亡。MAM是应激/伴侣蛋白和BCL2家族相互作用的关键位点，是决定脑缺血结局的核心。我们最近发现，过表达GRP78 （ER伴侣和HSP70家族成员）可以保护线粒体功能，减少线粒体Ca2+超载，并提高应激后脑细胞的存活率。我们还发现，在星形胶质细胞和中枢神经胶质细胞中，GRP78在应激后易位到线粒体内膜，而星形胶质细胞是神经元存活不可或缺的一部分。在本提案的目的1中，我们将通过比较野生型和线粒体靶向型GRP78来研究易位GRP78在线粒体功能中的作用。mirna转录后基因沉默的发现导致了人类疾病新假说的爆发。脑缺血中mirna的研究是最近才开始的，大多数研究都集中在mirna的变化分析上。我们最近报道，减少或阻断miR-181（一种富含大脑的miRNA）可以保护大脑在损伤后的初始阶段免受中风的影响。目的2是一个转化目的，我们将对这些研究进行随访，以确定改变miR-181对中风后长期行为结局的影响，治疗后测试，并确定miR-181是否对雌性动物也有效。我们最近证明miR-181可以靶向GRP78和抗凋亡BCL2家族成员BCL2和MCL1。因此，我们将研究miR-181在MAM中er -线粒体钙转移中的作用。通过计算miRNA靶标预测，我们发现miR-200可能靶向线粒体伴侣GRP75和BCL2。本提案的目标3将侧重于通过降低miR-181水平来研究对ER和线粒体Ca2+以及线粒体功能的详细影响的保护机制。总的来说，这一提议检验了一个新的假设：mirna作为伴侣和BCL2家族成员的主要调节剂，影响Ca2+稳态、线粒体-内质网串扰和脑缺血后的预后。