Stress Proteins in Brain Cell Injury

脑细胞损伤中的应激蛋白

• 批准号: 8580868
• 负责人: Rona G Giffard
• 金额: $ 34.43万
• 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/8580868
• 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; RNA Interference; Recovery; Regulation; Reporting; Research; Role; Site; Stress; Stroke; Testing; Translating; United States; Work; 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）：中风是全世界死亡的主要原因之一，也是长期残疾的主要原因。尽管许多临床试验都针对中风患者，但迄今为止只有溶栓治疗成为有效的治疗方法。尽管如此，新的治疗靶点已经出现。钙失调长期以来一直被认为与中风时的脑损伤有关。我们之前在这项资助上的工作以及其他人的工作表明，伴侣蛋白或应激蛋白的热休克蛋白 70 kDa (HSP70) 家族，即使在缺血后表达，也可以保护大脑。 BCL2 蛋白家族对细胞凋亡的调节是缺血结果的另一个重要决定因素。这些途径汇聚在一起控制细胞钙稳态以及细胞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) 的特定位点相互作用，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，对于确定脑缺血的结果至关重要。我们最近发现，过度表达 GRP78（ER 伴侣和 HSP70 家族成员）可以保留线粒体功能，减少线粒体 Ca2+ 过载，并提高应激后脑细胞的存活率。我们还发现，星形胶质细胞（已知对神经元存活至关重要的中枢神经系统胶质细胞）受到应激后，GRP78 易位至线粒体内膜。在本提案的目标 1 中，我们将通过比较野生型和线粒体靶向 GRP78，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引发了人类疾病新假说的爆发。脑缺血中 miRNA 的研究是最近才进行的，大多数都集中在 miRNA 谱的变化上。我们最近报道，减少或阻断 miR-181（一种大脑富集的 miRNA）可以保护大脑在损伤后初期免受中风。目标 2 是一个转化目标，我们将跟进这些研究，以确定改变 miR-181 对中风长期行为结果的影响，测试治疗后，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，将研究 miR-181 在 MAM 内质网线粒体钙转移中的作用。通过计算 miRNA 靶标预测，我们确定 miR-200 可能靶向线粒体伴侣 GRP75 和 BCL2。该提案的目标 3 将重点关注通过降低 miR-181 水平的保护机制，详细研究对 ER 和线粒体 Ca2+ 以及线粒体功能的影响。总体而言，该提案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因子，影响 Ca2+ 稳态、线粒体 - ER 串扰和脑缺血后的结果。