Astrocytes and ischemic brain injury

星形胶质细胞和缺血性脑损伤

• 批准号: 8440740
• 负责人: Rona G Giffard
• 金额: $ 35.33万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-03-15 至 2017-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8440740
• 关键词: Address; Affect; Aftercare; Algorithms; Apoptosis; Apoptotic; Astrocytes; BCL2 gene; Behavioral; Biological Assay; Brain; Brain Ischemia; Cell Death; Cells; Cerebral Ischemia; Cessation of life; Coculture Techniques; Failure; Family; Family member; Fright; Functional disorder; Funding; Gene Expression; Gene Family; Gene Targeting; Glial Fibrillary Acidic Protein; Heart Arrest; Hippocampus (Brain); Impairment; In Vitro; Individual; Injury; Ischemia; Ischemic Brain Injury; Label; Life; Luciferases; Matrix Metalloproteinases; Messenger RNA; MicroRNAs; Mitochondria; Modeling; Mutate; Neurologic; Neurological outcome; Neurons; Nucleotides; Outcome; Oxidative Stress; Patients; Prosencephalon; Protein Family; Proteins; Publishing; Reactive Oxygen Species; Regulation; Reperfusion Therapy; Reporter; Research; Resuscitation; Role; Seeds; Signal Transduction; Small Interfering RNA; Stress; Survivors; Testing; Time; Translating; Untranslated Regions; Up-Regulation; Work; cell type; clinical application; cytochrome c; dentate gyrus; disability; genetic regulatory protein; improved; in vivo; member; mitochondrial membrane; neuronal survival; neuroprotection; novel; prevent; response; small hairpin RNA; stroke therapy

DESCRIPTION (provided by applicant): One of the most feared disabilities in survivors of cardiac arrest is neurological impairment. Even brief global cerebral ischemia causes delayed loss of CA1 hippocampal neurons while sparing nearby hippocampal dentate gyrus (DG) neurons. Lack of consideration of the role of astrocytes is thought to be a factor in the failure o many potential stroke therapies aimed only at neuronal survival (Nedergaard and Dirnagl, 2005). In the prior funding period we demonstrated selective dysfunction of hippocampal CA1 astrocytes at early reperfusion times, long before CA1 neuron death. We published the first demonstration that potent protection of CA1 neurons can be achieved by targeting protective proteins selectively to hippocampal astrocytes. Although delayed CA1 neuronal death after transient ischemia is apoptotic, the regulation of the critically important BCL2 family following cerebral ischemia is not well understood. BCL2 family proteins are central regulators of the life/death decision in cells (Adams et al, 2007), and via their regulation of mitochondrial membrane integrity and function control apoptosis signaling. MicroRNAs (miRNAs) are a novel, abundant class of ~22-nucleotide RNAs that control gene expression post- transcriptionally. Although hundreds of miRNAs have been cloned, little is known about their real targets and functions. Numerous miRNAs are expressed in a cell-specific manner and the miR-29 family has been suggested to be astrocyte related (Smirnova et al, 2005). In this proposal we will investigate the role of miR-29 in brain ischemia and neuroprotection, evaluate manipulation of miR-29 as a novel target for neuroprotection, and assess potential targets of miR-29 in the BCL2 family. Using computational miRNA target prediction algorithms we found that miR-29 could potentially target messenger RNAs of several BCL2 family members, both pro- and anti-apoptotic. Aims 1 and 2 focus on the role of miR-29 in ischemic brain injury. We will first investigate how altering miR-29 expression changes ischemia-induced cell death in vivo and in vitro. Aim 2 will determine changes in miR-29 family member levels in response to ischemia, including cell type and hippocampal region specific changes with forebrain ischemia. We will then address the mechanisms of the miR-29 effects on cells focusing on mitochondrial function and oxidative stress. The role of miR-29 in selective CA1 astrocyte dysfunction will be tested. In addition to focusing on the role of miR-29 in outcome from injury, we need to investigate downstream targets. In Aim 3 we focus on 5 likely miR-29 targets, all members of the BCL2 family, focusing especially on pro-apoptotic PUMA. These are likely candidates for effects of miR-29 on mitochondrial function and oxidative stress. Overall this proposal will test the hypothesis that specific microRNAs are a novel and effective target for protection from ischemic brain injury, as well as advancing our understanding of the contributions of astrocyte regulation and impairment to neurological outcome following cardiac arrest and resuscitation.

描述（由申请人提供）：心脏骤停幸存者最害怕的残疾之一是神经损伤。即使是短暂的全脑缺血也会导致CA1海马神经元的延迟损失，而保留附近的海马齿状回（DG）神经元。缺乏对星形胶质细胞作用的考虑被认为是许多仅针对神经元存活的潜在卒中治疗失败的一个因素（Nedergaard和Dirnagl， 2005）。在之前的资助期内，我们证明了海马CA1星形胶质细胞在早期再灌注时的选择性功能障碍，早在CA1神经元死亡之前。我们发表了第一个证明，通过选择性地将保护蛋白靶向海马星形胶质细胞，可以实现对CA1神经元的有效保护。虽然短暂性缺血后延迟的CA1神经元死亡是凋亡，但脑缺血后至关重要的BCL2家族的调控尚不清楚。BCL2家族蛋白是细胞生死决定的中心调节因子（Adams等，2007），并通过调节线粒体膜完整性和功能控制细胞凋亡信号。MicroRNAs （miRNAs）是一类新的，丰富的约22个核苷酸的rna，在转录后控制基因表达。尽管已经克隆了数百种mirna，但人们对它们真正的目标和功能知之甚少。许多mirna以细胞特异性的方式表达，miR-29家族被认为与星形胶质细胞相关（Smirnova et al, 2005）。在本提案中，我们将研究miR-29在脑缺血和神经保护中的作用，评估miR-29作为神经保护新靶点的操作，并评估miR-29在BCL2家族中的潜在靶点。通过计算miRNA靶标预测算法，我们发现miR-29可能潜在地靶向几种BCL2家族成员的信使rna，包括促凋亡和抗凋亡。目的1和2关注miR-29在缺血性脑损伤中的作用。我们将首先在体内和体外研究改变miR-29表达如何改变缺血诱导的细胞死亡。Aim 2将确定miR-29家族成员水平在缺血反应中的变化，包括前脑缺血时细胞类型和海马区的特异性变化。然后，我们将重点讨论miR-29对线粒体功能和氧化应激细胞的影响机制。我们将测试miR-29在选择性CA1星形胶质细胞功能障碍中的作用。除了关注miR-29在损伤结果中的作用外，我们还需要研究下游靶点。在Aim 3中，我们关注了5个可能的miR-29靶点，它们都是BCL2家族的成员，特别关注促凋亡的PUMA。这些可能是miR-29对线粒体功能和氧化应激影响的候选者。总的来说，这一提议将验证特异性microrna是缺血性脑损伤保护的一个新的有效靶点的假设，并推进我们对星形胶质细胞调节和心脏骤停和复苏后神经预后损害的贡献的理解。