Neuroprotective Genes in Ischemic Tolerance

项目1：局灶性缺血影响脑内基因表达，影响预后。先兆亚致死性局灶性脑缺血可诱导神经保护基因产生耐受性。缺血耐受也可由全身缺血和全身内毒素引发的亚致死应激引起。我们的目标是发现在这些范例中产生耐受性的基因子集。我们以前研究脑缺血基因调控的方法是减法杂交和克隆。在过去的五年里，我们克隆了27个新的基因。我们现在将使用微阵列技术来提高基因发现的能力，并将重点放在缺血耐受模型中的保护基因上。我们已经开发了新的小鼠耐受性模型，以便使用小鼠基因芯片并利用小鼠基因组的力量。我们的方法是使用几个模型，每个模型都产生耐受性，但在大脑中有不同的刺激或区域目标。我们将描述受先前短暂局灶缺血或内毒素启动保护的皮质对局灶缺血的耐受性，并将描述受先前短暂全局缺血保护的ca1神经元对全局缺血的耐受性。多个模型和复制将允许对数据进行复杂的统计分析。

Project 1: Focal ischemia affects gene expression in brain, and such gene expression affects outcome. Neuroprotective genes are induced by premonitory sublethal focal ischernia resulting in tolerance. Ischemic tolerance also results from sublethal stress caused by global ischemia and by systemic endotoxin priming. Our goal is to discover the subset of genesthat produce tolerance in each of these paradigms. Our previous approach to gene modulation in brain ischenaia was subtractive hybridization and cloning. We cloned 27 new genes over the past five years. We will now use microarray technology to increase our capabilities for gene discovery and focus the efforts on protective genes modeled in ischemic tolerance. We have developed new mouse models of tolerance in order to use murine gene chips and exploit the power of the mouse genome. Our approach is to use several models, each of which results in tolerance but with distinct stimuli or regional targets in brain. We will characterize tolerance to focal ischemia in cortex protected by prior brief focal ischemia or endotoxin priming, and will characterize tolerance to global ischemia in CA 1 neurons protected by prior brief global ischemia. Multiple models and replicates will permit sophisticated statistical analysis of data. Established core facilities for genomics will enable optimal data management and analysis. New informatics and data mining collaborators at Pacific Northwest National Laboratories Supercomputer and Bioinformatics divisions adds substantial strength to our data analysis capabilities. Our goal is to find endogenous pathways of neuroprotection modeled in tolerance. To this end we will identify genes and pathways regulated during tolerance by preconditioning focal ischemia (Aim 1); identify genes and pathways regulated during tolerance by endotoxin priming (Aim 2); and identify genes and pathways regulated during tolerance in the selectively vulnerable neurons of the CA 1 sector following global ischemia (Aim 3). Genes of known and unknown function discovered in these tolerance models will be advanced to projects two and three for validation in in vitro and in vivo systems based on a specific strategic plan.The elucidation of endogenous protective systems should identify novel approaches for stroke treatment.