Novel Targets for Stroke Intervention - Gene Discovery for Modulators of Infarction

供应大脑的血管闭塞导致缺血性中风， 梗塞-脑组织的不可逆死亡。导致中风的危险因素，特别是涉及血脂的因素 代谢，形成目前治疗的基础，以减少中风的风险。尽管几十年来 对梗死期间发生的分子事件的研究，这些发现的翻译， 治疗中风结果（脑组织死亡）的“可药物化”目标相当令人失望。小说 将需要采取各种办法来确定新的和与生理学更相关的目标。科学 我们的建议的前提是，自然发生的等位基因变异的深刻差异的基础， 这些神经保护基因变异将提供一种新的途径， 中风治疗的新目标然而，在脑梗死患者中， 人（例如，缺血性卒中患者梗死体积的GWAS）本质上是有问题的， 闭塞血管的范围和位置的广泛变化，特别是时间的变化， 第一次识别症状和医疗干预之间的窗口。到目前为止，我们还没有发现任何出版的 缺血性卒中梗死体积的GWAS。Marchuk实验室采用了一种替代的， 发现新的调节梗死的基因的方法。我们已经通过手术封闭了 在超过35个近交系小鼠品系的脑动脉中，发现梗死体积差异超过50倍。 这些在梗死面积上的强有力的和高度可重复的差异至少比所看到的大10倍 但是，重要的是，是由小鼠中的天然等位基因变异引起的 基因组我们已经绘制了其中几个遗传基因座，目标1和2的目标是确定 这些新基因调节梗死面积。然而，这种基因发现方法需要在体内 在数千只成年动物身上进行的外科试验。我们需要一个更具可扩展性的生理相关的 筛选平台，将这种方法转化为全基因组范围的规模。Lo实验室开创了 开发脑梗死的发现平台，通过氧/葡萄糖模拟中风 离体脑组织外植体中的脱髓鞘（OGD;缺血性损伤的良好表征模型）。不像 在培养的分离神经元中，脑切片外植体保留了完整器官的复杂多细胞性质， 从而保留和代表在脑组织中发生的复杂的细胞间相互作用， 脑梗死在目标1和2中，该离体OGD平台将用于鉴定致病性 我们之前绘制的基因座中的基因。我们在这些目标中获得的经验将导致目标3，即 离体OGD测定将用于直接定位和鉴定新的脑梗死基因，使用 协作杂交的遗传作图资源群体。我们的研究利用了 联合PI开发的创新方法，以实施识别新药的新策略 治疗缺血性中风的靶点。

Occlusion of the blood vessels supplying the brain leads to ischemic stroke and infarction—irreversible death of brain tissue. Risk factors causing stroke, especially those involving lipid metabolism, form the basis of current therapies to reduce stroke risk. However, despite decades of research on the molecular events occurring during infarction, the translation of these discoveries to “druggable” targets to treat stroke outcome (death of brain tissue) has been quite disappointing. Novel approaches will be required to identify new and more physiologically relevant targets. The scientific premise of our proposal is that naturally occurring allelic variation underlies the profound differences in seen in stroke outcomes and that these neuro-protective gene variants would provide a novel path towards new targets for stroke treatment. However, genetic mapping approaches for infarct size in the human (e.g., GWAS of infarct volume among ischemic stroke patients) are intrinsically problematic due to wide variation in the extent and location of the occluded vessel, and especially, variation in the time window between first recognized symptoms and medical intervention. To date, we can find no published GWAS for infarct volume in ischemic stroke. The Marchuk lab has taken an alternative, forward genetic approach to discover novel genes modulating infarction. We have surgically occluded the distal middle cerebral artery in over 35 inbred mouse strains and found that infarct volume differs more than 50-fold. These robust and highly reproducible differences in infarct size are at least 10-fold larger than that seen in any engineered mouse lines but, importantly, are caused by natural allelic variation in the mouse genome. We have mapped several of these genetic loci and the goals of Aims 1 and 2 are to identify these novel genes regulating in infarct size. However, this gene discovery approach has required in vivo surgical assays in thousands of adult animals. We need a more scalable yet physiologically relevant screening platform to transform this approach to full genome-wide scale. The Lo lab has pioneered the development of such a discovery platform for cerebral infarction, simulating stroke by Oxygen/Glucose Deprivation (OGD; a well-characterized model for ischemic injury) in ex vivo brain tissue explants. Unlike isolated neurons in culture, brain slice explants retain the complex multicellular nature of the intact organ, and thus retain and represent the complex intercellular interactions occurring in brain tissue during cerebral infarction. In Aims 1 and 2, this ex vivo OGD platform will be used to identify the causative genes in our previously mapped loci. Our experience gained in these aims will lead to Aim 3, where the ex vivo OGD assay will be used to directly map and identify novel cerebral infarction genes, using the genetic mapping resource population of the Collaborative Cross. Our study takes advantage of innovative approaches developed by the co-PIs to implement a novel strategy for identifying novel drug targets to treat ischemic stroke.