Targeting the Pial Collateral Circulation for Mitigation of Cerebral Ischemia

摘要 大血管闭塞后卒中严重程度的主要决定因素是： 闭塞和侧支血流的量。不幸的是，软脑膜侧支血流在患者中变化很大 急性缺血性卒中（AIS）后，与梗死体积和出血性跨膜血管瘤风险呈负相关。 直接影响溶栓、取栓效果。在我们的研究之前， 这种广泛的变化在很大程度上是未知的。事实上，我们对血管生物学的了解要少得多。 与一般动静脉循环的血管相比，我们最近发现， 小鼠妊娠晚期通过独特的血管生成过程和信号通路形成侧支， 被称为“平行生成”。由于遗传背景的差异， 导致成人侧支范围和中风严重程度的巨大差异。利用遗传 通过作图，我们确定了四个与侧枝程度变异相关的基因座，并确定了致病基因 其最大基因座上的致病SNP是新基因Rabep 2。在中风遗传学的初步分析中 数据集，我们发现人类RABEP 2的多态性与急性缺血性卒中的发病率有关 和梗死面积。为了充分支持这些回顾性研究和前瞻性研究， 在招募阶段，我们需要确定三个额外的附属QTL的因果基因， 以及其他可能存在于小鼠物种中的大效应基因座。我们也取得了初步成果 回答了一个长期存在的问题--在成人中是否可以诱导形成额外的络脉。初步结果 表明全身性缺氧和MCA闭塞都是如此。而这两者都需要Rabep 2，概括其 在胚胎中的collaterogenesis中起关键作用。以下目标延续了我们的总体目标， 了解这些独特而重要的侧支血管的生物学，并将研究结果转化为 人类及其临床护理。目的我将鉴定先前鉴定的QTL的候选基因， Canq 2、Canq 3和Canq 4，以及额外的大效应QTL，使用最近开发的多样性远交 和协作交叉参考人群。方法包括高分辨率血管造影和遗传学检查。 作图、表达和计算机分析。目的II将使用基因靶向技术来确定致病基因， 目标I中鉴定的QTL。评价结果的方法包括测定脑血流量、梗死体积、恢复时间、 神经功能和缺血性血管生成。目的III将确定从头形成的机制， 新的侧支循环（NCF）诱导缺氧后持续减少吸入O2和MCA闭塞。 这些研究还将有助于确定驱动collaterogenesis的关键基因，这些基因含有变异， 是成年人侧枝丰富度变化很大的原因。他们还被要求为研究提供动力 目前正在测试人类的正向基因。此外，他们将开辟一个新的领域， 研究表明，缺血引起的NCF可能导致治疗阻塞性疾病的新疗法。

ABSTRACT The major determinants of stroke severity after large-vessel occlusion are the location and duration of occlusion and the amount of collateral blood flow. Unfortunately, pial collateral flow varies widely in patients following acute ischemic stroke (AIS), correlating inversely with infarct volume and risk of hemorrhagic trans- formation and directly with efficacy of thrombolysis and thrombectomy. Prior to our studies, clues to the cause of this wide variation were largely unknown. In fact, much less is known about the vascular biology of collaterals, compared to vessels of the general arterio-venous circulation. We recently identified that collaterals form late in gestation in mice by a unique angiogenic process and signaling pathway, which we termed “collaterogenesis”. And that collaterogenesis varies widely due to differences in genetic background, resulting, as in humans, in large differences in collateral extent and stroke severity in the adult. Using genetic mapping, we identified four loci that link to variation in collateral extent, and determined that the causal gene and its causal SNPs at the largest locus is the novel gene, Rabep2. In preliminary analyses of stroke genetics datasets, we have found that polymorphisms in human RABEP2 link to the incidence of acute ischemic stroke and infarct size in AIS patients. To fully power these retrospective studies and also prospective studies that are in the enrollment phase, we need to identify the causal genes for the three additional collateral QTL, as well as other large-effect loci likely extant in the mouse species. We have also obtained preliminary results answering a long-standing question—can additional collaterals be induced to form in adults. Preliminary results show that systemic hypoxia and MCA occlusion both do so. And that both require Rabep2, recapitulating its critical role in collaterogenesis in the embryo. The following Aims continue our overall goal to provide a deeper understanding of the biology of these unique and important collateral vessels, and to translate the findings to humans and their clinical care. Aim I will identify the candidate genes underlying the previously identified QTL, Canq2, Canq3 and Canq4, and additional large-effect QTL, using the recently developed Diversity Outbred and Collaborative Cross reference populations. Methods include high-resolution angiography and genetic mapping, expression and in silico analyses. Aim II will use gene targeting to ascertain the causal genes at the QTL identified in Aim I. Methods to assess outcome include determination of CBF, infarct volume, recovery of neurological function and ischemic angiogenesis. Aim III will determine mechanisms of de novo formation of new collaterals (NCF) induced by hypoxia following sustained decrease in inspired O2 and by MCA occlusion. These studies will also aid identifying the key genes that drive collaterogenesis that harbor variants that underlie the wide variation in collateral abundance in the adult. They are also required to power studies currently underway to test the orthologous genes in humans. In addition, they will open up a new area of basic research, NCF induced by ischemia, which may lead to novel therapies to treat obstructive disease.