Oligo-Vascular Signaling in Stroke

描述(由申请人提供)：中风白质损伤中的少血管信号是包括中风和血管性痴呆在内的各种中枢神经系统疾病的病理生理学中的中心事件。但在脑血管研究中，研究脑白质的研究相对较少。越来越多的证据表明，脑内皮细胞除了具有传导血流的功能外，还具有多种功能。这里，我们假设脑内皮细胞分泌营养因子以维持少突胶质细胞(OL)和少突胶质前体细胞(OPC)的生存和功能。我们的三个目标是：目标1：剖析内皮源性生长因子寡聚保护的细胞机制。将培养的大鼠OL/OPC置于缺氧缺糖状态。内皮条件培养液(Endo-CM)将用于检测来自内皮细胞的生长因子是否能保护OL/OPC培养。我们将研究Endo-CM如何促进OL/OPC中的生存信号(如Akt)和减少细胞凋亡。目的2：显示氧化应激减少内皮生长因子的产生。我们假设，即使没有明显的细胞死亡，氧化应激内皮细胞也会减少生长因子的产生。大脑内皮细胞将暴露在氧化应激下，我们比较了来自健康内皮细胞的条件培养液和氧化应激内皮细胞。氧化应激内皮细胞的条件培养基应含有较少的生长因子，因此，对OL/OPC的保护作用较差。目的3：在体内证实内皮生长因子在OL/OPC中的重要作用。在不同的小鼠脑缺血模型中，我们将评估胼胝体和纹状体的OL/OPC完整性(即脑白质损伤)。血管内皮细胞特异性基因传递方法(脂质体，scAAV9)将被用来调节体内的内皮营养因子。我们预测，选择性地增加内皮营养因子可以保护脑白质，而减少内皮营养因子会使脑白质更脆弱。我们的实验数据(ARAI和LO，J Neurosci 2009)表明，Endo-CM通过特定的信号通路支持OPC的增殖，氧化应激干扰少血管偶联。在这项提案中，我们将以这些初步发现为基础，证明Endo-CM可以真正防止OL/OPC中的细胞死亡。最重要的是，我们的目标是在体内证明寡血管偶联可以预防中风。这些实验应该提供证据，证明内皮和OL/OPC之间的营养偶联维持和保护白质。

DESCRIPTION (provided by applicant): Oligo-Vascular Signaling in Stroke White matter injury is a central event in the pathophysiology of diverse CNS disorders, including stroke and vascular dementia. But studies that investigate white matter are relatively uncommon in cerebrovascular research. Accumulating evidence suggest that cerebral endothelial cells have multiple functions in addition to conducting blood flow. Here, we hypothesize that cerebral endothelial cells secrete trophic factors to maintain oligodendrocyte (OL) and oligodendrocyte precursor cell (OPC) survival and function. Our 3 aims are: Aim 1: Dissect the cellular mechanisms of oligo-protection by endothelial-derived growth factors. Cultured rat OL/OPC will be subjected to oxygen-glucose deprivation. Endothelial conditioned media (Endo-CM) will be used to test whether growth factors from endothelial cells can protect OL/OPC cultures. We will investigate how Endo-CM promotes survival signaling (e.g. Akt) and decreases apoptosis in OL/OPC. Aim 2: Show that oxidative stress decreases endothelial growth factor production. We hypothesize that even without overt cell death, oxidatively-stressed endothelial cells will have reduced growth factor production. Cerebral endothelial cells will be exposed to oxidative stress, and we compare conditioned media from healthy endothelial cells vs oxidatively-stressed endothelial cells. Conditioned media from oxidatively-stressed endothelial cells should have less growth factors and thus, be less protective for OL/OPC against insults. Aim 3: Demonstrate that endothelial growth factors are important for OL/OPC in vivo. In various mouse models of cerebral ischemia, we will assess OL/OPC integrity in corpus callosum and striatum (i.e. white matter damage). Endothelial-specific gene delivery methods (liposome, scAAV9) will be used to modulate endothelial trophic factors in vivo. We predict that selectively increasing endothelial trophic factors protect white matter, whereas decreasing endothelial trophic factors makes white matter more vulnerable. Our pilot data (Arai and Lo, J Neurosci 2009) suggest that Endo-CM supports OPC proliferation via specific signaling pathways and oxidative stress interferes with oligovascular coupling. In this proposal, we will build on these initial findings to show that Endo-CM can truly prevent cell death in OL/OPC. And most importantly, we aim to show that oligovascular coupling protects against stroke in vivo. These experiments should provide evidence to show that trophic coupling between endothelium and OL/OPC maintains and protects white matter.