MicroRNAs and post-stroke angiogenesis

 描述（由申请人提供）：中风是美国死亡和残疾的主要原因。不幸的是，目前的治疗选择受到严重限制。新的证据表明，缺血后血管生成在脑卒中后血流恢复和神经元代谢中起着至关重要的作用。因此，通过各种方法促进血管生成似乎是实验性缺血性卒中的有希望的治疗方法。 microRNAs（miRs）是一类新型的非编码小分子RNA，能够负调控蛋白质的表达。miR与多种人类神经系统疾病有关。最近的研究表明miR在调节血管生成中具有重要作用。我们和其他人已经表明miR参与缺血性脑损伤的发病机制，表明miR作为卒中的潜在治疗靶点。然而，miR分子在脑卒中后血管生成过程中的调节作用和机制尚不清楚。miR-15 a/16-1簇是第一个被鉴定的与人类癌发生相关的miR组。最近，在中风患者中发现血浆miR-15 a/16-1水平失调，显示出作为诊断和预后生物标志物的巨大潜力。抑制miR-15已被证明可以防止心肌梗死（MI）。因此，几家制药公司（MiRagen therapeutics; Servier）认为miR-15是开发用于改善MI后恢复的基于miR的药物的重要miR靶标。我们首次报道了内皮miR-15 a可以显著抑制后肢缺血中的细胞自主血管生成。此外，我们的初步研究表明，miR-15 a/16-1簇的水平在小鼠大脑中动脉闭塞（MCAO）后7 d的半暗区的脑血管中显著增加。值得注意的是，EC选择性miR-15 a/16-1转基因过表达导致MCAO后7天小鼠脑微血管减少和脑梗死增加。此外，我们发现miR-15 a/16-1簇可以与血管内皮生长因子（VEGF）mRNA的3 '-UTR结合，并抑制其蛋白表达。这些发现提示了一个中心假设，即miR-15 a/16-1簇在缺血后脑血管生成中起关键调节作用，从而影响缺血性卒中后的长期神经学结局。本提案将实现三个目标。目标1：确定miR-15 a/16-1簇在调节卒中后血管生成中的功能作用;目的2：确定miR-15 a/16-1簇在调节卒中后血管生成中的分子靶点;目的3：确定miR-15 a/16-1介导的血管生成是否影响卒中的长期结局。

 DESCRIPTION (provided by applicant): Stroke is a major cause of death and disability in the US. Unfortunately, current therapeutic options are seriously limited. Emerging evidence shows that post-ischemia angiogenesis plays a crucial role in the recovery of blood flow and neuronal metabolism after stroke. Thus, promoting angiogenesis via various approaches appears as a promising treatment for experimental ischemic stroke. MicroRNAs (miRs) function as a novel class of noncoding small RNAs that negatively modulate protein expression. MiRs have been implicated in a variety of human neurological diseases. Recent studies have revealed important roles for miRs in regulating angiogenesis. We and others have shown the involvement of miRs in the pathogenesis of ischemic brain injury, suggesting miRs as potential therapeutic targets in stroke. However, the functional significance and mechanisms of miR molecules in the regulation of angiogenic processes after stroke are poorly understood. The miR-15a/16-1 cluster is the first identified miR group associated with human carcinogenesis. Recently, dysregulated plasma miR-15a/16-1 levels are found in stroke patients, showing great potential as diagnostic and prognostic biomarkers. Inhibition of miR-15 has been shown to protect against myocardial infarction (MI). Thus, several pharmaceutical companies (MiRagen therapeutics; Servier) consider miR-15 as an important miR target to develop miR-based drugs for improving post-MI recovery. We reported for the first time that endothelial miR-15a can significantly suppress cell-autonomous angiogenesis in hindlimb ischemia. Moreover, our preliminary studies showed that the levels of the miR-15a/16-1 cluster are significantly increased in the cerebral vasculature at the penumbral area 7d after middle cerebral artery occlusion (MCAO) in mice. Of note, EC-selective miR-15a/16-1 transgenic overexpression leads to reduced cerebral microvessels and increased brain infarction in mice 7d after MCAO. Furthermore, we found that the miR-15a/16-1 cluster can bind to the 3'-UTR of vascular endothelial growth factor (VEGF) mRNA and inhibit its protein expression. These findings prompt the central hypothesis that the miR-15a/16-1 cluster functions as a critical regulator in post-ischemic cerebral angiogenesis, thus affecting long-term neurological outcomes after ischemic stroke. Three aims will be performed in this proposal. Aim 1: Determine the functional role of the miR-15a/16-1 cluster in regulating post-stroke angiogenesis; Aim 2: Identify the molecular targets of the miR- 15a/16-1 cluster in regulating post-stroke angiogenesis; Aim 3: Determine whether miR-15a/16-1-mediated angiogenesis affects long-term stroke outcomes.