Mechanisms of retinal angiogenesis

描述(申请人提供)：在了解病理性视网膜血管生成的机制中，我们发现除了VEGFA、VEGFB、VEGFC、DLL4和裂解的NOTCH1诱导外，低氧还以一种强劲和持续的方式激活了小鼠视网膜中的CREB和STAT3。有趣的是，阻断CREB或STAT3均可抑制VEGFC和FGF2诱导的HRMVECs中DLL4和NOTCH1的表达。大量的研究表明，Notch信号通过调节茎尖细胞的分化在发育和出生后的血管生成中起着关键作用。CREB属于碱性/亮氨酸拉链(BZIP)转录因子家族，能够与TGAC/GTCA共同序列结合在基因启动子区域，增强其对多种刺激因子的反应，包括细胞因子、生长因子和激素，参与神经元可塑性、长期记忆和细胞代谢的调节。同样，STAT3属于转录因子家族，其活性受Tyr/Ser磷酸化和乙酰化调控，在诱导其表达的基因的启动子区域具有与TTCCGGGAA共同序列结合的能力，对包括细胞因子、生长因子和激素在内的广泛刺激做出反应，参与细胞生长和迁移的调节。虽然大量的数据表明这些转录因子在许多细胞过程的调节中的重要性，包括细胞的增殖、迁移和分化，但对于它们在病理性视网膜血管生成的调节中的作用，特别是在具有Notch信号的肿瘤中的作用，从字面上讲还不清楚。根据我们新的初步观察，我们假设CREB和STAT3通过增强DLL4的表达从而介导NOTCH1信号转导病理性视网膜新生血管。为了解决这一假设，我们将检验以下三个具体目标：目的1.检验CREB介导低氧诱导的病理性视网膜新生血管的假说。目的2.验证STAT3介导低氧诱导的病理性视网膜新生血管的假说。目的3.验证低氧通过CREB和STAT3诱导DLL4-NOTCH1信号通路导致渗漏和非图案性病理性视网膜新生血管的假说。这项拨款申请中提出的实验的实施将为我们提供关于Notch信号激活在介导病理性视网膜新生血管中的上游机制的新信息，这些知识将有助于开发治疗这种削弱眼部疾病的药物。

DESCRIPTION (provided by applicant): In understanding the mechanisms of pathological retinal angiogenesis, we have discovered that besides VEGFA, VEGFB, VEGFC, DLL4 and cleaved NOTCH1 induction, hypoxia activates CREB and STAT3 in the mouse retina in a robust and sustained manner. Interestingly enough, blockade of either CREB or STAT3 inhibits VEGFC and FGF2-induced expression of DLL4 and cleaved NOTCH1 in HRMVECs. A large number of studies suggest that NOTCH signaling by modulating the tip versus stalk cell differentiation plays a pivotal role in developmental and postnatal angiogenesis. CREB belongs to a basic/leucine zipper (bZIP) family of transcriptional factors and possess the ability to bind o the consensus sequence TGAC/GTCA in the promoter regions of genes enhancing their expression in response to a wide variety of stimulants, including cytokines, growth factors and hormones and is involved in the regulation of neuronal plasticity, long-term memory and cellular metabolism. Similarly, STAT3, which belongs to a family of transcriptional factors and whose activity is regulated by Tyr/Ser phosphorylation and acetylation, possess the ability to bind to th consensus sequence TTCCGGGAA in the promoter regions of genes inducing their expression in response to a large spectrum of stimulants, including cytokines, growth factors and hormones and is involved in the modulation of cellular growth and migration. While a large body of data demonstrates the importance of these transcriptional factors in the regulation of many cellular processes, including cell proliferation, migration and differentiation, literally nothing is known n regard to their role in the regulation of pathological retinal angiogenesis, particularly in concer with NOTCH signaling. Based on our novel preliminary observations, we hypothesize that CREB and STAT3 via enhancing the expression of DLL4 and thereby NOTCH1 signaling mediates pathological retinal neovascularization. To address this hypothesis, we will test the following three specific aims: Aim 1. To test the hypothesis that CREB mediates hypoxia-induced pathological retinal neovascularization. Aim 2. To test the hypothesis that STAT3 mediates hypoxia-induced pathological retinal neovascularization. Aim 3. To test the hypothesis that hypoxia induces DLL4-NOTCH1 signaling via CREB and STAT3 leading to leaky and non- patterning pathological retinal neovascularization. The execution of the experiments proposed in this grant application should provide us novel information on the upstream mechanisms of NOTCH signaling activation in mediating pathological retinal neovascularization and such knowledge would be useful in the development of therapeutic drugs against this debilitating ocular disease.