Mechanisms of retinal angiogenesis

描述（由申请人提供）：在理解病理性视网膜血管生成的机制中，我们发现除了VEGFA、VEGFB、VEGFC、DLL 4和裂解的NOTCH 1诱导之外，缺氧以稳健和持续的方式激活小鼠视网膜中的CREB和STAT 3。足够有趣的是，CREB或STAT 3的阻断抑制VEGFC和FGF 2诱导的DLL 4和切割的NOTCH 1在HRMVEC中的表达。大量的研究表明，NOTCH信号通过调节尖端与柄细胞分化在发育和出生后血管生成中起着关键作用。CREB属于碱性/亮氨酸拉链（basic/leucine zipper，bZIP）转录因子家族，具有结合基因启动子区域中的共有序列TGAC/GTCA的能力，从而响应于多种刺激物（包括细胞因子、生长因子和激素）增强其表达，并且参与神经元可塑性、长期记忆和细胞代谢的调节。同样，STAT 3属于转录因子家族，其活性受Tyr/Ser磷酸化和乙酰化调节，具有与基因启动子区中的共有序列TTCCGGGAA结合的能力，诱导基因的表达以响应大范围的刺激物，包括细胞因子、生长因子和激素，并且参与细胞生长和迁移的调节。虽然大量的数据证明了这些转录因子在许多细胞过程（包括细胞增殖、迁移和分化）的调节中的重要性，但是关于它们在病理性视网膜血管生成的调节中的作用，特别是与NOTCH信号传导有关的作用，实际上一无所知。基于我们新的初步观察，我们假设CREB和STAT 3通过增强DLL 4的表达，从而NOTCH 1信号传导介导病理性视网膜新生血管形成。为了解决这个假设，我们将测试以下三个具体目标：目标1。验证CREB介导缺氧诱导的病理性视网膜新生血管形成的假说。目标二。验证STAT 3介导缺氧诱导的病理性视网膜新生血管形成的假说。目标3：检验缺氧通过CREB和STAT 3诱导DLL 4-NOTCH 1信号传导，导致渗漏和非模式化病理性视网膜新生血管形成的假设。在本授权申请中提出的实验的执行应该为我们提供关于NOTCH信号传导激活在介导病理性视网膜新生血管形成中的上游机制的新信息，并且这样的知识将有助于开发针对这种使人衰弱的眼部疾病的治疗药物。

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.