Mechanisms of retinal angiogenesis

以前我们已经证明了src-pld1-pkc、γ-cplA2信号通路参与了视网膜的调节 新生血管。我们的研究还表明，pak1-cofilin信号轴的激活 视网膜新生血管所必需的。此外，我们还表明，除了VEGFA之外，VEGFC通过CREB- DLL4-NOTCH1信号介导的激活在增强HRMVEC中的重要作用 体外细胞培养模型和视网膜中的迁移、增殖、萌发和管状形成 OIR模型体内新生血管的研究。在这些研究过程中，我们发现VEGFA和 低氧对PKCθ的刺激非常强烈，其水平的耗竭完全阻止了细胞的迁移和萌发 高密度脂蛋白血管内皮细胞对VEGFA的反应能力。我们还观察到VEGFA和低氧都能诱导 Flt4的表达水平很高，这种反应需要PKCθ介导的JunB的诱导。此外，两者 VEGFA和缺氧触发了CDC6表达的诱导，CDC6是复制前的调节成分 复合体(前RC)，并参与将微小染色体维护蛋白(MCM2-7)加载到 DNA，它在复制中扮演着限速步骤。此外，CDC6水平完全下调 取消VEGFA诱导的DNA合成。基于这些新发现，我们提出了pkcθ通过 通过促进EC的增殖促进EC的迁移和萌发，CDC6在 调制视网膜新生血管。为了验证这一中心假设，我们将讨论以下四个具体问题 目标。目的1.验证PKCθ介导低氧诱导的视网膜新生血管的假说。目标2.目标 验证JunB介导低氧诱导的视网膜新生血管的假设。目标3.测试 假设PKCθ-JunB信号轴以Flt4诱导为靶点，介导视网膜新生血管形成。目标4. 验证CDC6在视网膜新生血管中起重要作用的假说。评选结果 拟议中的实验可能会提供有关以下分子机制的新信息 病理性视网膜新生血管，可能导致新的治疗化合物的开发 对抗这种眼病。

Previously we have demonstrated that Src-PLD1-PKCγ-cPLA2 signaling is involved in the modulation of retinal neovascularization. Our studies have also demonstrated that activation of Pak1-cofilin signaling axis is required for retinal neovascularization. Furthermore, we have shown that besides VEGFA, VEGFC via CREB- mediated activation of DLL4-NOTCH1 signaling plays an important role in the enhancement of HRMVEC migration, proliferation, sprouting and tube formation in vitro in a cell culture model and retinal neovascularization in vivo in an OIR model. During the course of these studies, we found that both VEGFA and hypoxia stimulate PKCθ very robustly and depletion of its levels completely blocks the migrating and sprouting capacity of HRMVECs in response to VEGFA. We have also observed that both VEGFA and hypoxia induce Flt4 expression profoundly and this response requires PKCθ-mediated JunB induction. Furthermore, both VEGFA and hypoxia triggered the induction of expression of CDC6, a regulatory component of pre-replication complex (pre-RC) and involved in loading the mini chromosome maintenance proteins (MCM2-7) onto the DNA, which plays a rate-limiting step in replication. In addition, down regulation of CDC6 levels completely abolished VEGFA-induced DNA synthesis. Based on these novel discoveries, we propose that PKCθ via enhancing EC migration and sprouting and CDC6 via enhancing EC proliferation play an integral role in the modulation retinal neovascularization. To test this central hypothesis, we will address the following four specific aims. Aim 1. To test the hypothesis that PKCθ mediates hypoxia-induced retinal neovascularization. Aim 2. To test the hypothesis that JunB mediates hypoxia-induced retinal neovascularization. Aim 3. To test the hypothesis that PKCθ-JunB signaling axis targets Flt4 induction in mediating retinal neovascularization. Aim 4. To test the hypothesis that CDC6 plays an essential role in retinal neovascularization. The results of the proposed experiments may provide novel information in regard to the molecular mechanisms underling pathological retinal neovascularization, which could lead to the development of new therapeutic compounds against this ocular disease.