G protein-coupled receptor regulation of transcriptional mechanisms in the retinal vasculature.

项目摘要 视网膜血管功能障碍导致视力损害和视力丧失，这种现象发生在 早产儿视网膜病变（ROP）、糖尿病视网膜病变（DR）和新生血管性年龄相关性黄斑变性 （AMD）。视网膜内皮细胞（REC）的损伤引发了一系列的病理事件，最终导致 加速视网膜疾病的发展。REC损伤如何导致转录变化，决定 视网膜血管功能是否恢复或导致病理变化尚不清楚。氨醇 1-磷酸盐（S1P），一种通过G蛋白偶联的S1P受体（S1PR1 - 5）发出信号的血液脂质介质。 申请人的实验室发现了第一个S1PR，并确定了其在血管屏障中的功能作用 维持、发育/成熟、抗炎过程、细胞存活和内皮/周细胞 交互.虽然FDA批准的两种靶向S1PR的药物在治疗多种 硬化、视网膜起泡和黄斑水肿是剂量限制性不良反应， 视网膜屏障我们最近发现S1PR信号抑制血管内皮生长因子， （VEGF）诱导的AP-1转录因子活性并允许Norrin/Wnt/β-连环蛋白依赖性REC基因 表达，从而导致视网膜REC特化。在AP-1因子中，JunB蛋白表达是 最显着地受到S1 PR信号传导的调节，S1 PR信号传导是最佳血管网络扩张所需的事件， 视网膜深血管丛的形成。该提案的中心假设是REC S1PR信号传导 建立JunB转录因子梯度并允许REC器官型特化机制。 以这种方式，减弱的S1PR信号传导轴驱动功能不良的视网膜血管网络， 血管增生性ROP。在本提案中，第一个具体目标将阐明 S1PR在REC中对JunB转录因子梯度的塑造。第二，S1PR信号在REC中是如何传递的？ 通过有效的Norrin/Wnt/β-连环蛋白依赖性信号转导促进器官型特化 并进行基因表达。具体重点将是omega-3脂肪酸转运蛋白（MFSD 2A）和 铁转运蛋白（TFRC）。这些机制在ROP小鼠模型中的相关性将得到解决 具体目标3.这些研究有望增强我们对视网膜病变基本机制的理解， 视网膜血管发育、特化和疾病，并最终导致驯服 通过靶向S1P脂质信号传导轴治疗视网膜疾病，并提供副作用较少的S1PR抑制剂。

PROJECT SUMMARY Retinal vascular dysfunction leads to visual impairment and loss of vision, a phenomenon that occurs in retinopathy of prematurity (ROP), diabetic retinopathy (DR) and neovascular age-related macular degeneration (AMD). Injury of the retinal endothelial cell (REC) initiates a series of pathogenetic events that ultimately lead to accelerated progression of retinal diseases. How REC injury leads to transcriptional changes that determine whether the retinal vascular function is restored or leads to pathological changes is not known. Sphingosine 1-phosphate (S1P), a blood-borne lipid mediator that signals via G protein-coupled S1P receptors (S1PR1-5). The applicant’s laboratory discovered the first S1PR and worked out its functional roles in vascular barrier maintenance, development/ maturation, anti-inflammatory processes, cell survival and endothelial/ pericyte interactions. Although two FDA-approved S1PR-targeted drugs are efficacious in the treatment of multiple sclerosis, retinal blistering and macular edema are dose-limiting adverse effects due to the impairment of retinal barriers. We recently showed that S1PR signaling suppresses vascular endothelial growth factor (VEGF)-induced AP-1 transcription factor activity and permits Norrin/Wnt/ß-catenin-dependent REC gene expression, thus leading to retinal REC specialization. Among the AP-1 factors, JunB protein expression is most prominently regulated by S1PR signaling, an event needed for optimal vascular network expansion and formation of deep retinal vascular plexus. The central hypothesis of the proposal is that REC S1PR signaling establishes JunB transcription factor gradients and permits the REC organotypic specialization mechanisms. In this manner, attenuated S1PR signaling axis drives poorly functional retinal vascular network and vasoproliferative ROP. In this proposal, the first specific aim will elucidate mechanisms and consequences of S1PR sculpting of JunB transcription factor gradients in REC. Second, how S1PR signaling in the REC promotes organotypic specialization by enabling efficient Norrin/Wnt/ß-catenin-dependent signal transduction and gene expression will be conducted. Specific focus will be on omega-3 fatty acid transporter (MFSD2A) and iron transporter (TFRC). The relevance of these mechanisms in the mouse models of ROP will be addressed in specific aim 3. These studies are anticipated to enhance our understanding of basic mechanisms of retinal vascular development, specialization and disease in the retina and ultimately lead to approaches that tame retinal disorders by targeting the S1P lipid signaling axis and to provide S1PR inhibitors with fewer side effects.