Ibuprofen-Caffeine Modulation of Retinal Endothelial Tip Cell Migration

早产儿视网膜病变（ROP）是所有极低胎龄新生儿（elgan, <28周/<1250g）中多达50%的患者。在elgan和动物模型中，布洛芬和咖啡因已被证明可以降低严重ROP的风险。使用我们实验室开发的独特技术，我们将检查对人类视网膜微血管内皮尖细胞（ECs）的特定影响，ECs是异常血管生成背后的驱动力。我们将研究它们与星形胶质细胞的动态相互作用，它们在氧化应激过程中的选择、激活和迁移。更重要的是，我们将研究布洛芬和/或咖啡因在阻止它们激活和降低它们感知血管生成信号的能力方面的功效。本研究的总体目标是研究EC尖端细胞在氧化应激（高氧/缺氧循环）环境下的行为及其与星形胶质细胞的关系；并确定布洛芬与咖啡因共同使用是否会保持细胞的静止。使用最先进的生物分析，蛋白质组学，药物基因组学，生物信息学和成像技术，我们将使用三个相互关联的特定目标：1)检查人类视网膜微血管内皮细胞和人类大脑星形胶质细胞在常氧和氧化应激（短暂高氧/缺氧循环）之间的关系。我们将重点关注VEGF和ECM蛋白水解，VEGF释放和梯度增加；尖端细胞的活化、VEGF的释放和再捕获以及迁移；VEGF和Notch信号机制；2)利用小干扰RNA确定VEGFR-2、VEGFR-3、NP-1、Notch 1、D1I4和Jagged 1在尖端细胞选择、激活和迁移中的作用

Retinopathy of prematurity (ROP) is afflicts as much as 50% of all extremely low gestational age neonates (ELGANs, <28 weeks/<1250g). Ibuprofen and caffeine have been shown to decrease the risk of severe ROP in ELGANs and in animal models. Using unique techniques developed in our laboratory, we will examine the specific effects on human retinal microvascular endothelial tip cells (ECs), the driving force behind aberrant angiogenesis. We will study their dynamic interaction with astrocytes, their selection, activation, and migration during oxidative stress. More importantly, we will examine the efficacy of ibuprofen and/or caffeine in preventing their activation and reducing their capacity to sense angiogenic cues. The overarching goal of this proposal is to study the behavior of EC tip cells and their relationship with astrocytes in the setting of oxidative stress (hyperoxia/hypoxia cycling); and to determine whether ibuprofen coadministered with caffeine will preserve fip cell quiescence. Using state-of-the-art bioanalytics, proteomics, pharmacogenomics, bioinformatics, and imaging techniques, we will use three interrelated specific aims: 1) to examine the relationship between human retinal microvascular ECs and human brain astrocytes in normoxia and in oxidative stress (brief hyperoxia/hypoxia cycling). We will focus on VEGF and ECM proteolysis, VEGF release and increased gradient; tip cell activation, release and recapture of VEGF, and migration; and VEGF and Notch signaling mechanisms; 2) to establish the roles of VEGFR-2, VEGFR-3, NP-1, Notch 1, D1I4, and Jagged 1 on tip cell selection, activation and migration using small interference RNA (SiRNA) knockdown of these specific genes in human retinal microvascular ECs. We will study the influence of astrocytes; and 3) to determine whether ibuprofen potentiated with caffeine will protect and preserve normal human retinal microvascular EC and astrocyte growth and function in oxidative stress. We will use state-of-the-art technologies to provide insights on the biomolecular mechanisms, pharmacokinetics, drug interactions; drug transport and metabolism.