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.
These specific aims will use a unique and novel model for oxidative stress (brief, frequent hyperoxia/hypoxia cycling) to study the effects on retinal microvascular endothelial fip cells selection, activation, and migration, and their relationship to astrocytes. We will also use a unique pharmacologic approach to prevent or curtail the biological cues responsible for aberrant tip cell migration and activation. ROP is the leading cause of childhood blindness and the epidemic is increasing. The current, popular use of intravitreal Avastin is highly invasive. In addition to the added pain and distress to the micropremie, it causes retinal hemorrhage, retinal detachment, and choroidal ruptures. More importantly, it may have adverse effects on associated retinal cells such as astrocyges and microglia and influence normal brain development. The need for other potential therapies is vital. Our proposed studies will provide further understanding of the mechanisms associated with aberrant fip cell migration and the role of genes involved in ECM and VEGF degradation, and VEGF and Notch signaling in oxidative stress. These projects will provide additional information and add to our current repertoire gained from previous experiments. We have all the necessary technologies and systems in order to successful accomplish these proposed proposals. They will provide the basis for clinical trials utilizing an alternate, safe, effective, and non-invasive pharmacologic approach to treatment of ROP.
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