The hyaloid vasculature functions to supply the highly metabolic eye with oxygen and nutrients during the early phases of eye development. While hyaloid formation has been described to various extents in humans, mice, and chick, all of these studies utilized fixed samples and therefore have been unable to generate a comprehensive understanding of the dynamic cellular and molecular underpinnings of hyaloid morphogenesis. Experiments in this proposal utilize the zebrafish as a model for studying hyaloid morphogenesis and preliminary data demonstrate that hyaloid formation is almost identical to that in humans. Given the strengths of the zebrafish system for in vivo imaging and genetic manipulations, it is possible to image the migration of hyaloid precursor cells into the eye and their subsequent morphogenetic movements to ultimately give rise to the functional hyaloid. Moreover, given the genetic amenability of zebrafish and the large quantity of loss of function mutations in the system, the effects of these mutations on hyaloid formation can be determined. Vascular development is thought to require interactions between vascular precursor cells and their extracellular matrix (ECM), both during migratory and morphogenetic phases. The role of ECM in these processes, however, has been difficult to study because mutations or knock-out of many ECM components (i.e. fibronectin and laminin) are embryonic lethal and embryos do not survive until hyaloid development. Zebrafish mutants and loss of function embryos for these ECM components, their cellular binding partners (integrin-alpha-5) and their intracellular signaling mediators (focal adhesion kinase) survive to hyaloid development, and preliminary analysis of each identifies defects in hyaloid formation. These data support the hypothesis that hyaloid formation requires a precise coordination of cell-ECM interactions, and that specific cell-ECM components mediate distinct events in hyaloid formation. Experiments in this proposal will test this hypothesis.
In Specific Aim 1, the morphogenetic and cell biological processes underlying hyaloid vasculature formation will be identified.
In Specific Aim 2, the role of Fibronectin/integrin-alpha-5 and FAK during hyaloid formation will be determined.
In Specific Aim 3, the role of laminin during hyaloid formation will be determined. When completed, data generated from these experiments will be significant as they will have identified the in vivo regulation of hyaloid formation and how cell-ECM interactions mediate formation of the hyaloid. Because of the conservation of gene function between zebrafish and humans, these data will directly translate into a better understanding of hyaloid formation in the human eye and how defects in cell-ECM interactions can lead to defects in hyaloid formation.
Experiments in this proposal will identify the cell biological and morphogenetic underpinnings of hyaloid vasculature formation. Hyaloid formation is critical to supply the eye with oxygen and nutrients during early eye development yet little is known about how the hyaloid is built. Thus, the results of these experiments will be significant because they will have identified the morphogenetic underpinnings of hyaloid formation and determined how cell-extracellular matrix interactions facilitate the process.