Cell migration is a fundamental process involved in both normal and pathological conditions including wound healing and inflammation. Despite recent progress in understanding the mechanisms that regulate cell motility in vitro, the mechanisms that regulate cell movement and chemotaxis in vivo remain largely unknown. This research proposal is focused on establishing a model system to examine the mechanisms that regulate leukocyte migration and chemotaxis in vivo. We propose to use zebrafish to dissect mechanisms involved in leukocyte motility and to establish disease models. Both the tractable genetics and the transparency of zebrafish provide a powerful system to study vertebrate gene function and to dissect the mechanisms involved in leukocyte motility. Several recent experimental developments indicate a high likelihood of successful investigation of the proposed objectives: 1. In response to superficial wounding the chemotaxis of leukocytes to the wound site may be visualized in live zebrafish using DIC microscopy. 2. Using in situ hybridization the presence of both macrophage and neutrophil-like cells have been demonstrated in response to wounding. We now propose to use zebrafish to identify mechanisms involved in leukocyte migration and chemotaxis in vivo. Accordingly, we propose to: I. Characterize the migration and chemotaxis of leukocytes in zebrafish in response to wounding. Establish zebrafish lines that express GFP driven by the myeloperoxidase (mpo) promoter to isolate the neutrophilic response; II. Identify mutant zebrafish with aberrant leukocyte migration and chemotaxis using mutagenesis. The proposed studies will likely provide important insight into the mechanisms that regulate leukocyte recruitment and chemotaxis in living organisms. The studies will also have important implications for the identification of agents that modulate leukocyte motility and the development of an inflammatory response in vivo.
