Identification and Characterization of Dendritic Cells in Fish

Preliminary studies from our laboratory provide strong evidence that rainbow trout have specialized cells that bear many of the hallmarks of mammalian dendritic cells (DCs) including a superior ability to initiate lymphocyte (specifically, T-cell) proliferation in vitro. To validate the precise role of these cells in antigen-presentation and to further characterize their behavior and activity we plan to examine how these cells respond to well characterized signals for the activation of mammalian DCs and whether they are capable of inducing lymphocyte responses in vivo with the ultimate goal of determining where in fish antigen presentation takes place. In so doing, we will collect valuable information on the transcriptional responses of these cells to pathogen-associated molecular patterns, or to infection with bacterial pathogens. Lastly, by loading trout DCs ex vivo with bacterial antigens and returning these to syngeneic and allogeneic fish we hope to demonstrate either correlates
of protection (in the form of T-cell responses) or frank protection following direct pathogen challenge. In line with these goals, the specific aims of this project are to: 1) Determine the response of trout DCs to activating signals from toll-like receptor (TLR) ligands and infectious disease agents; 2) Examine the trafficking of trout DCs in fish tissues, and determine where antigen-presentation is likely to take place; and, 3) Establish that antigen-loaded DCs can induce correlates of protection and/or frank protection against known bacterial pathogens of fish. In aim 1, we will investigate the phagocytic activity of trout DCs as well as their gene expression profiles in response to toll-like receptor ligands and bacterial infection. In aim 2, we will determine the homing patterns of cultured trout DCs in recipient fish and the ability of these cells to stimulate T-cell proliferation in vivo following treatment with bacterial superantigens. In aim 3, we will determine whether trout
DCs can stimulate antigen-specific responses to bacterial antigens by measuring T-cell recall responses to bacterial antigens in vitro, and levels of protection of live fish challenged with bacterial pathogens after vaccination with antigen-loaded DCs.