We hypothesize that memory T lymphocytes present after primary dengue virus infection are important for protection against subsequent infection with virus of the same dengue serotype but also are involved in the immunopathological processes which accompany the severe complications of secondary infection with a virus of another dengue serotype. The purpose of this project is to examine the memory helper T lymphocyte (Th) and cytotoxic T lymphocyte (CM) responses of murine spleen cells after primary dengue virus infection as a model of the human T cell responses to dengue virus. The goal of the project is to define the structure of the major T cell epitopes of dengue viruses and the function of the dengue virus-specific Th and CTL responses in protection against disease and in the immunopathology associated with dengue virus infections. This information will be used to identify important dengue viral proteins that will be included in candidate subunit vaccines and to speed the development of immunogenic vaccine preparations. Phase I of this project would allow the candidate to develop the techniques for demonstrating specific Th and CTL activity and long term culturing of virus-specific T lymphocytes. Didactic instruction will be included to bring up-to-date the candidate's knowledge in the fields of virology and immunology. In phase II of the project, dengue virus-immune spleen cells and T cell clones would be used to test the ability of candidate subunit vaccines- recombinant antigenic constructs and purified dengue viral proteins- to induce dengue virus-specific T cell responses. Computer algorithms will be used to select the likely T cell epitopes from the published dengue virus protein sequences. The peptides identified will be synthesized and tested for the ability to stimulate Th and CTL activity. Those preparations which show the best in vitro activity will then be tested for the ability to induce dengue virus-specific Th and CTL activity in vivo. Mice will then be challenged with dengue virus, and we will attempt to correlate the results of virus challenge with the results of dengue virus-specific T cell assays performed before challenge.
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