The use of DNA for immunizations represents a new approach to raising immune responses. In this continuation, studies are proposed to (i) study the role of professional antigen presenting cells in the initiation of DNA-raised responses, (ii) to define parameters that determine whether a DNA-raised response will be biased towards Th1 or Th2 T-cell help, (iii) to study the role of germinal centers in DNA raised antibody responses, (iv) to characterize the cytolytic T-cells (CTL) raised by """"""""Th1"""""""" and """"""""Th2"""""""" DNA immunizations, and (v) to study the consequences of different types of DNA-raised T-help for challenge infections. Studies on antibody and antibody-mediated protection will use DNAs expressing normal (membrane-bound) and secreted forms of the influenza H1 hemagglutinin glycoprotein and the A/PR/8/34 (H1N1) murine influenza virus model. Studies on CTL and CTL-mediated protection will use DNA expressing the LCMV nucleoprotein (NP) and LCMV infections of mice. Antigen presenting cells will be examined in draining lymphoid tissue using DNA-expressed green fluorescent protein and in vitro restimulation of H1-specific Th cell lines. Co-transfected lymphokines, lymphokine knock out mice, and lymphokine neutralizing antibodies will be used to analyze the dependence on lymphokines of the Th-biases of DNA raised responses. DNA immunizations of germinal center knockout (CD40 knock out) and intact mice will be used to study the differentiation and function of Th cells and B cells that do, and do not enter the germinal center reaction. Studies on CTL will test for differences in frequency and function of CD8+ cells raised by Th1 and Th2-biased DNA immunizations. Post challenge studies will examine the effects of Th1- and Th2-biased responses on the control of infection and post challenge immunopathology.
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