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Plasmid DNA encoding Plasmodium yoelii circumsporozoite protein (PyCSP) followed by boosting with recombinant vaccinia virus containing the PyCSP elicited significant protective immunity in mice that was primarily mediated by CD8+ T-cell responses directed to P. yoelii -infected hepatocytes. This study was to further explore protection using in vitro cultures of P. yoelii parasites in mouse hepatocytes. Spleen cells from DNA/vaccinia virus-immunized mice were co-cultured in vitro with mouse hepatocytes containing developing P. yoelii liver stage parasites. A semipermeable membrane separating spleen cells and hepatocytes was used to demonstrate if cell-to-cell contact was required. Inhibitors of mediators likely involved in spleen cell killing were added to these co-cultures. Spleen cells from immunized mice inhibited in vitro P. yoelii parasite development, and inhibition was eliminated by separating effectors and targets with the semipermeable membrane. Additionally, inhibitors of inducible nitric oxide synthase, caspase activation, NF-κB activation as well as antibodies against interferon-gamma (IFN-γ) and ICAM-1 reduced parasite inhibition. These findings suggest that direct contact between spleen cells from immunized mice and P. yoelii-infected hepatocytes is required for eliminating liver stage parasites and provide more insight into CD8+ T-cell-mediated inhibition of malaria liver stage development.

Cellular immune response to DNA and vaccinia prime-boost immunization kills Plasmodium yoelii-infected hepatocytes in vitro