ATTENUATED LISTERIA-MONOCYTOGENES AS A LIVE VECTOR FOR INDUCTION OF CD8(-CELLS IN-VIVO - A STUDY WITH THE NUCLEOPROTEIN OF THE LYMPHOCYTIC CHORIOMENINGITIS VIRUS() T)

Listeria monocytogenes spends most of Its intracellular life cycle in the cytosol of the infected eucaryotic cells. Within this cellular com partment originates the endogenous pathway of antigen processing and p resentation. We thus assumed that recombinant L. monocytogenes express ing an heterologous protein, the nucleoprotein of the lymphocytic chor iomeningitis virus (LCMV), should be able to induce antigen-specific C D8(+) T cells in vivo, The LCMV nucleoprotein gene was-inserted in pha se with the sequence coding for the putative signal sequence of the he molysin of L. monocytogenes in order to target its secretion into the cytosol of the infected cell. The ability of this recombinant bacteriu m to induce LCMV-reactive CD8(+) T cells was then monitored in BALB/c mice. The immune status of the immunized BALB/c mice was studied on th e seventh day after a single i.v. injection of a sublethal dose of the recombinant bacteria: (i) cytotoxic CD8(+) T cells were detected in l iver; (ii) using in vitro re-stimulation with PMA and ionomycin, secon dary cytotoxic CD8(+) T cells were detected in spleen; (iii) an early inflammatory reaction dependent on the presence of CD8(+) T cells occu rred in the footpad after intraplantar inoculation of live LCMV; and ( iv) mice were protected against an otherwise lethal intracerebral LCMV challenge; the protection was accompanied by elimination of the virus , When the immune status of the immunized hosts was monitored for a lo nger period post-immunization, the balance between immune protection a nd immunopathology described for the anti-LCMV immune responses was ob served; two phases of protection were detected, flanking a transitory phase of exacerbation of the lymphocytic choriomeningitis disease (wee ks 2-5). Taken together, these results indicate the feasibility of usi ng attenuated L. monocytogenes as a model of a live vector to induce i n vivo CD8-dependent immune responses against intracellular pathogens.