Splenocytes from mice primed with influenza virus envelope proteins incorporated in iscoms, as micelles or as infectious virus, were restimulated in vitro with the same antigen. Interleukin-2 (IL-2) and interferon-gamma (IFN-gamma) were assayed in the supernatants of such cultures. Influenza virus iscoms induced IL-2 and IFN-gamma responses in restimulation experiments that were antigen specific and significantly higher than those induced by micelles or infectious virus. Serum samples collected at the end of the experiments were analysed for the antibody response and profile. The antibody titres induced by iscoms were of a similar order of magnitude as those induced by infectious virus, and were about 18 times higher than the titres induced by micelles. In mice immunized with iscoms or infectious virus the most abundant antibodies were of the IgG1 and IgG2a isotype, and the IgE response was low. We conclude that immunization with iscoms stimulates the Th1-like subtype of murine T lymphocytes.

To understand how a presentation system for antigens initiates an immune response and why it has a strong adjuvant activity, a number of parameters need to be analysed. In this study the frequency of spleen cells expressing MHC class II (Ia antigen) was determined after immunization of mice and restimulation of their spleen cells, in vitro, with influenza virus envelope proteins in different physical forms, namely iscoms, micelles and virus particles. All three forms of the antigen stimulated, in an antigen-specific manner, an increased proportion of spleen cells expressing MHC class II in the restimulation experiments. The induction of increased MHC class II expression was at least partly dependent on antigen-specific induction of IFN-gamma since an antibody to IFN-gamma partly inhibited the increase of MHC class II+ cells induced by iscom or by Concanavalin A. The iscom-borne antigens were superior to micelles to prime the immune response in vitro, indicating a capacity to induce memory cells. This primed immune response was readily recalled in vitro, as measured by IFN-gamma production and an increased number of MHC class II positive cells.

SUMMARYThe kinetics ofthe expression of membrane-associated IL-1 (mIL-1) and soluble IL-1 (sIL-1) was studied in in vitro stimulated spleen cells from non-primed mice or from mice primed with influenza virus antigens incorporated in the immuno-stimulating complexes (iscoms) or as micelles. Matrix, which is the carrier structure for the antigens in the iscom, was used as a non-antigen stimulus. The IL-1 produced was assayed in an I L-l-dependent cell line and the specificity was demonstrated in a blocking experiment with antiserum to IL-la. Soluble IL-la was also quantified in ELISA. Iscoms and matrix induced production of mIL-1 and sIL-l in cultures from non-treated mice as well as from mice primed 4 days before with iscoms or micelles. Micelles were a less strong stimulus and did not induce production of sIL-1. Micelles induced production of mIL-1 in cultures from non-primed mice or from mice which were recently immunized with micelles. No mIL-l expression was induced by micelles if the spleen cells originated from mice immunized shortly before with iscoms. Depletion experiments demonstrated that sIL-1 was produced by adherent cells upon stimulation with iscoms or matrix. However, factor(s) from the non-adherent cells seem to be necessary for optimal secretion ofslL-1.