A novel immunization method to induce cytotoxic T-lymphocyte responses (CTL) against plasmid-encoded herpes simplex virus type-1 glycoprotein D.

DNA molecules complexed with an asialoglycoprotein-polycation conjugate, consisting of asialoorosomucoid (ASOR) coupled to poly-L-lysine, can enter hepatocytes which bear receptors for ASOR. We used this receptor-mediated DNA delivery system to deliver plasmid DNA encoding glycoprotein D (gD) of herpes simplex virus type 1 to ASOR-positive cells. Maximum expression of gD protein was seen at 3 days after injection of this preparation in approximately 13% of cells from BALB/c mice [hepatocytes from mice injected intravenously (i.v.) or peritoneal exudate cells from mice injected intraperitoneally (i.p.)]. In comparison with mice injected with either the plasmid vector alone or the gD-containing plasmid uncomplexed to ASOR, mice immunized with gD-containing plasmid complexed with ASOR-poly-L-lysine induced marked antigen-specific CTL responses. BALB/c mice immunized with gD-DNA developed a T-cell-mediated CTL response against target cells expressing gD and MHC class II glycoproteins, but not against cells expressing only gD and MHC class I molecules. In C3H mice, gD-DNA induced a T-cell-mediated CTL response against target cells expressing gD and class I MHC molecules. Serum anti-gD antibody in low titers were produced in both strains of mice. DNA complexed with ASOR-poly-L-lysine induced CTL responses in mice.

Staphylococcal enterotoxin B primes cytokine secretion and lytic activity in response to native bacterial antigens.

Superantigens stimulate T-lymphocyte proliferation and cytokine production, but the effects of superantigen exposure on cell function within a complex, highly regulated immune response remain to be determined. In this study, we demonstrate that superantigen exposure significantly alters the murine host response to bacterial antigens in an in vitro coculture system. Two days after exposure to the superantigen staphylococcal enterotoxin B, splenocytes cultured with Streptococcus mutans produced significantly greater amounts of gamma interferon (IFN-gamma) and interleukin-12 than did sham-injected controls. The majority of IFN-gamma production appeared to be CD8(+) T-cell derived since depletion of this cell type dramatically reduced the levels of IFN-gamma. To study host cell damage that may occur following superantigen exposure, we analyzed cytotoxicity to "bystander" fibroblast cells cultured with splenocytes in the presence of bacterial antigens. Prior host exposure to staphylococcal enterotoxin B significantly enhanced fibroblast cytotoxicity in the presence of bacteria. Neutralization of IFN-gamma decreased the amount of cytotoxicity observed. However, a greater reduction was evident when splenocyte-bacterium cocultures were separated from the bystander cell monolayer via a permeable membrane support. Increased cytotoxicity appears to be primarily dependent upon cell-cell contact. Collectively, these data indicate that overproduction of inflammatory cytokines may alter the activity of cytotoxic immune cells. Superantigen exposure exacerbates cytokine production and lytic cell activity when immune cells encounter bacteria in vitro and comparable activities could possibly occur in vivo.

Cytokines produced early in picornavirus infection reflect resistance or susceptibility to disease.

Gender bias favoring female resistance to picornavirus disease is not seen in ICR Swiss mice following infection with the MM strain of encephalomyocarditis virus (EMCV) (causing encephalitis and death) as it is with D variant of EMCV (causing diabetes in males). To define this difference, an in vitro virus-infected splenocyte culture system was used to explore virus effects on lymphoid cells. Infected and sham-infected splenocyte cultures, prepared from both genders of mice and infected with either virus variant, were examined for immunoregulatory cytokines in the first 24 h of infection using ELISA or bioassays. Disease resistance was associated with increased levels of interferon-y (IFN-gamma) and undetectable levels of interleukin-10 (IL-10) by 12 h postinfection in splenocytes from ICR Swiss females infected with EMCV-D. Disease susceptibility was associated with high levels of IL-10 at 12 h after infection of spleen cells from ICR Swiss males infected with EMCV-D or from both genders infected with EMCV-MM. This information was used to protect susceptible mice against picornavirus disease (either diabetes or death) by giving them an inducer of IFN-alpha/beta, to induce natural killer (NK)-like cells to produce high levels of IFN-gamma and rat monoclonal anti-IL-10 to neutralize the effects of mouse IL-10.