Anti-interleukin 10 receptor monoclonal antibody is an adjuvant for T helper cell type 1 responses to soluble antigen only in the presence of lipopolysaccharide

Abstract

Soluble foreign antigen usually leads to a transient clonal expansion of antigen-specific T cells followed by the deletion and/or functional inactivation of the cells. As interleukin (IL)-10 is a key immunoregulatory cytokine, we questioned whether neutralization of IL-10 during priming with soluble antigen could prime for a subsequent T helper cell type 1 (Th1) effector recall response. By using an adoptive transfer model to track the fate of antigen-specific T cell receptor (TCR)-transgenic CD4(+) T cells, we show that administration of soluble ovalbumin (OVA) protein, but not OVA(323-339) peptide antigen, together with an anti-IL-10 receptor (R) mAb led to the enhancement of a Th1 response upon rechallenge. Lipopolysaccharide (LPS) present in the protein was necessary for priming for Th1 recall responses in the presence of anti-IL-10R mAb, as removal of LPS abrogated this effect. Moreover, addition of LPS to the peptide did not itself allow priming for recall Th1 effector responses unless endogenous levels of IL-10 were neutralized with an anti-IL-10R mAb. A significant increase in OVA-specific IgG1 and IgG2a isotypes was observed when the protein antigen was administered with anti-IL-10R mAb; however, this was not the case with peptide antigen administered together with anti-IL-10R and LPS. Our data, showing that LPS receptor signaling and neutralization of endogenous immunosuppressive cytokines is essential for Th1 priming, has important implications for the design of relevant vaccines for effective in vivo immunotherapy.

Figure 1

Treatment of OVA-primed mice with anti–IL-10R mAb before rechallenge potentiates the production of IFN-γ. CD4⁺ T cells were purified from the draining lymph nodes of mice primed and rechallenged as indicated in Materials and Methods and stimulated in vitro with OVA_323–339 and APCs. Supernatants were harvested after 24 (IL-2) or 48 h (IFN-γ) of culture.

Figure 2

Neutralization of IL-10 allows priming for Th1 recall responses by soluble protein or peptide antigen only when low levels of LPS are present. CD4⁺ T cells were purified from the draining lymph nodes of mice primed with OVA protein, OVA protein that had been purified as free of LPS (*OVA), or OVA_323–339 peptide antigen in the presence or absence of LPS (140 EU [14 ng]) plus or minus anti–IL-10R mAbs as shown. Mice were then rechallenged with OVA and CFA as indicated in Materials and Methods and stimulated in vitro with OVA_323–339 and APCs. Supernatants were harvested after 24 (IL-2) or 48 h (IFN-γ) of culture. Results were reproducible in greater than three experiments.

Figure 3

Anti–IL-10R mAb plus LPS enhances antigen-specific IgG1 and IgG2a levels with protein and not peptide antigen. Mice were primed and rechallenged as indicated in Materials and Methods. Serum was collected 7 d after rechallenge with OVA in CFA. The average concentration in nanograms per milliliter for each isotype is indicated in parentheses. Each symbol represents data from an individual mouse. Results were reproducible in greater than three experiments.