CD4+ T cell-independent vaccination against Pneumocystis carinii in mice

Abstract

Host defenses are profoundly compromised in HIV-infected hosts due to progressive depletion of CD4+ T lymphocytes. Moreover, deficient CD4+ T lymphocytes impair vaccination approaches to prevent opportunistic infection. Therefore, we investigated a CD4+ T cell-independent vaccine approach to a prototypic AIDS-defining infection, Pneumocystis carinii (PC) pneumonia. Here, we demonstrate that bone marrow-derived dendritic cells (DCs) expressing the murine CD40 ligand, when pulsed ex vivo by PC antigen, elicited significant titers of anti-PC IgG in CD4-deficient mice. Vaccinated animals demonstrated significant protection from PC infection, and this protection was the result of an effective humoral response, since adoptive transfer of CD4-depleted splenocytes or serum conferred this protection to CD4-deficient mice. Western blot analysis of PC antigen revealed that DC-vaccinated, CD4-deficient mice predominantly reacted to a 55-kDa PC antigen. These studies show promise for advances in CD4-independent vaccination against HIV-related pathogens.

Figure 1

(a) Outline of double vaccine protocol. *Vaccine groups: PBS only, DCs only, DC/AdCD40L, DC/AdLuc, DC/AdCD40L/PC, DC/AdLuc/PC. (b) Induction of anti-PC IgG after two rounds of DC-based PC vaccine. Mice were vaccinated as outlined in b, and serum was obtained at 4 weeks to measure induction of anti-PC IgG. CD4-intact mice were used as a positive control and received PC antigen in CFA followed by IFA 2 weeks later. This serum was harvested at 4 weeks for anti-PC IgG determination (n = 5–6 per group; **P < 0.05). BM, bone marrow; i.t., intratracheal; i.v., intravenous.

Figure 2

Protection of DC-vaccinated mice to in vivo challenge with PC. Mice were vaccinated as outlined in Figure 1b and then challenged with 2 × 10⁵ PC cysts intratracheally and sacrificed at 2 weeks (a) or 4 weeks (b) after PC challenge (n = 5–6 per group). *P < 0.05, **P < 0.001 versus controls.

Figure 3

(a) Adoptive transfer of serum or splenocytes from DC/AdCD40L/PC-vaccinated mice confers protection to CD4-depleted mice from in vivo challenge with PC. Mice were passively immunized with serum or splenocytes from vaccinated or control CD4-depleted mice. Twenty-four hours later, mice were challenged with PC; they were sacrificed 4 weeks later to assess intensity of PC infection (n = 5–6 per group; *P < 0.01 versus controls). (b) Immunoreactivity of serum from vaccinated mice. The two lanes of control mice represent one each of DCs alone or AdLuc-modified DCs pulsed with PC. Serum from CFA/IFA-immunized mice resulted in a polyclonal response, whereas serum from two representative AdCD40L/DC/PC mice reacted with a 55-kDa PC antigen.

Figure 4

Representative GMS stains from vaccinated or control mice. (a) Lung section from a mouse that received splenocytes from CD4-depleted mice vaccinated with AdLuc-modified DCs pulsed with PC. (b) Lung section from a mouse that received serum from mice vaccinated with AdCD40L-modified DCs pulsed with PC. (c) Lung section of a mouse that received serum from mice vaccinated with AdLuc-modified DCs pulsed with PC. (d) Lung section of a mouse that received splenocytes from mice vaccinated with AdCD40L-modified DCs pulsed with PC.