Combined NKT cell activation and influenza virus vaccination boosts memory CTL generation and protective immunity - PubMed (original) (raw)

Combined NKT cell activation and influenza virus vaccination boosts memory CTL generation and protective immunity

Carole Guillonneau et al. Proc Natl Acad Sci U S A. 2009.

Abstract

Current influenza A virus vaccines do not generate significant immunity against serologically distinct influenza A virus subtypes and would thus be ineffective in the face of a pandemic caused by a novel variant emerging from, say, a wildlife reservoir. One possible solution would be to modify these vaccines so that they prime cross-reactive CD8(+) cytotoxic T lymphocytes (CTL) cell-mediated immunity directed at conserved viral epitopes. A further strategy is to use novel adjuvants, such as the immunomodulatory glycolipid alpha-galactosylceramide (alpha-GalCer). We show here that giving alpha-GalCer with an inactivated influenza A virus has the paradoxical effect of diminishing acute CTL immunity via natural killer T (NKT) cell-dependent expression of indoleamine 2,3-dioxygenase (IDO), an important mediator of immune suppression, while at the same time promoting the survival of long-lived memory CTL populations capable of boosting protection against heterologous influenza A virus challenge. This enhancement of memory was likely due to the alpha-GalCer-induced upregulation of prosurvival genes, such as bcl-2, and points to the potential of alpha-GalCer as an adjuvant for promoting optimal, vaccine-induced CD8(+) T cell memory.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.

Fig. 1.

Quantitative analysis of DbNP366+ and DbPA224+ CD8+ T cells following influenza vaccination with an adjuvant. The B6 mice were immunized with iIAV ± α-GalCer, and lymphocytes from the spleen and brachial lymph node (BLN) were sampled 7 days later. The percentage (A and B) and absolute number (C and D) of tetramer-positive populations was evaluated among the CD8+ T cells in the spleen (A and C) and BLN (B and D). Results are expressed as mean ± SD for groups of 5 mice. *P < 0.05; **P < 0.01.

Fig. 2.

Fig. 2.

α-GalCer promotes the generation of long-term CD8+ T cell memory. The B6 mice were immunized with iIAV ± α-GalCer and killed 6 weeks later. Splenocytes were harvested and analyzed for the percentage (A) and absolute number (B) of tetramer-positive CD8+ CTLs. (C) The proportion of TCM vs. TEM cells was evaluated at early and late time points after immunization. Results are expressed as mean ± SD for groups of 5 mice. *P ≤ 0.05; **P < 0.01

Fig. 3.

Fig. 3.

NKT cells limit the acute CTL response but are essential for memory. Groups of CD1d+/+ and CD1d−/− mice were immunized with iIAV ± α-GalCer and analyzed 7 days (A and C) or 6 weeks later (B and D) for the percentage (A and B) and absolute number (C and D) of DbNP366+CD8+ T cells in the spleen. Results are expressed as mean ± SD. *P ≤ 0.05; **P < 0.01.

Fig. 4.

Fig. 4.

Increased protection of mice vaccinated with α-GalCer after infection. Mice were challenged i.n. with 104 pfu of live HKx31 IAV at 6 weeks after priming with iIAV + (black bars) or − (white bars) α-GalCer. The percentage (A) and absolute numbers (B) of influenza-specific CTLs were evaluated for the bronchoalveolar lavage (BAL), spleen, and mediastinal lymph node (MLN). Cells were stained for IFNγ and TNFα after stimulation with NP366 peptide. Shown is the percentage of IFNγ+ CD8+ T cells (C) and the proportion of IFNγ+ T cells that are also TNFα+ (D). *P ≤ 0.05 and **P < 0.01 comparing iIAV ± α-GalCer. (E) Virus clearance from the lung of mice that had been primed with iIAV ± α-GalCer. The graphs show the mean pfu per lung (n = 5) ± SD. **P < 0.01.

Fig. 5.

Fig. 5.

Increased expression of IFNγ and IDO inhibit T cell proliferation. (A) Quantitative analysis of IFNγ and IDO transcript accumulation was performed for the LNs of CD1d+/+ and CD1d−/− mice given iIAV ± α-GalCer. Results are expressed in arbitrary units (log2) of molecules normalized to hypoxanthine phosphoribosyltransferase ± SD for groups of 9 to 3 mice. (B) Mice were vaccinated with iIAV ± α-GalCer, and some were given 1-MT for 7 days. The proportion of CTLs was evaluated in the spleen (Left) and the LN (Right). Results are expressed as mean ± SD for groups of 10 mice. (C) Mice were analyzed 6 weeks after treatment for the percentage and number of CTLs in the spleen. Results are expressed in mean ± SD for groups of 5 mice. **P ≤ 0.001.

Fig. 6.

Fig. 6.

Enhanced memory-cell generation correlates with increased survival signals. (A) Representative histograms of intracellular Bcl-2 expression by splenic CD8+CD44+ T cells from mice given iIAV + (Right) or − (Left) a-GalCer. (B) Quantitation of the proportion of CD8+CD44+ CTLs expressing intracellular Bcl2 at 7 days after iIAV ± α-GalCer. Results are expressed as mean ± SD for groups of 5 mice. *P ≤ 0.05; ***P < 0.001.

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