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. 2015 May 15;194(10):4657-67.
doi: 10.4049/jimmunol.1403080. Epub 2015 Apr 8.

Impaired NK Cell Responses to Pertussis and H1N1 Influenza Vaccine Antigens in Human Cytomegalovirus-Infected Individuals

Carolyn M Nielsen  1 , Matthew J White  1 , Christian Bottomley  2 , Chiara Lusa  1 , Ana Rodríguez-Galán  1 , Scarlett E G Turner  1 , Martin R Goodier  1 , Eleanor M Riley  3
Affiliations

Impaired NK Cell Responses to Pertussis and H1N1 Influenza Vaccine Antigens in Human Cytomegalovirus-Infected Individuals

Carolyn M Nielsen et al. J Immunol. .

Abstract

NK cells contribute to postvaccination immune responses after activation by IL-2 from Ag-specific memory T cells or by cross-linking of the low-affinity IgG receptor, CD16, by Ag-Ab immune complexes. Sensitivity of NK cells to these signals from the adaptive immune system is heterogeneous and influenced by their stage of differentiation. CD56(dim)CD57(+) NK cells are less responsive to IL-2 and produce less IFN-γ in response to T cell-mediated activation than do CD56(bright) or CD56(dim)CD57(-) NK cells. Conversely, NK cell cytotoxicity, as measured by degranulation, is maintained across the CD56(dim) subsets. Human CMV (HCMV), a highly prevalent herpes virus causing lifelong, usually latent, infections, drives the expansion of the CD56(dim)CD57(+)NKG2C(+) NK cell population, skewing the NK cell repertoire in favor of cytotoxic responses at the expense of cytokine-driven responses. We hypothesized, therefore, that HCMV seropositivity would be associated with altered NK cell responses to vaccine Ags. In a cross-sectional study of 152 U.K. adults, with HCMV seroprevalence rate of 36%, we find that HCMV seropositivity is associated with lower NK cell IFN-γ production and degranulation after in vitro restimulation with pertussis or H1N1 influenza vaccine Ags. Higher expression of CD57/NKG2C and lower expression of IL-18Rα on NK cells from HCMV seropositive subjects do not fully explain these impaired responses, which are likely the result of multiple receptor-ligand interactions. This study demonstrates for the first time, to our knowledge, that HCMV serostatus influences NK cell contributions to adaptive immunity and raises important questions regarding the impact of HCMV infection on vaccine efficacy.

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Figures

FIGURE 1.
FIGURE 1.
NK cell responses to pertussis and H1N1 are inhibited by IL-2 neutralization and IgG depletion. PBMCs were cultured in vitro for 18 h with medium alone, killed whole-cell pertussis (Per), and inactivated whole H1N1 influenza virus (H1N1), pertussis or H1N1 with blocking Ab to IL-2 (Per α-IL-2, H1N1 α-IL-2), or pertussis or H1N1 in IgG-depleted plasma (Per IgG depl., H1N1 IgG depl.). The isotype control Ab (IgG2A) for the IL-2 blocking Ab was included in the medium, pertussis, and H1N1 wells. Representative flow cytometry plots show gating of CD3−CD56+ NK cells and expression of CD25, IFN-γ, and CD107a (A). Responses to pertussis (B–D) and H1N1 (E–G) were measured by the percentage of NK cells expressing CD25 (B and E), coexpressing CD25/IFN-γ (C and F), and expressing CD107a (D and G). Data were analyzed in Prism using paired, one-tailed Wilcoxon signed-rank tests. Each data point represents one donor, n = 100 (B–D) or n = 16 (E–G), and bar graphs denote medians. ****p ≤ 0.0001, ***p < 0.001, **p < 0.01, *p < 0.05.
FIGURE 2.
FIGURE 2.
NK cell responses to vaccine Ag are affected by HCMV infection. PBMCs were cultured in vitro for 18 h with medium alone, LCC, killed whole-cell pertussis (Per), inactivated whole H1N1 influenza virus (H1N1), Per + LCC, H1N1 + LCC, or HCC. Donors were stratified into HCMV− (−) and HCMV+ (+) groups. Responses were measured as the percentage of NK cells expressing CD25 (A), coexpressing CD25/IFN-γ (B) or CD107a (C). Bivariate regression of age against responses to Per + LCC was performed for the percentage of NK cells expressing CD25 (D), CD25/IFN-γ (E), and CD107a (F). Each data point represents one donor, n = 152, except for H1N1 and H1N1 + LCC where n = 52. Bar graphs denote medians. NB, all Ag stimulations induced statistically significant increases in expression of CD25, CD25/IFN-γ, and CD107a over background (medium alone for pertussis/H1N1, or LCC for pertussis+LCC/H1N1+LCC; p < 0.05 in all cases), except that H1N1 did not induce a significant increase in CD25+IFN-γ+ NK cells in HCMV+ donors (p = 0.416). Data were analyzed in Prism using, one-tailed Mann–Whitney U tests. ****p ≤ 0.0001, ***p < 0.001, **p < 0.01, *p < 0.05.
FIGURE 3.
FIGURE 3.
Comparison of ex vivo expression of NK cell markers and receptors in HCMV− and HCMV+ donors. PBMCs were analyzed ex vivo for surface expression of CD56, CD57, CD16, NKG2C, and NKG2A, as shown by representative flow cytometry plots (A). Proportions of total NK cells in the CD56bright, CD56dimCD57−, CD56dimCD57int, and CD56dimCD57+ subsets were compared between HCMV− and HCMV+ donors (B), as was expression of CD16 (C), NKG2C (D), NKG2A (E), and CD57/NKG2C (F, CD56dim only). The percentages of cells expressing each marker in HCMV− (−) and HCMV+ (+) donors were compared using two-tailed Mann–Whitney U tests. Each data point represents one donor, n = 152; bar graphs denote medians. ****p ≤ 0.0001, ***p < 0.001, **p < 0.01, *p < 0.05.
FIGURE 4.
FIGURE 4.
HCMV infection affects vaccine Ag responses of all NK cells, irrespective of their differentiation status. PBMCs were cultured in vitro for 18 h with killed whole-cell pertussis with LCC (pertussis + LCC) (A–C and G–I) or inactivated whole H1N1 influenza virus with LCC (H1N1 + LCC) (D–F). Responses were measured as the percentage of cells expressing CD25 (A, D, and G), CD25/IFN-γ (B, E, and H), and CD107a (C, F, and I) by CD56/CD57-defined subsets (A–F) or CD56dim CD57/NKG2C-defined subsets (G–I) and compared between HCMV− (−) and HCMV+ donors (+). Data were analyzed using one-tailed Mann–Whitney U tests. Each data point represents one donor, n = 152 (A–C and G–I) or n = 52 (D–F); bar graphs denote medians. NB, for CD57/NKG2C-defined subsets, CD57int cells were grouped together with CD57− cells. ****p ≤ 0.0001, ***p < 0.001, **p < 0.01, *p < 0.05.
FIGURE 5.
FIGURE 5.
NK cell responses of HCMV+ donors with or without the characteristic CD56dimCD57+NKG2C+ expansion. PBMCs were cultured in vitro for 18 h with medium alone, LCC, killed whole-cell pertussis (Per), inactivated whole H1N1 influenza virus (H1N1), Per + LCC, H1N1 + LCC, or HCC. Donors were stratified into HCMV− (−), HCMV+ without expansion of CD56dimCD57+NKG2C+ cells (+), and HCMV+ with expansion of CD56dimCD57+NKG2C+ cells (++). Responses are expressed as the percentage of total NK cells expressing CD25 (A), coexpressing CD25/IFN-γ (B), or expressing CD107a (C). CD57-defined (D–F) or CD57/NKG2C-defined subsets (G–I) were analyzed for responses to pertussis with LCC for CD25 (D and G), CD25/IFN-γ (E and H), and CD107a (F and I). Data were analyzed in Prism using one-tailed Mann–Whitney U tests to compare responses between HCMV+ donors and either HCMV− donors or HCMV++ donors. ANOVA for linear trend (from − to + to ++) was also performed for each functional readout. Each data point represents one donor, n = 152, except for H1N1 and H1N1 + LCC where n = 52. Bar graphs denote medians. ****p ≤ 0.0001, ***p < 0.001, **p < 0.01, *p < 0.05.
FIGURE 6.
FIGURE 6.
Decreased cytokine responsiveness and decreased cytokine receptor expression by NK cells from HCMV+ donors. (A–C) PBMCs were cultured in vitro for 18 h with an HCC. Responses were measured as the percentage of CD56dim CD57/NKG2C-defined cells expressing CD25 (A), CD25/IFN-γ (B), and CD107a (C), and compared between HCMV− (−) and HCMV+ donors (+). (D–F) NK cells were analyzed for surface expression of IL-12Rβ2 using an mIgG1 PECy5-conjugated isotype control to set the gate (D). Total NK cells (E and F) and CD56/CD57-defined subsets (E) were analyzed ex vivo (E) and after 18 h culture in vitro with LCC or HCC (F). (G and H) NK cells were also analyzed for IL-18Rα surface expression using the T cell population to set the IL-18Rα gate (G), for total NK cells and CD56/CD57-defined subsets ex vivo (H). HCMV− and HCMV+ donors were compared using one-tailed (A–C) or two-tailed (E, F, and H) Mann–Whitney U tests. Each point represents one donor, n = 152 (A–C, E, and H) or n = 16 (F); bar graphs denote medians. ****p ≤ 0.0001, **p < 0.01, *p < 0.05.
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References

    1. He X. S., Draghi M., Mahmood K., Holmes T. H., Kemble G. W., Dekker C. L., Arvin A. M., Parham P., Greenberg H. B. 2004. T cell-dependent production of IFN-gamma by NK cells in response to influenza A virus. J. Clin. Invest. 114: 1812–1819. - PMC - PubMed
    1. Long B. R., Michaelsson J., Loo C. P., Ballan W. M., Vu B. A., Hecht F. M., Lanier L. L., Chapman J. M., Nixon D. F. 2008. Elevated frequency of gamma interferon-producing NK cells in healthy adults vaccinated against influenza virus. Clin. Vaccine Immunol. 15: 120–130. - PMC - PubMed
    1. Horowitz A., Behrens R. H., Okell L., Fooks A. R., Riley E. M. 2010. NK cells as effectors of acquired immune responses: effector CD4+ T cell-dependent activation of NK cells following vaccination. J. Immunol. 185: 2808–2818. - PubMed
    1. Evans J. H., Horowitz A., Mehrabi M., Wise E. L., Pease J. E., Riley E. M., Davis D. M. 2011. A distinct subset of human NK cells expressing HLA-DR expand in response to IL-2 and can aid immune responses to BCG. Eur. J. Immunol. 41: 1924–1933. - PMC - PubMed
    1. Horowitz A., Hafalla J. C., King E., Lusingu J., Dekker D., Leach A., Moris P., Cohen J., Vekemans J., Villafana T., et al. 2012. Antigen-specific IL-2 secretion correlates with NK cell responses after immunization of Tanzanian children with the RTS,S/AS01 malaria vaccine. J. Immunol. 188: 5054–5062. - PMC - PubMed

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