Adults 65 years old and older have reduced numbers of functional memory T cells to respiratory syncytial virus fusion protein

Abstract

Respiratory syncytial virus (RSV) infects elderly (≥65 years) adults, causing medically attended illness and hospitalizations. While RSV neutralizing antibody levels correlate inversely with RSV-associated hospitalization in the elderly, the role of RSV-specific T cells in preventing disease in the elderly remains unclear. We examined RSV-specific humoral, mucosal, and cellular immune profiles in healthy elderly (65 to 85 years) and young (20 to 30 years) adults. RSV neutralization antibody titers in the elderly (10.5 ± 2.2 log(2)) and young (10.5 ± 2.1 log(2)) were similar. In contrast, levels of RSV F protein-specific gamma interferon (IFN-γ)-producing T cells were lower in elderly (180 ± 80 spot-forming cells [SFC]/10(6) peripheral blood mononuclear cells [PBMC]) than in young adults (1,250 ± 420 SFC/10(6) PBMC). Higher levels of interleukin-13 (IL-13; 3,000 ± 1,000 pg/ml) in cultured PBMC supernatants and lower frequency of RSV F-specific CD107a(+) CD8(+) T cells (3.0% ± 1.6% versus 5.0% ± 1.6%) were measured in PBMC from elderly than young adults. These results suggest that deficient RSV F-specific T cell responses contribute to susceptibility to severe RSV disease in elderly adults.

Fig 1

Equivalent RSV-specific antibody levels in plasma of young and elderly donors. (A) RSV-specific antibody titers in plasma from young (n = 30) and elderly (n = 30) donors were compared by microneutralization of GFP-RSV A2 virus. (B) F-specific IgG ELISA. (C) Correlation analysis between neutralizing antibody titers and F-specific IgG titers. (D) RSV F-specific and total IgA titers in nasal washes of young (n = 10) and elderly (n = 20) adults.

Fig 2

RSV F-specific memory B cells in young and elderly donors. RSV F-specific memory B cells were measured in young (n = 10) and elderly (n = 20) donor PBMC. PBMC cultures were stimulated ex vivo with a mixture of pokeweed mitogen, S. aureus protein A, and CpG DNA. The expanded RSV F-specific antibody-secreting cells (ASC) were measured in a memory B cell ELISPOT assay. Data are expressed as percent RSV F-specific IgG ASC/total IgG ASC.

Fig 3

Elderly have significantly lower numbers of RSV F-specific IFN-γ-producing cells. PBMC from young and elderly donors were stimulated ex vivo using (A) wt RSV A2 at 1 PFU/10 cells (n = 12 young and n = 20 elderly), (B) 5 μg/ml of RSV F protein (n = 20 young and n = 20 elderly), (C) 2 μg/ml of RSV F-specific CD4⁺ T cell peptide pools (n = 12 young and n = 12 elderly), or (D) 2 μg/ml of RSV F-specific CD8⁺ T cell peptide pools (n = 12 young and n = 12 elderly). The IFN-γ secreted by T cells was measured by ELISPOT assay, and data are expressed as spot-forming cells (SFC)/10⁶ PBMC. (E) Correlation between RSV F-specific CD4⁺ and CD8⁺ T cell frequencies.

Fig 4

Elderly PBMC have lower numbers of RSV F-specific CD107a⁺ CD8⁺ T cells. PBMC cultures were incubated in medium alone or were stimulated with RSV F protein or an RSV F-specific CD8⁺ T cell peptide pool. (A) CD107a gating strategy and acquisition data plots for medium, PMA/ionomycin, RSV F, or CD8⁺ T cell peptide pool stimulation of representative young (ME25) and elderly (ME26) donor PBMC. (B) The percentages of CD107a⁺ CD8⁺ T cells were quantified using multiparameter flow cytometry in young (n = 14) and elderly (n = 14) donor PBMC. (C) Fold CD107a⁺ CD8⁺ T cell numbers relative to medium following stimulation of young (n = 14) and elderly (n = 14) donor PBMC samples with RSV F protein or CD8⁺ T cell peptide pool.

Fig 5

Elderly PBMC secrete relatively low levels of IFN-γ in conjunction with secreting high basal levels of IL-13 ex vivo. PBMC cultures (n = 10 young and n = 10 elderly) were incubated in medium alone or with RSV F protein, and the resulting cytokines secreted into the supernatant were measured using a multiplex Th1-Th2 cytokine bead array panel (Luminex). (A) IFN-γ. (Β) IL-13.