Frequency of epitope-specific naive CD4(+) T cells correlates with immunodominance in the human memory repertoire - PubMed (original) (raw)

Frequency of epitope-specific naive CD4(+) T cells correlates with immunodominance in the human memory repertoire

William W Kwok et al. J Immunol. 2012.

Abstract

The frequency of epitope-specific naive CD4(+) T cells in humans has not been extensively examined. In this study, a systematic approach was used to examine the frequency of CD4(+) T cells that recognize the protective Ag of Bacillus anthracis in both anthrax vaccine-adsorbed vaccinees and nonvaccinees with HLA-DRB1*01:01 haplotypes. Three epitopes were identified that had distinct degrees of immunodominance in subjects that had received the vaccine. Average naive precursor frequencies of T cells specific for these different epitopes in the human repertoire ranged from 0.2 to 10 per million naive CD4(+) T cells, which is comparable to precursor frequencies observed in the murine repertoire. Frequencies of protective Ag-specific T cells were two orders of magnitude higher in immunized subjects than in nonvaccinees. The frequencies of epitope-specific memory CD4(+) T cells in vaccinees were directly correlated with the frequencies of precursors in the naive repertoire. At the level of TCR usage, at least one preferred Vβ in the naive repertoire was present in the memory repertoire. These findings implicate naive frequencies as a crucial factor in shaping the epitope specificity of memory CD4(+) T cell responses.

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Figures

Figure 1

Figure 1

Frequencies and phenotypes of DR0101 restricted PA reactive T cells in DR0101 AVA vaccinees. A. PBMC from a DR0101 subject were stained with DR0101/PA401-420, DR0101/PA505-524 and DR0101/PA713-732 PE-tetramers and anti-CD45RA antibody, and with gating antibodies ex vivo. The cells were subsequently incubated with anti-PE magnetic beads and enriched with a magnetic column. Frequencies of PA specific T cells were determined by dividing the number of tetramer positive cells in the enriched fraction by the total number of CD4+ T cells in the sample before fractionation, determined to be 63, 22 and 1,000 per million total CD4+ T cells for PA401-420, PA505-524 and PA713-732 epitopes, respectively. B. Frequencies of PA401-420, PA505-524 and PA713-732 T cells in four different DR0101 AVA vaccinees. Each symbol represents the epitope specific T cell frequency for a single subject. The p values were determined by using two-tailed unpaired t tests. C. CD27 and CCR4 expression of PA713-732 reactive CD4+ T cells. For each panel (A-C), we gated on CD4+, CD14-, CD19- and Via-Probe negative cells. The percentages of tetramer positive cells that expressed the surface marker are as indicated. Data as shown are representative results from two or more experiments in each of the 4 subjects.

Figure 2

Figure 2

Frequencies and phenotypes of PA713-732 reactive T cells in non-AVA vaccines. A. PBMC from a DR0101 subject were stained with DR0101/PA713-732 PE-tetramers, CD4 antibody, and gating antibodies (anti-CD3, anti-CD14 and anti-CD19) ex vivo. B. PBMC were stained with DR0101/PA713-732 PE-tetramers, CD8 antibody, and gating antibodies ex vivo. Samples from A and B were obtained from the same subject and the frequency was determined to be 6 per million total CD4+ T cells. C. PBMC from a second subject were co-stained with DR0101/PA713-732 tetramers labeled with either PE or allophycocyanin and stained with gating antibodies (anti-CD4, anti-CD14 and anti-CD19) ex vivo. The frequency was determined to be 3 per million total CD4+ T cells. D. E and F. PBMC from DR0101 subjects were stained with DR0101/PA713-732 PE-tetramers, gating antibodies and anti-CD45RA, anti-CCR4 and anti-CD27 antibodies ex vivo. For each panel (A-F), we enriched using anti-PE magnetic beads and examined tetramer positive cells ex vivo, gating on forward and side scattering, and then on cells that were Via-Probe-, CD3+ or CD4+ , CD14- and CD19-. The percentages of tetramer positive cells that expressed the surface marker are as indicated. Data shown for each panel are representative results of more than 4 independent experiments.

Figure 3

Figure 3

Frequencies of DR0101 restricted PA401-420 reactive T cells and PA505-524 reactive T cells in DR0101 non-AVA vaccinees. A. PBMC from a DR0101 subject were stained with DR0101/PA401-420 PE-tetramers, DR0101/PA401-420 allophycocyanin -tetramers and gating antibodies ex vivo followed by enrichment with anti-PE magnetic beads. The frequency of PA401-420 specific T cells was determined to be 1.3 per million total CD4+ T cells. B. PBMC were stained with DR0101/PA401-420 PE-tetramers and CD45RA antibody, and gating antibodies ex vivo then enriched. The frequency of PA401-420 specific T cells was determined to be 0.5 per million total CD4+ T cells. C. PBMC from a DR0101 subject were stained with DR0101/PA505-524 PE-tetramers, DR0101/PA505-524 allophycocyanin-tetramers and gating antibodies ex vivo then enriched. The frequency of PA401-420 specific T cells was determined to be 0.2 per million total CD4+ T cells. Each plot represents staining from a different DR0101 subject. For each panel (A-C), we gated on forward and side scattering and then on cells that were Via-Probe-, CD4+ , CD14- and CD19-. Data shown in A and B are representative of more than 11 experiments in 11 subjects, and the data shown in C are representative of data from 7 different subjects.

Figure 4

Figure 4

Frequencies of DR0101 restricted PA reactive T cells in DR0101 non-AVA vaccinees. Each symbol within each PA epitope represents staining for a different subject. Frequencies less than 0.1 per million were below the threshold of detection. The p values were determined by using two-tailed unpaired t tests.

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