Antiretroviral therapy reduces the magnitude and T cell receptor repertoire diversity of HIV-specific T cell responses without changing T cell clonotype dominance

Abstract

After initiation of antiretroviral therapy (ART), HIV loads and frequencies of HIV epitope-specific immune responses decrease. A diverse virus-specific T cell receptor (TCR) repertoire allows the host to respond to viral epitope diversity, but the effect of antigen reduction as a result of ART on the TCR repertoire of epitope-specific CD8(+) T cell populations has not been well defined. We determined the TCR repertoires of 14 HIV-specific CD8(+) T cell responses from 8 HIV-positive individuals before and after initiation of ART. We used multiparameter flow cytometry to measure the distribution of memory T cell subsets and the surface expression of PD-1 on T cell populations and T cell clonotypes within epitope-specific responses from these individuals. Post-ART, we noted decreases in the frequency of circulating epitope-specific T cells (P = 0.02), decreases in the number of T-cell clonotypes found within epitope-specific T cell receptor repertoires (P = 0.024), and an overall reduction in the amino acid diversity within these responses (P < 0.0001). Despite this narrowing of the T cell response to HIV, the overall hierarchy of dominant T cell receptor clonotypes remained stable compared to that pre-ART. CD8(+) T cells underwent redistributions in memory phenotypes and a reduction in CD38 and PD-1 expression post-ART. Despite extensive remodeling at the structural and phenotypic levels, PD-1 was expressed at higher levels on dominant clonotypes within epitope-specific responses before and after initiation of ART. These data suggest that the antigen burden may maintain TCR diversity and that dominant clonotypes are sensitive to antigen even after dramatic reductions after initiation of ART.

Fig 1

Epitope-specific responses contract while clonotypic dominance remains intact. (A) Epitope-specific T cell populations and constituent clonotypes were identified by colabeling with tetramer and anti-TRBV antibodies. Frequencies of epitope-specific populations as part of the CD8⁺ T cell population are noted at pre-ART and post-ART time points. (B) Dominant clonotypes were identified by sequencing and labeled with anti-TRBV antibodies, and the frequencies of dominant clonotypes as part of the epitope-specific population are noted at corresponding pre-ART and post-ART time points. The dotted vertical line at day 0 indicates initiation of ART. Subject, epitope identifier, and dominant TRBV family are indicated for each response studied.

Fig 2

Diversity in the HIV epitope-specific TCR repertoire is reduced after initiation of ART. (A) At pre-ART (unshaded charts) and post-ART (shaded charts) time points, Shannon's entropy (plotted on the y axis) was calculated and shown for amino acid positions within the TRBV CDR3 region (plotted on the x axis) for each of 12 individual epitope-specific T cell responses. Mean entropy for the TCR repertoire is shown in the upper left corner of each chart. The P value for statistical comparison (Wilcoxon matched-pairs test) is shown in the upper right corner of each chart. Pre-ART and post-ART comparisons between independent diversity measurements are shown in panels B to D. (B) Number of distinct clonotypes within the TCR repertoire identified at each time point. (C) Mean Shannon entropy value for each TCR repertoire. (D) Simpson's diversity index for each clonotypic repertoire.

Fig 3

T cell memory populations are repopulated after initiation of ART. Memory cell subsets were identified based on expression of CD45RO and CCR7. (A) Representative memory cell distributions on CD8⁺ (blue) and epitope-specific (orange) T cell populations are shown pre-ART and post-ART. (B) Frequencies of CD45RO⁺ CCR7⁺ T_cm and CD45RO⁻ CCR7⁺ T_naive cell populations as proportions of the overall CD8⁺ T cell population are increased post-ART.

Fig 4

CD8⁺ T cell activation is reduced after ART, while epitope-specific changes are variable. T cell activation was measured by surface expression of CD38 and PD-1 molecules. (A) Representative plots are shown for CD8⁺ (blue) and epitope-specific (orange) populations. (B) Expression of CD38 and PD-1 as measured by MFI was reduced on CD8⁺ T cell populations after initiation of ART.

Fig 5

Dominant clonotypes express higher levels of PD-1 than subdominant clonotypes before and after ART. PD-1 expression was measured on dominant and subdominant clonotypes within epitope-specific responses before and after initiation of ART. (A) Representative histograms show PD-1 expression on dominant (blue) and subdominant (green) clonotypes before and after ART. The MFI value is shown in the upper right corner of each histogram for each clonotypic population. (B) PD-1 expression levels on dominant and subdominant clonotypes of 13 epitope-specific responses were compared at pre-ART and post-ART time points (Wilcoxon matched-pairs test).