Differential regulatory T cell activity in HIV type 1-exposed seronegative individuals

Abstract

The potential role of conventional and regulatory T cells (Tregs) in protection from HIV-1 infection remains unclear. To address this question, we analyzed samples from 129 HIV-1-exposed seronegative individuals (HESN) from an HIV-1-serodiscordant couples cohort. To assess the presence of HIV-specific T cell responses and Treg function, we measured the proliferation of T cells in response to HIV-1 peptide pools in peripheral blood mononuclear cells (PBMCs) and PBMCs depleted of Tregs. We identified HIV-specific CD4(+) and CD8(+) T cell responses and, surprisingly, the overall CD4(+) and CD8(+) T cell response rate was not increased when Tregs were removed from cell preparations. Of the 20 individuals that had HIV-1-specific CD4(+) T cell responses, only eight had Tregs that could suppress this proliferation. When compared with individuals whose Tregs could suppress HIV-1-specific CD4(+) T cell proliferation, individuals with Tregs unable to suppress showed a trend toward increased T cell activation and Treg frequency and a significant increase in HIV-1-specific production of microphage inflammatory protein-1β (MIP-1β) by CD4(+) T cells, autocrine production of which has been shown to be protective in terms of HIV-1 infection of CD4(+) T cells.

Figures

FIG. 1.

HIV-driven T cell proliferation in HIV-exposed seronegatives. (A) The response rates for total peripheral blood mononuclear cells (PBMCs) (white bars) and PBMCs-Tregs (black bars) for CD4+ and CD8+ T cells were calculated as the percentage of positive proliferative responses out of the total number of analyzed samples. The McNemar test was used to compare the frequency of individuals with any proliferative response in PBMCs vs. in PBMCs-Tregs: for CD4+ T cells, p=0.37 and for CD8+ T cells, p=0.74. (B) A FACS-based sorting scheme was used for the T cell proliferation assay. The PBMCs from each sample were stained with propidium iodide (PI) for dead cell exclusion and with anti-CD4, CD8, CD127, and CD25, and divided in half. The PI-negative fraction was sorted from one aliquot, and considered the “PBMCs.” To prepare the “PBMCs-Treg” fraction, the second aliquot was also sorted based on PI negativity. Next, the CD4-negative cells were sorted and combined with the CD4+CD127+CD25– (T conv) cells to obtain the PBMCs-Tregs.

FIG. 2.

Treg depletion can result in an increased or decreased CD4+ and CD8+ T cell proliferation in different subjects. The ratios between the percentage of cells proliferating in response to a peptide pool and to DMSO are shown for samples where the Treg depletion caused an increased (A, C) and decreased (B, D) CD4+ and CD8+ T cell proliferation, respectively. Each data pair represents the response to one peptide pool. N=number of responses. The p value was calculated by the Wilcoxon signed rank test.

FIG. 3.

Lack of Treg suppressive capacity results in increased CD4+ T cell activation and microphage inflammatory protein-1β (MIP-1β) secretion. (A) Frequencies of Tregs and (B) activated CD4+ T cells are compared between the groups where the Tregs could or could not suppress CD4+ T cell proliferation. (C) Linear correlation between Treg frequency and CD4+ T cell activation (p=0.0008 by Spearman test). (D) Interferon (IFN)-γ, (E) tumor necrosis factor (TNF)-α, and (F) MIP-1β secretion by CD4+ T cells in response to stimulation by HIV-1 peptide pools are compared between the two groups. After background subtraction, the values in the graphs were calculated as the average between the responses upon stimulation with different peptide pools. The bars represent the mean between the samples. p=0.02, Wilcoxon rank sum test.