Changes in function of HIV-specific T-cell responses with increasing time from infection

Abstract

Recently HIV-infected individuals have virus-specific responses characterized by IFN-gamma/IL-2 secretion and proliferation rarely seen in chronic infection. To investigate the timing of loss of HIV-specific T-cell function, we screened cells from 59 treatment-naïve HIV-infected individuals with known dates of infection for proteome-wide responses secreting IFN-gamma/IL-2 and IFN-gamma alone by ELISPOT. HIV peptide-specific proliferation was assessed by carboxyfluorescein diacetate succinimidyl ester (CFSE) dilution. The contribution of IFN-gamma/IL-2 and IFN-gamma-only secretion to the total HIV-specific response was compared in subjects infected <6, 6-12, and 12-36 mo earlier. The frequency of IFN-gamma/IL-2-secreting cells fell, while that of IFN-gamma-only secretion rose with time from infection. HIV peptide-specific proliferative responses were almost exclusively mediated by CD8(+) T cells, and were significantly lower in cells obtained from the 12-36 mo versus < 6 mo post-infection groups. By the second year of infection there was a significant difference in these functions compared to those assessed within 6 mo.

FIG. 1.

HIV-specific functional subsets in subjects categorized by duration of infection. Thirty-five subjects infected <6 mo, 19 infected between 6 and 12 mo, and 35 infected between 12 and 36 mo were screened for immune responses to the entire HIV proteome using a dual-color ELISPOT assay. The magnitude of proteome-wide HIV-specific responses characterized by secretion of IFN-γ/IL-2 is shown in panel A, and IFN-γ only in panel B, for the three groups categorized by time post-infection. The y-axis shows for each individual tested the magnitude of the HIV-specific response in spot-forming cells per million peripheral blood mononuclear cells (SFC/10⁶ PBMCs). The percentage contribution of HIV-specific responses characterized by IFN-γ/IL-2 secretion and IFN-γ-only secretion to the total response is shown in panels C and D, respectively. Panels E and F show the same analyses as in panels C and D, but with individuals carrying protective HLA alleles excluded from the analysis. The line through each scatterplot indicates the median response for the groups.

FIG. 2.

HIV-specific proliferation of CD4⁺ and CD8⁺ T cells in samples obtained at times within 6 mo and after 6 mo of infection to peptides stimulating either IFN-γ/IL-2- or IFN-γ-only secretion. Panel A shows an example of a CFSE dilution experiment. Peripheral blood mononuclear cells were cultured with the stimuli shown on the right for 6d. The boxes show the percentage of CFSE^lo cells in the CD4 (left) and CD8 (right) T cell compartments. The scatterplots in panel B show the percentage of CD3⁺ CFSE^lo cells detected following a 6-day stimulation with HIV peptides. The left-hand plots show the percentage of CFSE^lo T cells in the CD4⁺, while the right-hand plots show the percentage of CFSE^lo T cells in the CD8⁺ compartments following stimulation of cells from an individual infected <6 mo. Panel C shows the percentage of CFSE^lo CD4⁺ and CD8⁺ T cells generated following the stimulation of PBMCs from 17 individuals with a panel of 81 peptides that stimulated IFN-γ/IL-2 (right-hand scatterplots), or 22 peptides that induced IFN-γ-only secretion (left-hand scatterplots) in these subjects. Panel C shows the percentage of CFSE^lo CD4⁺ and CD8⁺ T cells generated following stimulation of cells from six individuals infected more than 6 mo with a panel of 28 peptides that stimulated IFN-γ/IL-2 (right-hand scatterplots), or 10 peptides that induced IFN-γ-only secretion (left-hand scatterplots). Panel D shows changes in HIV-peptide-specific proliferation as measured by the percentage CFSE^lo cells. Paired samples from two time points from five study subjects were stimulated with a panel of 27 HIV peptides that were stimulatory at both time points in a dual-color ELISPOT assay. One sample was from a time point within 6 mo of infection, the other from a later time point in infection. A Wilcoxon matched pairs signed-ranks test was used to assess the significance of between time-point changes in the percentage of CFSE^lo cells as a measure of HIV-peptide-specific proliferative capacity.