Immunodominant HIV-1-specific HLA-B- and HLA-C-restricted CD8+ T cells do not differ in polyfunctionality

Abstract

HIV-1 specific HLA-B-restricted CD8+ T cell responses differ from HLA-C-restricted responses in antiviral effectiveness. To investigate possible reasons for these differences, we characterized the frequency and polyfunctionality of immmunodominant HLA-B*57/B5801- and HLA-Cw*07-restricted CD8+ T cells occurring concurrently in nine study subjects assessing IFN-gamma, TNF-alpha, IL-2, MIP-1beta, and CD107a by flow cytometry and analyzed sequence variation in targeted epitopes. HLA-B*57/5801 and HLA-Cw*07 restricted CD8+ T cells did not differ significantly in polyfunctionality (p=0.84). Possession of three or more functions correlated positively with CD4+ T cell counts (r=0.85; p=0.006) and monofunctional CD8+ T cells inversely correlated with CD4 cell counts (r=-0.79; p=0.05). There were no differences in polyfunctionality of CD8+ T cells specific to wildtype versus mutated epitopes. These results suggest that loss of polyfunctionality and increase in monofunctional HIV-1-specific CD8+ T cells are associated with disease progression independent of restricting HLA allele. Furthermore, sequence variation does not appear to significantly impact CD8+ T cell polyfunctionality in chronic HIV-1 infection.

Fig. 1

Measurement of HLA-B*57/5801 and HLA-Cw*07-restricted T cell responses by Interferon-gamma ELISPOT assay. (A) Hierarchy of dominant epitopes presented by the study subjects expressing HLA-B*57/5801 and HLA-Cw*07 alleles (n = 9). Dotted line indicates responses above 500 SFCs from ELISPOT assay that were further tested in subsequent multicolor assays. (B) Combined total magnitude of T cell responses presented by both HLA-B*57/B5801 and HLA-Cw*07 restricted epitopes in the study individuals (n = 9). (C) Percentage of epitope recognition (>100 SFCs/10⁶ ELISPOT responses) of the dominant epitopes presented by different HLA-B57/5801 alleles (B*5701, B*5703 and B*5801) in all chronically infected subjects coexpressing HLA-Cw*07 (n = 37).

Fig. 2

Assessment of HIV-1 specific CD8+ T cell polyfunctionality by multicolor staining for HLA-B*57/5801 and HLA-Cw*07-restricted epitopes. (A) Magnitude of HLA-B*57/5801 and HLA-C restricted epitope responses using multicolor staining, single CD8+ T cell function responses are shown after background subtraction. (B) Comparison of the contribution of individual functions between HLA-B*57/B5801 (○) and HLA-C (●) restricted epitopes in the study subjects (n = 9). The fractions of the response patterns are grouped and color-coded by the number of functions and summarized in pie chart form where each slice of the pie represents the fraction of the total epitope-specific response that consist of CD8+ T cells with the respective number of functions.

Fig. 3

Relationship between the fractions of monofunctional, bi-functional and polyfunctional HIV-1 specific CD8+ T cell responses and the (A) CD4+ T cell counts and (B) viral loads.

Fig. 4

Evaluation of the relationship between epitope sequence variation and polyfunctionality of HIV-specific CD8+ T cell responses. HLA- B*57/5801 and HLA-Cw*07 restricted epitope responses with and without sequence variation were plotted against the percentage of total HIV-1 specific CD8+ T cells with ≥ 3 functions, bi-functional and monofunctional CD8+ T cell responses generated from the study individuals (n = 9).