2B4+ CD8+ T cells play an inhibitory role against constrained HIV epitopes

Abstract

Cytotoxic T cells play a critical role in the control of HIV and the progression of infected individuals to AIDS. 2B4 (CD244) is a member of the SLAM family of receptors that regulate lymphocyte development and function. The expression of 2B4 on CD8+ T cells was shown to increase during AIDS disease progression. However, the functional role of 2B4+ CD8+ T cells against HIV infection is not known. Here, we have examined the functional role of 2B4+ CD8+ T cells during and after stimulation with HLA B14 or B27 restricted HIV epitopes. Interestingly, IFN-γ secretion and cytotoxic activity of 2B4+ CD8+ T cells stimulated with HIV peptides were significantly decreased when compared to influenza peptide stimulated 2B4+ CD8+ T cells. The expression of the signaling adaptor molecule SAP was downregulated in 2B4+ CD8+ T cells upon HIV peptide stimulation. These results suggest that 2B4+ CD8+ T cells play an inhibitory role against constrained HIV epitopes underlying the inability to control the virus during disease progression.

Fig. 1

HIV antigen pulsed APC stimulation inhibits IFN-g production in 2B4⁺ CD8⁺ T cells. 2B4⁻ and 2B4⁺ CD8⁺ T cells were primed either without peptide or with Influenza and HIV-peptide pulsed APCs. IFN-γ released was measured by ELISA after first and second stimulation with APCs, approximately 8 and 16 days respectively. PBMCs from HIV-negative HLA B14 donor (BLCL-2) after 8 days (A) and 16 days of stimulation (B). (*- p<0.05; 2 way ANOVAs detected significant differences between 2B4⁺ CD8⁺ T cells stimulated with HIV peptides and FL-8).

Fig. 2

Cytotoxic activity against peptide pulsed targets is decreased after HIV-peptide pulsed APC stimulation. 2B4⁺ and 2B4⁻ CD8⁺ T cells primed with two rounds of HIV-peptide pulsed APCs were analysed for their cytotoxic activity against peptide pulsed BLCLs in a standard 4 hr⁵¹Cr release assay. CD8⁺ T cells used as effectors and the target cells BLCL-2 were from HLA B14 donor (A), CD8⁺ T cells used as effectors and the target cells BLCL-3 were from HLA B27 donor (B). Data shown is at an E:T ratio of 20:1. Each data point is the mean ± SEM of two independent experiments done in triplicates. (*- p<0.05; 2 way ANOVAs detected significant differences between 2B4⁺ CD8⁺ T cells stimulated with HIV peptides and FL-8).

Fig. 3

SAP is downregulated in 2B4⁺ CD8⁺ T cells upon HIV peptide stimulation. 2B4⁺ and 2B4⁻ CD8⁺ T cells before stimulation (day 0) and after HIV peptides and influenza peptide (FL-8) stimulation (after day 16) were harvested and total RNA was extracted and SAP expression was evaluated by RT-PCR for HLA B14, B8 donor (A) and HLA B27, B44 donor (B). The bands were subjected to densitometry analysis and the relative density of HIV peptide and influenza peptide stimulated samples were normalized with respect to no peptide sample and the data was represented as fold change.