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. Author manuscript; available in PMC: 2019 May 9.
Published in final edited form as: Sci Immunol. 2018 Nov 9;3(29):eaao2892. doi: 10.1126/sciimmunol.aao2892

Figure 3. Impact of iKIR-HLA interaction on CD8+ T cell survival in vitro & iKIR expression ex vivo.

Figure 3

A. CD8+ T cell lines (plots from left ED, KIR2DL3+; VTE1 KIR2DL3+; SK1 KIR3DL1+) were cocultured with superantigen either without adding ligand for KIR i.e. with 221 cells or with 221 cells transfected to express irrelevant, non-cognate HLA (open bars); with ligand i.e. 221 transfected to express cognate HLA with or without isotype control antibody (black bars) and with KIR-HLA blocking i.e. 221 transfected to express cognate HLA with addition of antibody to block KIR (GL183, DX9) or to block HLA (DX17) (red bars). The count of live CD8+ T cells after 5 days is shown (mean± SE). ED: 5 experiments each consisting of 3 replicates; VTE1: 1 experiment, 4 replicates; SK1: 1 experiment, 2 replicates. It is consistently seen that there are greater numbers of viable T cells with isotype control than when the KIR-HLA interaction is blocked or when cognate ligand is absent.

B. Metaanalysis of survival of the 3 KIR-expressing CD8+ T cell lines. Block HLA. Coculture with 221 cells transfected to express cognate HLA with isotype control antibody compared to coculture with 221 cells transfected to express cognate HLA with anti-HLA antibody. Median difference = 48% P = 0.001. Block KIR. Coculture with 221 cells transfected to express cognate HLA with isotype control antibody compared to coculture with 221 cells transfected to express cognate HLA with anti-KIR antibody. Median difference = 40% P = 0.001. Irrelevant HLA. Coculture with 221 cells transfected to express cognate HLA compared to coculture with 221 cells transfected to express irrelevant, non-cognate HLA. Median difference = 57% P =1.5x10-5. 221 only. Coculture with 221 cells transfected to express cognate HLA compared to coculture with 221 cells. Median difference = 53% P = 0.002. Survival of VTE1 plotted on left hand y axis (for all conditions), ED and SK1 plotted on right hand y axis (for all conditions). We consistently found that absence of KIR:HLA ligation (either by blocking HLA, blocking KIR, adding a non-ligating HLA or not adding HLA) resulted in significantly decreased CD8+ T cell survival.

C. PBMCs from 3 HLA-B27+ individuals with ankylosing spondylitis were left untreated (open bars); cultured with superantigen (SEB) with or without isotype control antibody (black bars) or cultured with SEB with antibodies that block KIR3DL2 or homodimeric HLA-B27 (red bars). The count of live KIR3DL2+ CD4- T cells after 5 days is shown, (mean± SE). Blocking the iKIR-HLA interaction with specific antibodies decreased the count of live KIR3DL2+ T cells compared to isotype control (block KIR: median difference=63%; block HLA: median difference=50%).

D. The KIR2DL3-expressing CD8+ T cell line VTE1 was cultured as in A. The number of live CD8+ CD107a+ T cells was quantified by flow cytometry at day 5.

E. The fraction of pentamer+ CD8+ T cells expressing iKIRs (KIR2DL1, KIR2DL2/L3, KIR3DL1 or KIR3DL2) directly ex vivo is plotted for HLA-A2+ individuals infected with HIV-1 (N = 16) or HTLV-1 (N = 9).