CD3⁺CD4^low and CD3⁺CD8^low cells are suppressor T-cell subsets whose CD45RA, CD62L, and HLA-DR expression is modulated. (A-C) T cells primed with either APCs (left panel) or APC-HLA-G1s (right panel) were analyzed by 4-color flow cytometry at day 6 of MLR for expression of (A) CD45RA, (B) CD62L, and (C) HLA-DR among CD4⁺ and CD8⁺ T cells. Results are shown as dot plots. Numbers represent the percentage of cells within the corresponding region. One representative experiment of 3 is shown. (D-E) T cells were primed either with (D) APCs or APC-HLA-G1s or with (E) HLA-G5⁻ or HLA-G5⁺ SNs for 6 days. Then, CD4⁺ and CD8⁺ T cells were sorted. These purified cells were used in an immunosuppressive assay as γ-irradiated third-party cells in MLR with HLA-mismatched PBMCs at a responder–stimulator–third-party cell ratio of 1:1:1. Results are expressed as percentage of alloproliferation inhibition (%) ± SEM in triplicate wells. Alloproliferation inhibition percentage observed with CD3⁺CD4^low or CD3⁺CD8^low T cells was calculated, based on the alloproliferation observed with CD3⁺CD4^high and CD3⁺CD8^high T cells, respectively.