Treg cell effect on M CD8⁺ T-cell apoptosis is rather cytokine dependent. (A) Histogram overlays representing PD-L1 expression on CD8⁺ T cells stimulated with anti-CD3 for 48 hours in the presence of CD4⁺CD25⁻ (open histogram) or Treg cells (gray histogram), either mixed in the lower chamber of a transwell plate (left) or separated by a semipermeable membrane (right). (B) Annexin V staining of anti-CD3–activated M CD8⁺ T cells. Cocultures of CD8⁺ T cells with either CD4⁺CD25⁻ T cells (left) or Treg (right) were set in transwell plates as follows: CD8⁺ and CD4⁺ T cells were either mixed in the lower chamber, or separated by a semipermeable membrane. Histograms represent CD27⁺CD45RA⁻CD8⁺-gated cells. Percentage of annexin V⁺ cells is shown, based on negative control. One representative experiment of 2 with similar results is shown. The numbers correspond to the percentage of annexin V⁺ cells based on negative control staining.