DZNep inhibits histone methylation, reduces cellular Ezh2, and alters gene expression in activated CD4⁺ T cells. (A-B) Naive CD4⁺T cells were isolated from B6 mice and then stimulated with anti-CD3 and anti-CD28 Abs. Five days later, those activated T cells were further treated with different doses of DZNep for 24 hours. Live cell number was counted, and the recovery rate (output/input) was calculated (A). (B) Dot plots show the expression of annexin V, and the accompanying graph shows the percentage of annexin V⁺ cells. (C) CD4⁺ T_N cells were stimulated with IL-7 or IL-2 in the presence or absence of DZNep for 24 hours. Graphs show the percentage of dead cells after culture (mean ± SD). (D-G) CD4⁺ T_N cells were stimulated with anti-CD3 and anti-CD28 Abs for 5 days later and then treated with DZNep. Western blots show the amount of trimethylated histones (D) and Ezh2 protein (F) in activated T cells treated with DZNep for 72 hours. Real-time RT-PCR analyses show the expression genes in T cells treated with or without DZNep (200nM) for 24 hours. (H) Lethally irradiated BALB/C recipients [8 Gy (800 rads)] were transplanted with B6 mice–derived TCD BM (5 × 10⁶) or with TCD BM + B6 wild-type (WT) or Bim^−/− mice–derived CD4⁺ T_N cells (1 × 10⁶) + CD8⁺ T_N cells. T-cell recipients were treated with vehicle (Control) or 12 doses of DZNep (DZNep) from day 0 to day 27 after transplantation. Survival was monitored.