GPR109A^−/− T cells undergo increased apoptosis. Lethally irradiated BALB/c recipients received B6 WT TCD BM and 5 × 10⁶ WT or KO T cells. (A) Donor CD4⁺ and CD8⁺ T cell numbers were analyzed from spleen on day 3 post allo-HCT. (B) Donor regulatory T cells from spleen on day 3 post allo-HCT. (C) Expression of LPAM-1 on splenic donor CD4⁺ and CD8⁺ T cells on day 3 post allo-HCT. (D) T cells were labeled with CFSE prior to transplant and proliferation determined by measuring CFSE^low cells. (E-H) Early timecourse at day 1, 2, and 3 post allo-HCT measuring numbers of CD4⁺ and CD8⁺ T cells (E), LPAM-1⁺ CD4⁺ and CD8⁺ T cells (F) CD4⁺ and CD8⁺ T cells expressing activation markers, CD25 (G) and PD-1 (H). (I) Frequency of Annexin⁺DAPI^- apoptotic donor CD4⁺ T cells were analyzed on day 3 post alloHCT. (J-K) Frequency of donor CD4⁺ (top) and CD8⁺ (bottom) T cells on day 3 post allo-HCT, expressing other apoptotic markers Bim, cleaved Caspase-3 (J) and Fas(K). (L-N) MACS sorted splenic T cells were stimulated in vitro with anti-CD3/CD28 for 24, 48, or 72 hours and frequency of apoptotic cells measured by flow cytometry gating on live CD3⁺ and Annexin⁺DAPI^- (L), cleaved caspase-3⁺, Bim⁺ (M) or Fas⁺ (N). All comparisons in (A) to (M) were performed by two-tailed unpaired Mann-Whitney Test. Values are means ± standard deviation. *P < 0.05, **P < 0.01, ***P < 0.001, n = 15 mice per group. All results from two to three independent experiments.