CD4⁺ iNKT cells inhibit proliferation and activation of alloreactive T cells. (A) Representative bioluminescence images during GVHD initiation (day +5, upper row) and affection of GVHD target sites (day +22, lower row). Bioluminescence signal derived from luc⁺ Tcons injected on day 0. (B) Bioluminescence signals throughout the experiment. Shown are 5 animals per group from 1 of 2 independent experiments. (C) Graphs depict the percentage of proliferating Thy1.1⁺ T cells from different secondary lymphoid organs on day +3 as measured by CFSE proliferation assay. Shown is 1 of 3 independent experiments performed in triplicate. (D) Absolute number of live Thy1.1⁺CD44^high T cells re-isolated from BALB/c recipient animals on day +6. Shown is 1 of 2 independent experiments performed in triplicate. (E) Expression of CD25 on live Thy1.1⁺CD44^high T cells. Gates were set on isotype controls. One representative example per group of 3 mice from 1 of 3 independent experiments is shown. †Indicates all animals from the respective group died or needed to be euthanized. Error bars indicate standard error of the mean. LVR, liver; mLN, mesenteric lymph nodes; pLN, peripheral lymph nodes; SPN, spleen. *P ≤ .05; **P ≤ .01; ***P ≤ .001.