WT Treg and Lag-3^−/− Treg show similar protection against GVHD.

<p>(A) Foxp3 staining (upper panel) and Lag-3 staining (lower panel) of CD4⁺CD25⁺ T cells isolated from WT mice (black histogram) and Lag-3^−/− mice (dashed histogram). For Lag-3 staining, lower panel, regulatory T cells were activated for 2 days with anti-CD3/anti-CD28 beads. Cells were gated on CD4⁺Foxp3⁺. The shaded histogram represents the isotype staining control. The histograms are representative of 2 independent experiments. (B) ³H-thymidine incorporation of WT C57BL/6J responders to Balb/c stimulators in the presence of either WT Treg or Lag-3^−/− Treg at different Treg to responder ratios, R: responders, S: stimulators. (C–D) Lethally irradiated Balb/c recipients were co-transplanted with 5×10⁶ TCD-BM and 5×10⁵ WT or Lag-3^−/− Treg on day 0 followed by infusion of 1×10⁶<i>luc⁺</i> WT Tcon on day 2. (C) Percent survival of mice after transplantation. Graph contains data pooled from 3 independent experiments (n = 15). P = 0.034 for WT Treg+Tcon vs. Tcon and P = 0.033 for Lag-3^−/− Treg+Tcon vs. Tcon. (D) <i>In vivo</i> BLI images of mice receiving WT Tcon alone, WT Tcon+WT Treg and WT Tcon+Lag-3^−/− Treg. Upper panels: BLI images taken at different time points after transplantation; lower panels: the corresponding quantitative analysis of the BLI signal. The BLI images are representative of 3 independent experiments.