Id3 maintains the GVHD-inducing capacity of alloreactive T cells. Balb/c mice were subjected to total body irradiation (4.5 Gy on day −1 and 4 Gy on day 0) followed by infusion of 5 × 10⁶ B6 × B6/SJL F1 mouse (CD45.1⁺CD45.2⁺) TCD bone marrow (TCD-BM) alone or together with 5 × 10⁵ B6 (CD45.1^-CD45.2⁺) naïve WT or Id3-cKO CD4⁺ T cells. (A) Survival of Balb/c recipients. (B) Cutaneous GVHD in Balb/c mice receiving WT and Id3-cKO CD4⁺ T cells, day 22 after transplantation in at least 8 mice per group. (C) Intestine from Balb/c recipients were harvested from day 14 to 22, sectioned, and stained with hematoxylin and eosin. Slides were scanned with Leica Aperio VERSA slide scanner and visualized with Aperio ImageScope (20×). N = 8. (D-G) Balb/c recipients were euthanized at the indicated time points after HSCT. Tissues were harvested for the analysis of donor T-cell immune response using flow cytometry. Id3-cKO donor CD4⁺ T cells exhibited normal cytokine-producing capacity represented by the similar percentage (D) and number (E) of IFN-γ⁺ cells in the spleen, mesenteric lymph node (mLN), and liver. (F) Graphs show the number of intestine-infiltrated donor CD4⁺ T cells and those with IFN-γ–producing capacity recovered in Balb/c mice that underwent transplantation with Id3-cKO T cells. (G) Graphs show the frequency of donor T cells expressing α4β7, CCR5, and CCR9. ∗P < .05; ∗∗∗P < .001; Student t test was used for 2-group comparison. At least 4 mice per group were analyzed.