Increased gene expression of IL-23 and IL-23R in the colon during GVHD. (A,B) Lethally irradiated (900 cGy) Balb/c mice were transplanted with B6 BM (10 × 10⁶) and spleen cells (0.4-0.6 × 10⁶; n = 9-14, ■) or IL-23^−/− BM (10 × 10⁶) and spleen cells adjusted to yield an equivalent number of mature T cells (n = 9-14, □). Mice were killed 28 days after transplantation. RNA was extracted from spleen, colon, and liver tissues obtained from these animals as well as from normal nontransplanted Balb/c mice (n = 4-6, ▫). Gene expression of IL-23 (A) or IL-23R (B) was analyzed by real-time quantitative PCR as described in “Methods.” Data are cumulative results from 3 independent experiments for mice transplanted with B6 or IL-23^−/− marrow grafts. (C,D) Lethally irradiated Balb/c mice were transplanted with B6 BM (10 × 10⁶) and spleen cells (0.4-0.6 × 10⁶) and killed 28 days posttransplantation. LPMCs and spleen cells were isolated from pooled colons and spleens (n = 4-5/group per experiment), respectively. Spleen cells from normal nontransplanted B6 mice served as controls. CD11c⁺/CD11b⁺ and CD11b⁺/CD11c⁻ cells were sorted from LPMCs (■) and spleen cells (□) of GVHD mice and from normal splenocytes (▫). CD4⁺ T cells were sorted from splenocytes, liver lymphocytes, and colon LPMCs of GVHD animals and from spleen cells of normal mice. RNA was extracted from these cells, and gene expression of IL-23 (C) and IL-23R (D) was analyzed by real-time quantitative PCR. Data are normalized for 18S ribosomal RNA and presented as fold increase over 18S RNA (± SEM). Cumulative results from 3 independent experiments are shown. Statistics: *P ≤ .05; **P < .01.