Interleukin 23-mediated colonic damage is dependent upon donor-derived IFN-γ, but not IL-17. (A-C) Lethally irradiated (900 cGy) Balb/c mice were transplanted with B6 BM and spleen cells (0.4-0.6 × 10⁶; ■) or IL-23^−/− BM and spleen cells adjusted to yield an equivalent number of mature T cells (□). Mice in each cohort were killed 28-30 days after transplantation. Lymphocytes were isolated from the spleen (A), liver (B), or colon (C) of mice (n = 4-5/group per experiment) and pooled suspensions of these cells were stimulated with PMA and ionomycin in the presence of GolgiStop and then intracellularly stained with IFN-γ and IL-17–specific antibodies. Data are presented as the mean absolute number of CD4⁺ IFN-γ⁺ or CD4⁺ IL-17⁺ cells (± SEM) and are cumulative results from 4 independent experiments. (D) Lethally irradiated Balb/c mice were transplanted with marrow grafts from either B6 or IL-23^−/− donor mice as in panels A through C. Cohorts of mice (n = 8/group) were killed at the indicated time points (days 4, 7, 14, 21, and 28), and colonic tissue supernatants from individual mice were collected and analyzed for IL-17A by multiplex. Data are presented as mean amount of cytokine divided by the weight of cultured colon tissue (± SEM) and are cumulative results from 2 experiments. (E) Lethally irradiated Balb/c mice were transplanted with B6 BM (10 × 10⁶) and spleen cells (0.4-0.6 × 10⁶; ■, n = 9) or IL-17^−/− BM (10 × 10⁶) and spleen cells adjusted to yield the same T-cell dose (□, n = 9). Mice were killed 35 days posttransplantation and examined for overall (lung, liver, and colon) and colon-specific pathology using the semiquantitative scoring system detailed in “Histologic analysis.” Data are cumulative results from 2 independent experiments. (F-I) Lethally irradiated Balb/c mice were transplanted with B6 BM (10 × 10⁶) and spleen cells (adjusted to yield a T-cell dose of 0.225 × 10⁶ per mouse; ■, n = 6) or IFN-γ^−/− BM (10 × 10⁶) and spleen cells adjusted to yield the same T-cell dose (□, n = 5). Mice were killed 30 to 33 days posttransplantation, and the colon was examined for pathologic damage (F). Data are presented as the mean plus or minus SEM and are the cumulative results from 2 experiments. Colon tissue from the same mice was also cultured overnight in medium as described in “Methods” and assayed for levels of LPS (G), IL-23p19 (H), and proinflammatory cytokines (I). Data are presented as the mean amount of cytokine or LPS divided by the weight of cultured colon tissue (± SEM). *P ≤ .05; **P < .01.