TNF-TNFR2 interactions promote MHC class II-stimulated alloresponses while TNF-TNFR1 interactions promote MHC class I-stimulated alloresponses. The present studies were designed to evaluate whether TNF-TNFR2 interactions were involved in the in vivo generation of CD4+ T cell-mediated intestinal graft-versus-host disease (GVHD) in the (C57BL/6J (hereafter called B6)→ B6× B6. CH-2 bm12 (bm12)) F 1 GVHD model. Briefly, 5× 10 6 splenic CD4+ T lymphocytes from B6. TNFR2−/− or control B6 mice were transferred with 1–2× 10 6 T cell-depleted B6 bone marrow cells (BMC) to irradiated MHC class II-disparate (bm12× B6) F 1 mice. Weight loss, intestinal inflammation, and the surface expression of CD45RB (memory marker) on intestinal and splenic lymphocytes were assessed. IL-2 and IFN-α mRNA levels in intestinal lymphocytes were assessed by nuclease protection assays. A significant reduction in weight loss and intestinal inflammation was observed in recipients of the TNFR2−/− CD4+ SpC. Similarly, a significant decrease was noted in T cell numbers and in CD45RB low (activated/memory) expression on intestinal but not CD4+ T cells in recipients of TNFR2−/− CD4+ spleen cells. IL-2 and IFN-α mRNA levels were reduced in the intestine in the recipients of TNFR2−/− splenic CD4+ T cells. These results indicate that TNF-TNFR2 interactions are important for the development of intestinal inflammation and activation/differentiation of Th1 cytokine responses by intestinal lymphocytes in MHC class II-disparate GVHD while playing an insignificant role in donor T cell activation in the spleen.