The application of allogeneic stem cell transplantation is limited by graft-vs.-host disease (GVHD). GVHD can be divided into acute and chronic forms that likely have different initiation requirements and pathogenic mechanisms. Previously we demonstrated that residual host antigen presenting cells (APC) were required for CD8-mediated acute GVHD (aGVHD). In contrast, here we show that either donor or host APCs can initiate CD4-mediated GVHD in the B10.D2(H-2d)→BALB/c (H-2d) model of chronic GVHD (cGVHD). We selectively impaired donor APCs, host APCs or both by using donors and/or hosts deficient in both costimulatory molecules CD80 and CD86 (CD80/86−/−). When all CD28:CD80/86 signaling was prevented by using CD80/86−/− donors and hosts, wild type (WT) CD4 cells were unable to induce cGVHD. Therefore donor CD4 cells absolutely require signals from CD80/86 to mediate cGVHD. Next we compared cGVHD in CD80/86−/− BM + CD4→WT vs. WT BM + CD4→CD80/86−/− hosts, to debilitate antigen presentation by donor or host APCs, respectively. Cutaneous cGVHD developed in both groups, demonstrating that donor or recipient APCs are sufficient to initiate disease. However, the incidence of cutaneous cGVHD in CD80/86−/− recipients was less than that in WT→WT recipients (P<0.01), suggesting that recipient APCs are more important than donor APCs for skin cGVHD. In support of this, cGVHD was not reduced inCD80/86−/− BM + CD4 →WT recipients. Therefore, reconstitution with defective donor APCs does not affect disease when host APCs are intact. To address whether host APCs were dispensable, we compared cGVHD in donor→host (APCs >98% donor) and host→host chimeric recipients. Similar cGVHD developed in both chimeras; thus donor APCs alone are sufficient, consistent with the data in CD80/86−/− hosts. Interestingly, cGVHD was initiated in the absence of significant host hematopoietic antigen. Thus, exogenously acquired parenchymal antigen was sufficient. The above experiments focused on cutaneous GVHD. However, recipients of WT BM versus non-WT BM consistently developed diarrhea and increased weight loss. While CD4 recipients of CD80/86−/− or CD40−/− BM regained and maintained their original weights, CD4 recipients of WT BM never returned to their pre-transplant weight (P<0.01 after day 15). Consistent with the clinical finding, recipients of CD4 cells and WT BM had higher bowel pathology scores than equivalent mice that received CD80/86−/− BM (P<0.01). Recipients of CD80/86−/− BM + WT CD4 cells had statistically indistinguishable path scores from recipients of either type of BM without CD4 cells (P=0.19 and 0.73); therefore, without CD80/86 expression on donor BM, there was no statistical evidence that donor T cells caused bowel GVHD. These results provide new insights into the role of donor APCs in GVHD and demonstrate that APC requirements differ depending on the site of disease, both novel findings. They also identify differences in APC requirements between CD8-mediated aGVHD and CD4-mediated cGVHD and further highlight APCs as additional targets for GVHD prevention and therapy.