Background.Graft-versus-host disease (GVHD) occurs in the recipient after small bowel transplantation (SBT). Proinflammatory cytokines, such as tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ), and interleukin 6 (IL-6), may be important mediators of GVHD. Increased expression of these cytokines might precede the clinical manifestations of GVHD induced by SBT.Methods.Heterotopic SBT was performed using Lewis donors into Lewis × Brown Norway F1 (LBN-F1) recipients. The isograft control was performed from LBN-F1 into LBN-F1. Animals were killed on the 5th and 11th postoperative day (POD). mRNA was isolated from recipient native small bowel, colon, spleen, liver, and mesenteric lymph nodes and from nonsurgical controls as baseline. Semiquantitative reverse transcriptase polymerase chain reaction was performed to amplify mRNA transcripts for TNF-α, IFN-γ, and IL-6 using α32P-dATP incorporation. Clinical signs, histologic assessment, and cytokine expression were correlated.Results.On POD 5, there were neither clinical signs nor histologic features of GVHD, but mRNA expression of TNF-α and IL-6 in small bowel, IL-6 in spleen, and IFN-γ in mesenteric lymph nodes were significantly increased in allograft animals when compared with normal and isograft tissues. On POD 11, both the clinical signs and histologic features of GVHD were seen, and TNF-α and IL-6 in native small bowel, TNF-α in colon, IFN-γ in spleen, and IL-6 in mesenteric lymph nodes were significantly increased in allograft animals when compared with that in normal and isograft tissues.Conclusions.In conclusion, TNF-α, IFN-γ, and IL-6 expression precede clinical onset and histologic evidence of GVHD in specific tissues. Therefore, increased expression of these cytokines is correlated with the development of GVHD in this model of SBT.