Abstract: Background/Aims: Tumor necrosis factor-α (TNF-α) is believed to play a role in hepatic allograft rejection. However, the specific cellular population responsible for TNF-α production during hepatic allograft rejection is not known. Circulating monocyte-macrophage cells are the primary systemic sources of TNF-α. In the liver, Kupffer cells are the main producers of TNF-α. In this study, we determined which cells are involved in TNF-α production during allograft rejection after orthotopic liver transplantation. Methods: In situ hybridization was used to identify cells with TNF-α mRNA in the liver. Immunohistochemical staining with ED2 and ED3 was used to differentiate between cellular types (Kupffer cells versus infiltrating monocytes). To detect DNA fragmentation in liver cells, TdT-mediated biotin-dUTP nick-end labeling (TUNEL) was done. Studies were performed in the rat liver transplant model using rejecting (ACI to LEW) and non-rejecting (ACI to ACI) donor/recipient combinations. Results: In the control group, cells with TNF-α mRNA were rarely observed. In the rejection group, TNF-α mRNA was observed in mononuclear cells that were mainly within the vessels of the portal region and occasionally in the sinusoids. The cells with the signals for TNF-α mRNA were ED2-negative and ED3-positive. DNA fragementation was observed in hepatocytes as well as infiltrating mononuclear cells. Conclusions: The main producer of TNF-α may be infiltrating mononuclear cells such as monocyte-macrophage cells rather than Kupffer cells during allograft rejection after liver transplantation. Circulating monocyte-macrophages may play a role in the control of allograft rejection.