Portal venous transfusions (PVTs) of blood have been shown to induce significant immunosuppression in animal models of organ transplantation. A proposed mechanism of PVT-induced immunosuppression is via alteration of Kupffer cell arachidonic acid metabolism with increased secretion of the suppressive metabolite prostaglandin E2(PGE2). This study assessed the hypothesis that PVT increases Kupffer cell PGE2production via up-regulation of Kupffer cell phospholipase A2(PLA2) as well as constitutive (COX1) and inducible (COX2) cyclooxygenase. Kupffer cells from Lewis rats were harvested 1 hr after PVT with either 1 ml of Wistar-Furth blood, systemic transfusion (SVT), or saline via portal vein (PVSal). After lipopolysaccharide stimulation, 24-hr Kupffer cell supernatant fractions were assayed for PGE2. PGE2was increased after SVT (1465±234 pg/ml) compared with PVSal (597±99;P<0.01). PVT increased Kupffer cell PGE2(5370±533;P<0.001 vs. SVT and vs. PVSal) even more substantially. Kupffer cells from PVT-treated rats were then cultured in the presence of inhibitors of PLA2, COX1, or COX2. When Kupffer cells were treated with mepacrine to inhibit PLA2(5575±453 pg/ml), PGE2production was not different from that by PVSal-treated controls (6467±614 pg/ml), but when Kupffer cells were incubated in the presence of the COX1 inhibitor flurbiprofen (3512±407 pg/ml) or the COX2 inhibitor nimesulide (2800±830 pg/ml), production was decreased 46.7% and 56.7%, respectively, over control activity without added inhibitor. PVT also increased Kupffer cells COX1 and COX2 mRNA as measured by Northern blot. Heart transplants were then performed from Wistar-Furth donors into Lewis recipients at the time of PVT, SVT, PVSal, or PVT + indomethacin (COX1/2 inhibitor). PVT prolonged allograft survival (12.0±0.9 days) compared with PVSal (6.3±0.3;P<0.01) or SVT (6.3±0.3;P<0.04). Indomethacin shortened graft survival when given with PVT (6.5±0.3 days). In summary, PVT increased Kupffer cell PGE2production, up-regulated transcription of Kupffer cells COX1 and COX2 mRNA, and prolonged cardiac allograft survival. COX1/2 inhibition abrogated the effect of PVT. The results indicated that the immunosuppressive effect of PVT may be mediated by up-regulation of Kupffer cell COX1 and COX2. Manipulation of Kupffer cell arachidonic acid metabolism may be useful in augmentation of PVT-induced immunosuppression.