Diabetes mellitus is frequently associated with coagulation disorders such as coronary heart disease and stroke. We aimed to clarify the molecular mechanism whereby hyperglycemia causes the procoagulant state. HuH7 human hepatocyte cells were treated with high glucose alone or in combination with proinflammatory cytokines, and the effects on the activity of the transcription factor nuclear factor kappa-B (NF-kappaB), which mediates the expression of acute-phase and coagulation-related genes, were examined. The results showed that increasing the medium glucose concentration from 3 to 24 mM significantly enhanced NF-kappaB-luciferase activity by 40% in the presence of insulin. The effect was promoter specific and not mimicked by comparable hyperosmolality with L-glucose. Proinflammatory cytokines such as interleukin-1 and tumor necrosis factor-alpha (TNF-alpha) also stimulated NF-kappaB-dependent transcription and showed an additive effect with high glucose. Similar effects were obtained on acute-phase or coagulation/fibrinolysis-related gene promoters such as fibrinogen or plasminogen activator inhibitor-1, all of which are shown to have NF-kappaB-mediated transcription. Finally, pretreatment of the cells with an antioxidant PDTC completely abolished the effect of high glucose and markedly attenuated that of TNF-alpha, suggesting the involvement of reactive oxygen species. These results suggest that (1) high glucose as well as proinflammatory cytokines have positive effects on NF-kappaB-mediated transcription in an additive manner and enhance coagulation-related gene expression and (2) the effects are mediated, at least partly, by the generation of oxidative stress and may be responsible for the high prevalence of thrombotic disorders in the metabolic syndrome with diabetes, hyperinsulinemia, obesity, and/or inflammation.