HEMOSTASIS, THROMBOSIS, AND VASCULAR BIOLOGY Regulation of endothelial thrombomodulin expression by inflammatory cytokines is mediated by activation of nuclear factor-kappa B

Inflammation and thrombosis are increasingly recognized as interrelated biologic processes. Endothelial cell expression of thrombomodulin (TM), a key component of the anticoagulant protein C pathway, is potently inhibited by inflammatory cytokines. Because the mechanism underlying this effect is largely unknown, we investigated a potential role for the inflammatory transcription factor nuclear factor-kappa B (NFB). Blocking NFB activation effectively prevented cytokineinduced down-regulation of TM, both in vitro and in a mouse model of tumor necrosis factor(TNF)–mediated lung injury. Although the TM promoter lacks a classic NFB consensus site, it does contain tandem Ets transcription factor binding sites previously shown to be important for both constitutive TM gene expression and cytokine-induced repression. Using electrophoretic mobility shift assay and chromatin immunoprecipitation, we found that multiple Ets species bind to the TNFresponse element within the TM promoter. Although cytokine exposure did not alter Ets factor binding, it did reduce binding of p300, a coactivator required by Ets for full transcriptional activity. Overexpression of p300 also prevented TM repression by cytokines. We conclude that NFB is a critical mediator of TM repression by cytokines. Further evidence suggests a mechanism involving competition by NFB for limited pools of the transcriptional coactivator p300 necessary for TM gene expression. (Blood. 2005;105:3910-3917)