NF-κB activation and modulation in hepatic macrophages during cholestatic injury

Cholestatic liver injury induces an inflammatory response that follows the activation of hepatic macrophages. Constitutive activation of the transcription factor, NF-κB, was found in these macrophages over the course of hepatic injury. Since NF-κB activation has been shown to have a key role in the inflammatory process, the modulatory effects of the antioxidant, α- tocopherol succinate, and the glucocorticoid, dexamethasone, on NF-κB activation were examined in this study. Male Sprague Dawley rats underwent 2- 7 days of common bile duct division and ligation (CBDL) or sham laparotomy. Hepatic macrophages were isolated by collagenase Pronase perfusion and purified by centrifugal elutriation. Activation was determined by electrophoretic mobility shift assay and ELISA. We determined that NF-KB activation in injured hepatic macrophages could only be inhibited by dexamethasone. Dexamethasone-mediated inhibition of NF-κB activation required the synthesis of a regulatory protein since cycloheximide-treated cells were resistant to its effects. Furthermore, dexamethasone-treated hepatic macrophages showed elevated steady-state levels of IκB-α mRNA, suggesting the role of IκB-α as a potential regulatory mediator. Consistent with constitutive transcriptional activation we showed constitutive secretion of TNF-α from injured hepatic macro-phages which could be inhibited by dexamethasone. These data show for the first time, in a biologically significant model of hepatic injury, constitutive activation of the key inflammatory transcription factor NF-κB and cytokine TNF-α. These results support an approach focused on the NF-κB/IκB-α pathway as a critical target for therapeutic intervention during hepatic injury, and the consideration of possible steroid-based therapies.