Chronic ethanol (EtOH) abuse results in the development of steatosis, alcoholic hepatitis, and cirrhosis. Augmented TNF-α production by macrophages and Kupffer cells and signaling via the p55 TNF receptor have been shown to be critical for these effects of chronic EtOH; however, the molecular mechanisms leading to augmented TNF-α production remain unclear. Using cell culture models and in vivo studies we demonstrate that chronic EtOH results in increased TNF-α transcription, which is independent of NF-κB. Using reporter assays and chromatin immunoprecipitation we found that this increased transcription is due to increased IRF-3 binding to and transactivation of the TNF promoter. As IRF-3 is downstream from the TLR4 adaptor TIR-domain-containing adapter-inducing IFN-β (Trif), we demonstrate that macrophages from Trif−/− mice are resistant to this dysregulation of TNF-α transcription by EtOH in vitro as well as EtOH-induced steatosis and TNF dysregulation in vivo. These data demonstrate that the Trif/IRF-3 pathway is a target to ameliorate liver dysfunction associated with chronic EtOH.

The metabolic sensor AMP-activated kinase (AMPK) inhibits both the cystic fibrosis (CF) transmembrane conductance regulator (CFTR) Cl -channel and epithelial Na 1 channel (ENaC), and may inhibit secretion of proinflammatory cytokines in epithelia. Here we have tested in primary polarized CF and non-CF human bronchial epithelial (HBE) cells the effects of AMPK activators, metformin and 5-aminoimidazole-4-carboxamide-1-b-D-riboside (AICAR), on various parameters that contribute to CF lung disease: ENaC-dependent short-circuit currents (I sc ), airway surface liquid (ASL) height, and proinflammatory cytokine secretion. AMPK activation after overnight treatment with either metformin (2-5 mM) or AICAR (1 mM) substantially inhibited ENaC-dependent I sc in both CF and non-CF airway cultures. Live-cell confocal images acquired 60 minutes after apical addition of Texas Red-dextran-containing fluid revealed significantly greater ASL heights after AICAR and metformin treatment relative to controls, suggesting that AMPK-dependent ENaC inhibition slows apical fluid reabsorption. Both metformin and AICAR decreased secretion of various proinflammatory cytokines, both with and without prior LPS stimulation. Finally, prolonged exposure to more physiologically relevant concentrations of metformin (0.03-1 mM) inhibited ENaC currents and decreased proinflammatory cytokine levels in CF HBE cells in a dose-dependent manner. These findings suggest that novel therapies to activate AMPK in the CF airway may be beneficial by blunting excessive sodium and ASL absorption and by reducing excessive airway inflammation, which are major contributors to CF lung disease.Keywords: metformin; cystic fibrosis transmembrane conductance regulator; ENaC; airway surface liquid; inflammationThe serious genetic disease cystic fibrosis (CF) is caused by mutations in the CF transmembrane conductance regulator (CFTR) Cl -channel expressed at the apical membrane in a variety of epithelial tissues, including the lung, where CF lung disease causes considerable morbidity and mortality (1). Defective CFTR function prevents airway epithelium from adequately regulating the airway surface liquid (ASL) volume, which has an adverse effect on mucus clearance from the lung. The CF airway invariably becomes colonized by bacteria such as Pseudomonas aeruginosa and develops excessive inflammation, which can cause extensive damage to lung architecture and eventuate in respiratory failure (2). There is growing evidence that a lack of functional CFTR promotes an exaggerated or prolonged inflammatory response to bacterial and other insults, although the underlying pathophysiologic mechanisms are unclear (3-5). Both up-regulation of proinflammatory mediators (e.g., IL-8, IL-6, TNF-a, and GM-CSF) and down-regulation of anti-inflammatory mediators (e.g., IL-10 and inducible nitric oxide synthase) in the CF airway may play an important role in this process (6-8).Excessive activity of the epithelial sodium channel (ENaC) on the luminal airway surface is a key factor involved in th...

To determine whether early acute cellular rejection (ACR) is associated with sub-optimal immunosuppression in children with liver transplants (LTx). Methods: Twenty-five children with primary LTx after pre-transplant rabbit anti-thymocyte globulin (rATG), and steroid-free tacrolimus (TAC) were evaluated.

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