Ethanol disrupts hepatic function with the eventual appearance of alcoholic liver disease. Activation of Kupffer cells by bacterial endotoxins is a critical step in the etiology of fibrosis. The investigators have found that Kupffer cells isolated from rats fed ethanol in their diet for 4 weeks are hyper-responsive to stimulation by lipopolysaccharide, a component of bacterial cell walls, and phagocytosis, secreting a greater amount of the inflammatory cytokine, tumor necrosis factor alpha (TNFalpha), in response to stimulation. This response is dependent on activity of protein kinase C. Exposure of Kupffer cells to LPS also elicits an inhibitory feedback signal mediated by cAMP. In Kupffer cells from ethanol-fed rats, hormone-stimulated cAMP production is profoundly decreased. This desensitization is associated with a decrease in the stimulatory guanine nucleotide regulatory protein, G alpha S. Despite this decrease in cAMP production, inhibition of TNF production by hormones is normal, indicating that there are adaptations in the cAMP signal transduction cascade distal to cAMP production that potentiate cAMP-mediated responses in Kupffer cells from ethanol-fed rats. Here the investigators will determine the mechanisms and functional consequences of this potentiation of cAMP-mediated responses in isolated Kupffer cells and the perfused liver in situ in both male and female rats after chronic ethanol exposure. First, they will ascertain whether additional cAMP- dependent responses are maintained after ethanol feeding. Finally, the investigators will also determine whether the potentiated cAMP signal inhibits LPS-mediated responses by rapidly terminating the LPS signal at the plasma membrane, preventing the transduction of LPS signals from cytosol to nucleus and/or inhibition of LPS- activated gene transcription. Importantly, since the anti- inflammatory cascade mediated by cAMP remains functional after ethanol exposure, activation of cAMP production may provide a potential therapeutic strategy for minimizing the damaging consequences of endotoxin exposure associated with chronic ethanol consumption.
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