Acute Ethanol Binge Followed by Withdrawal Regulates Production of Reactive Oxygen Species and Cytokine-Induced Neutrophil Chemoattractant and Liver Injury During Reperfusion After Hepatic Ischemia

Abstract

This work tests the hypothesis that withdrawal from an acute ethanol binge regulates the production of reactive oxygen species (ROS) and chemokines by Kupffer cells, and as a result compromises or protects the liver from injury. Male Sprague-Dawley rats received an intravenous ethanol bolus (1.75 g/kg), followed by an intravenous infusion of 200-300 mg/kg/h for 12 h. At 12 h, ethanol infusion was stopped and replaced by saline. At 18 h, rats were subjected to 45 min of partial hepatic ischemia, followed by 0-24 h of reperfusion (I/R). At specific time points, Kupffer cells were isolated for superoxide anion assay and CINC (cytokine-induced neutrophil chemoattractant) and MIP-2 (macrophage inflammatory protein-2) production in vitro. Alanine transferase (ALT) activity, endotoxin, CINC, and MIP-2 were measured in serum samples taken at appropriate intervals. Results show that at 3 h post reperfusion, serum ALT was significantly elevated in the ethanol-treated group + I/R, compared with the saline + I/R group. ROS production by Kupffer cells at this time was also significantly increased compared with the saline + I/R group. However, ethanol withdrawal + I/R did not significantly alter CINC and MIP-2 production at 3 h of reperfusion. After 24 h, serum ALT was lower in the ethanol + I/R group than in the saline + I/R group. Superoxide anion and MIP-2 releases by Kupffer cells were not statistically significantly different between these two groups at this time. CINC production by Kupffer cells from the ethanol-treated + I/R group was significantly lower than in the saline + I/R group. Concomitantly, CINC and nuclear factor-κB (NF-κB) mRNAs and NF-κB translocation and binding in Kupffer cells in this treatment group were down-regulated. Moreover, the number of polymorphonuclear neutrophils (PMNs) sequestered in the liver was significantly lower in the ethanol + I/R group than in the saline-treated group. ROS and chemokine productions in sham animals with or without ethanol were lower than in the I/R group. These data suggest that acute ethanol binge followed by withdrawal may compromise the liver to injury during the early phase, whereas in the later phase it may be protective. Furthermore, these results support the notion that Kupffer cells are involved in hepatic injury in the early phase, whereas PMNs participate more actively during the later phase of reperfusion.