• Hepatocytes;
  • Kupffer Cells;
  • Sinusoidal Endothelial Cells;
  • Tumor Necrosis Factor-α;
  • Binding;
  • Lipopolysaccharide

Tumor necrosis factor-α (TNF-α) has been shown to contribute to the alcohol [ethanol (ETOH)]-induced alteration of hepatic function. Therefore we tested the hypothesis that the hepatic action of TNF-α could be due, at least in part, to alterations in TNF-α cell-surface receptors of hepatic parenchymal (hepatocytes) and nonparenchymal (Kupffer and sinusoidal endothelial) cells. Rats were either acutely treated with ETOH by a primed, continuous 7-hr intravenous infusion of 20% (w/v) ETOH (30 mg/100 g body weight/h) or chronically fed an ETOH-containing liquid diet (5.2% ETOH, w/v, with ETOH as 36% of total calories) for 14 weeks. Control rats in the acute group were infused with sterile saline, whereas control rats in the chronic group were fed liquid diet containing dextrin to replace ETOH in isocaloric amounts. Three hr before killing, the rats were injected intravenously with Gram-negative bacterial lipopolysaccharide [(LPS) 100 /μg/100 g body weight] or saline. Hepatocytes, Kupffer cells, and sinusoidal endothelial cells were isolated after liver perfusion with collagenase (without pronase), separated by centrifugal elutriation, and used to determine the affinity (Kd) and capacity (Bmax) of binding sites, using recombinant human-[125l]TNF-α as the ligand. Two binding sites were detected on Kupffer cells and sinusoidal endothelial cells isolated from control animals: a high-affinity (Kd1, in the range of 150–200 PM), low-capacity (Bmax1, in the range of 2–3 fmol/106 cells) binding site and a low-affinity (Kd2, in the range of 2–9 nm), high-capacity (Bmax2, in the range of 3–15 fmol/106 cells) binding site. One binding site was detected on hepatocytes isolated from control rats (Kd1= 1.0 nm and a Bmax= 95 fmol/106 cells). Acute ETOH administration caused an increase in Kd1 on both Kupffer and sinusoidal endothelial cells; a decrease in Kd1 on hepatocytes; and an increase in Kd2, Bmax1, and Bmax2 on endothelial cells. A second binding site on hepatocytes (Kd2= 5.8 nm, Smax2= 186 fmol/106 cells) was observed only in the ETOH-treated group after LPS administration. Chronic alcohol exposure markedly elevated Kd1 and Smax1 on hepatocytes. Overall, LPS-induced changes mimicked those induced by alcohol, except for a decrease in Bmax1 on Kupffer cells of rats in the acute treatment group. Also, the liquid diet containing alcohol or dextrin abrogated the high-affinity, low-capacity binding sites on both Kupffer cells and sinusoidal endothelial cells, and low-affinity, high-capacity binding sites on the hepatocyte. After LPS administration, the high-affinity binding sites on Kupffer and sinusoidal endothelial cells were reexpressed. These data show that: (1) ETOH induces changes in both the capacity (Bmax) and affinity (Kd) of TNF-α cell-surface receptors of hepatocytes, Kupffer cells, and sinusoidal endothelial cells; and (2) ETOH-induced changes are consistent with an increased sensitivity of these cells to the action of TNF-α.