Enhanced lnterleukin-8 Production by Cultured Chang Liver Cells Subjected to Ethanol Exposure and Subsequent Stimulation with Tumor Necrosis Factor-α

Abstract

We investigated the production of interleukin-8 (IL-8) and chemotactic activity released from Chang liver cells subjected to long-term treatment with ethanol (Et) and subsequent stimulation with tumor necrosis factor-α (TNF-α). Chang liver cells were cultured in the presence of 5, 50 or 100 mmol/l Et for 4 weeks and then treated with recombinant TNF-α (1, 10, 100 U/ml). The culture supernatants were assayed for IL-8 using a sandwich ELISA and chemotactic activity was measured using a chemotactic chamber. Total RNA was also extracted from these cells and IL-8 mRNA was assayed by RT-PCR. In addition, TNF-receptor expression on the Et-treated cells was analyzed by flow cytometry. IL-8 levels in supernatants of cells stimulated with 100 U/ml of TNF-α for 48 h rose significantly with increasing concentrations of Et and values obtained were as follows: 4,918 ± 244.4 pg/ml at 0 mmol/l Et, 5,335 ± 266.2 pg/ml at 5 mmol/l Et, 8,726 ± 873.4 pg/ml at 50 mmol/l Et and 9,134 ± 866.0 pg/ml at 100 mmol/l Et. The chemotactic activity also increased with increasing concentrations of Et and was almost completely suppressed by anti-IL-8 antibody. Using semiquantitative analysis of radioactivity of IL-8 mRNA using a ³²P γATP-labeled primer for IL-8 mRNA, Et-treated cells were shown to have markedly higher levels of radioactivity than untreated cells. In addition, TNF-receptor expression was significantly higher in cells treated with 100 mmol/l Et. These data suggest that long-term Et treatment of Chang liver cells stimulated with TNF-α may enhance transcription of the IL-8 gene with up-regulation of the TNF receptor.

Toxic effects caused by rhubarb (Rheum palmatum L.) are reversed on immature and aged rats

“…In our experiment, increased incidences of activated Kupffer cells and lymphocytic infiltration, indicating activation of inflammation course, were found at hepatic sinusoid in aged rats treated with high-dosage rhubarb. In addition, it was reported that IL-8 was activated in alcoholic hepatitis (Maher, 1995) and the transcription of IL-8 gene was enhanced with upregulation of the TNF receptor (Ohira et al, 1997). It was also found that the IL-8 levels were closely correlated with liver injury and could reflect the stage and severity in patients with alcoholic hep- a All survived animals were tested.…”

The role of chemokines in the immunopathology of the liver

“…Levels of IL-8 were found to be increased in alcoholic hepatitis (70)(71)(72) and correlated to increased ICAM-1 and vascular cell adhesion molecule-1 expression (73) and the infiltration of neutrophils (73)(74)(75). In vitro studies have shown that IL-8/CINC can be produced directly by ethanol-challenged hepatocytes (76,77) or as a result of cell to cell interactions with Kupffer cells. This was demonstrated when conditioned medium from ethanol-challenged hepatocytes was given to unchallenged Kupffer cells, which resulted in CINC secretion (78).…”