In this study, we observed the level of normal murine hepatocyte inducible NOS (iNOS) mRNA by semi-quantitative polymerase chain reaction (SQ-PCR) analysis after stimulation with ES products (ESP) and/or ESP fractions from the plerocercoids. We found that ESP are able to induce the expression of iNOS gene in a dose-dependent fashion. Treatment of ESP with polymyxin B did not affect their ability to induce the expression of iNOS gene, suggesting that bacterial lipopolysaccharide (LPS) is not involved. The iNOS-inducing factor (a) is soluble, and may be a component whose molecular mass exceeds 94 kDa as analyzed by a combination of SDS-PAGE and SQ-PCR. The peak of iNOS mRNA level was detected 3 h after stimulation with ESP; the mRNA level decreased sharply from 9 h. Dexamethasone inhibited the induction of mRNA for hepatocyte iNOS. In contrast, cycloheximide stimulated the induction; this suggests that de nova protein synthesis is important in the regulation of the ESP-induced expression of iNOS mRNA. Actinomycin D blocked the induction. In addition, the results of Northern blot analysis showed that ESP suppressed the LPS (10 micrograms/ml) and interferon-gamma (IFN-gamma, 100 U/ml)-induced hepatocyte iNOS mRNA expression in a dose-dependent fashion and the suppressing effect was more marked when hepatocytes were exposed to ESP 3 h prior to LPS and IFN-gamma. These results demonstrate that the soluble factor(s) of ESP is capable of inducing murine iNOS gene expression in hepatocytes in the absence of added cytokines.
Animals Cells, Cultured