Induction of inducible nitric oxide synthase expression by 18beta-glycyrrhetinic acid in macrophages

Glycyrrhizin (GL), a triterpenoid saponin fraction of licorice, is reported to have anti-viral and anti-tumor activities and is metabolized to 18beta-glycyrrhetinic acid (GA) in the intestine by intestinal bacteria. However, the mechanism underlying its effects is poorly understood. To further elucidate the mechanism of GA, the aglycone of GL, we investigated the effects of GA on the release of nitric oxide (NO) and at the level of inducible NO synthase (iNOS) gene expression in mouse macrophages. We found that GA elicited a dose-dependent increase in NO production and in the level of iNOS mRNA. Since iNOS transcription has been shown to be under the control of the transcription factor nuclear factor kappaB (NF-kappaB), the effects of GA on NF-kappaB activation were examined. Transient expression assays with NF-kappaB binding sites linked to the luciferase gene revealed that the increased level of iNOS mRNA, induced by GA, was mediated by the NF-kappaB transcription factor complex. By using DNA fragments containing the NF-kappaB binding sequence, GA was shown to activate the protein/DNA binding of NF-kappaB to its cognate site, as measured by electrophoretic mobility shift assay. These results demonstrate that GA stimulates NO production and is able to up-regulate iNOS expression through NF-kappaB transactivation in macrophages.