Inhibition of Inducible Nitric Oxide Synthase Gene Expression and Enzyme Activity by Epigallocatechin Gallate, a Natural Product from Green Tea

Abstract

Chronic inflammation has been implicated as the underlying factor in the pathogenesis of many disorders. In the past decade, inflammation-related endogenous production of reactive nitrogen species, similar to oxygen free radicals, has also been suggested as a risk factor for cancer, in addition to the well-studied exogenous nitroso compounds. Epidemiological, in vitro, and animal model studies have implicated green tea to be protective against nitroso compound-induced and inflammation-related cancer. Therefore, we investigated the effect of epigallocatechin-3-gallate (EGCG), one of the known biologically active catechins contained in green tea, on the production of nitric oxide (NO ·). We have shown previously that EGCG reduces NO · production as measured by nitrite accumulation in the culture medium. Expanding on this finding, in this report we show that EGCG may do so by two mechanisms: reduction of inducible nitric oxide synthase (iNOS) gene expression and inhibition of enzyme activity. Addition of 1–10 μM EGCG to lipopolysaccharide- and interferon-γ-activated mouse peritoneal cells reduced iNOS mRNA expression concentration dependently, to 82–14%, as measured by relative reverse transcription–polymerase chain reaction. Addition of 50–750 μM EGCG, in a concentration-dependent manner, inhibited the enzyme activity of iNOS, to 85–14%, and neuronal nitric oxide synthase (nNOS), to 93–56%, as measured by citrulline formation. EGCG competitively inhibited binding of arginine and tetrahydrobiopterin, and the gallate structure is important for this action.