Carnosol, an Antioxidant in Rosemary, Suppresses Inducible Nitric Oxide Synthase through down-Regulating Nuclear Factor-Kappa B in Mouse Macrophages

Carnosol is a naturally occurring phytopolyphenol found in rosemary. Carnosol functions as antioxidant and anticarcinogen. In the present study , we compared the antioxidant activity of carnosol and other compounds extracted from rosemary. Carnosol showed potent antioxidative activity in alpha,alpha-diphenyl-beta- picrylhydrazyl (DPPH) free radicals scavenge and DNA protection from Fenton reaction. High concentrations of nitric oxide (NO) are produced by inducible NO synthase ( iNOS) in inflammation and multiple stages of carcinogenesis. Treatment of mouse macrophage RAW 264.7 cell line with carnosol markly reduced lipopolysaccharide (LPS)- stimulated NO production in a concentration-related manner with an IC50 of 9.4 muM; but other tested compounds had slight effects. Western blot, reverse transcription-polymerase chain reaction, and northern blot analyses demonstrated that carnosol decreased LPS-induced iNOS mRNA and protein expression. Carnosol treatment showed reduction of nuclear factor- kappaB (NF-kappaB) subunits translocation and NF- kappaB DNA binding activity in activated macrophages. Carnosol also showed inhibition of iNOS and NF-kappaB promoter activity in transient transfection assay. These activities were referred to down-regulation of inhibitor kappaB (IkappaB) kinase (IKK) activity by carnosol (5 muM), thus inhibited LPS-induced phosphorylation as well as degradation of IkappaBalpha. Carnosol also inhibited LPS- induced p38 and p44/42 mitogen-activated protein kinase ( MAPK) activation at a higher concentration (20 muM). These results suggest that carnosol suppresses the NO production and iNOS gene expression by inhibiting NF-kappaB activation, and provide possible mechanisms for its anti-inflammatory and chemopreventive action