Inhibition of lipopolysaccharide-induced nitric oxide production by flavonoids in RAW264.7 macrophages involves heme oxygenase-1

Biochem Pharmacol. 2003 Nov 1;66(9):1821-32. doi: 10.1016/s0006-2952(03)00422-2.


The role of heme oxygenase-1 (HO-1) played in the inhibitory mechanism of flavonoids in lipopolysaccharide (LPS)-induced responses remained unresolved. In the present study, flavonoids, including 3-OH flavone, baicalein, kaempferol, and quercetin, induced HO-1 gene expression at the protein and mRNA levels in the presence or absence of LPS in RAW264.7 macrophages. This effect was associated with suppression of LPS-induced nitric oxide (NO) production and inducible nitric oxide synthase (iNOS) protein expression. Hemin induced HO-1 protein expression and this was associated with the suppression of LPS-induced NO production and iNOS protein expression in a dose-dependent manner. In addition, an increase in bilirubin production was found in flavonoid- and hemin-treated cells. Hemin, at the doses of 10, 20, and 50 microM, dose-dependently stimulated the flavonoid (50 microM)-induced HO-1 protein expression, and enhanced their inhibitory effects on LPS-induced NO production and iNOS protein expression. Pretreatment of the HO-1 inhibitor, tin protoporphyrin (10 microM), attenuated the inhibitory activities of the indicated flavonoids on LPS-induced NO production. Morphologic analysis showed that 3-OH flavone, baicalein, kaempferol, quercetin, hemin, and tin protoporphyrin did not cause any change in cell viability in the presence or absence of LPS. In contrast, only 3-OH flavone showed a significant inhibition of cell growth using the MTT assay. Transfection of an HO-1 vector in macrophages (HO-1/RAW264.7) resulted in a 3-fold increase in HO-1 protein compared with that the parental RAW264.7 cells. NO production mediated by LPS in HO-1 over-expressed RAW264.7 cells (HO-1/RAW264.7) was significant less than that in parental RAW264.7 cells. 3-OH Flavone, baicalein, kaempferol, and quercetin showed a more significant inhibition on LPS-induced NO production in HO-1/RAW264.7 cells than in parental RAW264.7 cells. These results provide evidence on the role of HO-1 in the inhibition of LPS-induced NO production by flavonoids. A combination of HO-1 inducers (i.e. hemin) and flavonoids might be an effective strategy for the suppression of LPS-induced NO production.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Line
  • Drug Interactions
  • Enzyme Induction / drug effects
  • Flavanones*
  • Flavones
  • Flavonoids / chemistry
  • Flavonoids / pharmacology*
  • Gene Expression / drug effects*
  • Heme Oxygenase (Decyclizing) / biosynthesis*
  • Heme Oxygenase (Decyclizing) / genetics
  • Heme Oxygenase-1
  • Hemin / pharmacology
  • Kaempferols / pharmacology
  • Lipopolysaccharides / pharmacology*
  • Macrophages / drug effects*
  • Macrophages / enzymology
  • Macrophages / metabolism
  • Membrane Proteins
  • Metalloporphyrins / pharmacology
  • Mice
  • Nitric Oxide / metabolism*
  • Nitric Oxide Synthase / metabolism
  • Nitric Oxide Synthase Type II
  • Protoporphyrins / pharmacology
  • Quercetin / pharmacology


  • Flavanones
  • Flavones
  • Flavonoids
  • Kaempferols
  • Lipopolysaccharides
  • Membrane Proteins
  • Metalloporphyrins
  • Protoporphyrins
  • tin protoporphyrin IX
  • Nitric Oxide
  • baicalein
  • kaempferol
  • Hemin
  • Quercetin
  • Nitric Oxide Synthase
  • Nitric Oxide Synthase Type II
  • Nos2 protein, mouse
  • Heme Oxygenase (Decyclizing)
  • Heme Oxygenase-1
  • Hmox1 protein, mouse
  • flavone