Lin, HY; Juan, SH; Shen, SC; Hsu, FL; Chen, YC
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
muM, dose-dependently stimulated the flavonoid (50 muM)-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 muM), 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. (C) 2003 Elsevier Inc. All rights reserved.