Abstract: Purpose: The aim of this study was to evaluate the inhibitory effects of six flavonoids (luteolin, quercetin, myricetin, apigenin, kaempferol and chrysin) on M1 polarization of RAW264.7 macrophages and to delineate the structural determinants involved in their activity. Methods: Lipopolysaccharide (LPS) at 100 ng/mL was used to stimulate RAW264.7 macrophages to establish an inflammation model. Cell proliferation activity was detected by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide assay. The expression of CD274 and CD38 was detected by flow cytometry. The expression of proteins involved in the inflammatory signaling pathway was detected by Western blotting. The expression levels of M1-type macrophage marker genes were detected by real-time polymerase chain reaction (PCR). Results: All these flavonoids had no significant effect on cell proliferation at 20 μmol/L, but inhibited the expression of CD274 and CD38, with luteolin, apigenin and chrysin being more effective than quercetin, myricetin and kaempferol. The inhibitory activity of luteolin on activation of the nuclear factor κB signaling pathway was stronger than that of apigenin and quercetin, while the other flavonoids had no significant effect. The inhibitory effect of luteolin, apigenin and chrysin on the mRNA expression of inducible nitric oxide synthase (iNOS) was stronger than that of the other three compounds, and the inhibitory effect of luteolin, quercetin and apigenin on the mRNA expression of interleukin-1β (IL-1β) and monocyte chemotactic protein 1 (MCP1) was stronger than that of the other compounds. Conclusion: Hydroxylation at position 3 on the C ring is not conducive to the inhibition of flavonoids on macrophage polarization, while hydroxylation at positions 3’ and 4’ on the B ring are beneficial to enhance their activity.

Key words: flavonoids, macrophages, polarization, inflammation, structure-activity relationship