Watermelon Flower Petal-derived Apigenin Exhibits Anti-inflammatory Effects by Suppressing M1 Macrophage Polarization in Lipopolysaccharide/Interferon-gamma-co-stimulated RAW264.7 Cells

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Background Watermelon fruits contain various bioactive compounds. However, the bioactivity of watermelon flower petals, specifically in the context of the anti-inflammatory effects, remains unknown. Objectives In this study, we aimed to evaluate the effects of watermelon flower petal-derived components on nitric oxide (NO) production and elucidate the mechanisms underlying this process in murine macrophage RAW264.7 cells that were co-stimulated with lipopolysaccharide (LPS) and interferon-gamma (IFNγ). Materials and methods Phenolic compounds (specifically p-hydroxybenzoic acid, protocatechuic acid, apigenin, and rhoifolin) were first isolated from watermelon flower petals. NO production was assessed using a Griess reagent kit. The mRNA expression of Nos2, Cox2, Tnfα, Il6, and Cd80 was determined using quantitative real-time polymerase chain reaction. The protein expression levels of the following molecules were assessed using western blotting: inducible NO synthase (iNOS); cyclooxygenase-2; heme oxygenase-1; toll-like receptor 4; phosphorylated-extracellular signal-regulated kinase (p-ERK); ERK; p-p38; p38; p-Jun amino terminal kinase (p-JNK); JNK; p-nuclear factor-kappa B (p-NF-κB); NF-κB; p-signal transducer and activator of transcription 1 (p-STAT1); and STAT1. Furthermore, the nuclear translocation of NF-κB was observed using fluorescence microscopy. Results NO production was suppressed in LPS/IFNγ-co-stimulated RAW264.7 cells treated with apigenin. Additionally, iNOS mRNA and protein expression were reduced in LPS/IFNγ-co-stimulated RAW264.7 cells. Apigenin resulted in a slight reduction in the levels of phosphorylated p38, JNK, and STAT1; on the other hand, phosphorylation and nuclear translocation of NF-κB remaining unchanged in the LPS/IFNγ-co-stimulated RAW264.7 cells. mRNA expression of Cd80, a notable cell surface M1 macrophage marker, was suppressed by apigenin in LPS/IFNγ-co-stimulated RAW264.7 cells. Conclusion Apigenin was found to suppress M1 macrophage polarization in LPS/IFNγ-co-stimulated RAW264.7 cells. The study findings suggest that watermelon flower petals represent a promising and valuable source of anti-inflammatory agents.