Protective Effect of Black Rice-Derived Cyanidin-3-O-β-Glucoside on Lipopolysaccharide-Induced Inflammatory Injury in Human Monocyte Cell Line THP-1

Monocyte-mediated inflammatory responses play a key role in the progression of atherosclerosis. Anthocyanins are polyphenolic flavonoids that are abundant in various dark-colored vegetables, fruits and cereals, which possess a wide range of biological activities. Cyanidin-3-O-β-glucoside (Cy-3-g) is an important anthocyanin monomer. The current study aimed to explore the effects of black rice-derived Cy-3-g on lipopolysaccharide (LPS)-induced inflammatory responses in Tohoku hospital pediatrics-1 (THP-1) as well as its possible mechanism of action. Crudeanthocyanins from black rice were obtained by extraction with ethanol-hydrochloric acid solution and macroporous resin adsorption and purified by medium-pressure liquid chromatography with UV detection. Cy-3-g with a purity greater than 96.5% was obtained. THP-1 monocytes were sequentially incubated with different concentrations (0, 0.1, 0.25 and 0.5 μg/mL) Cy-3-g for 4 h, and LPS (50 ng/mL) for additional 48 h. The levels of interleukin (IL)-1β, IL-6, IL-8, IL-10 and tumor necrosis factor (TNF)-α in the culture were determined by enzyme-linked immunosorbent assay (ELISA), and the mRNA expression levels of the genes encoding IL-1β, IL-6, IL-8, IL-10, TNF-α, Toll-like receptor 4 (TLR4) and nuclear factor kappa B (NF-κB) p65 were measured by quantitative polymerase chain reaction (PCR). Moreover, the protein expression levels of TLR4, NF-κB p65 and inhibitor of NF-κB (IκBα) were detected by Western blot. The LPS-induced increase in the relative mRNA expression levels and release levels of IL-1β, IL-6 and TNF-α in THP-1 cells was significantly inhibited by all concentrations of Cy-3-g tested (P < 0.05, P < 0.01 and P < 0.001, respectively). Cy-3-g at 0.25 and 0.50 μg/mL significantly blunted the inhibitory effect of LPS on IL-10 mRNA expression (P < 0.01 and P < 0.001, respectively). Cy-3-g at 0.10, 0.25 and 0.50 μg/mL significantly mitigated the inhibitory effect of LPS on IL-10 release (P < 0.05, P < 0.01 and P < 0.001, respectively), but had no significant influence on LPS-induced promotion of IL-8 mRNA expression and release levels (P > 0.05). Cy-3-g at 0.25 and 0.50 μg/mL significantly inhibited the mRNA and protein expression of TLR4 and NF-κB p65 (P < 0.05, P < 0.01 and P < 0.001, respectively), suppressed the expression of phosphorylated IκBα and NF-κB, and inhibited LPS-induced IκBα degradation. Bay11-7082, a specific inhibitor of NF-κB, significantly decreased the release of IL-1β, IL-6 and TNF-α in LPS-stimulated THP-1 cells (P < 0.001), but had no synergistic effects with 0.50 μg/mL Cy-3-g (P > 0.05). Black rice-derived Cy-3-g exerts a protective effect on LPS-induced inflammatory injury of THP-1 monocytes possibly through down-regulating the TLR4/NF-κB signaling pathway.