Vicenin-2 Exerts Bidirectional Immunomodulation via p38MAPK Phosphorylation in RAW264.7 Macrophages

ABSTRACT

Background: Inflammatory diseases, including autoimmune disorders, and chronic inflammatory conditions, are driven by dysregulated immune responses, particularly involving macrophage activation. Macrophages play a critical role in modulating inflammation through the production of pro-inflammatory mediators such as nitric oxide (NO), reactive oxygen species (ROS), tumor necrosis factor-alpha (TNF-α), and interleukin-6 (IL-6). Vicenin-2 (VCN-2), a flavonoid with potential immunomodulatory properties, has garnered attention for its therapeutic potential. This study investigates the bidirectional immunomodulatory effects of VCN-2 on lipopolysaccharide (LPS)-induced RAW264.7 mouse macrophage cells to establish a foundation for its development as a novel anti-inflammatory agent. Objectives: This study investigates the bidirectional immunomodulatory effects of vicenin-2 (VCN-2) on lipopolysaccharide (LPS)-induced mouse macrophage RAW264.7 cells, aiming to establish a basis for developing VCN-2 as a novel anti-inflammatory agent in diseases such as skin inflammation, autoimmune disorders, and chronic inflammatory conditions. Materials and Methods: LPS-induced mouse macrophage RAW264.7 cells were used as an inflammation model. The CCK-8 method was used to detect the activity and toxicity of VCN-2. The neutral red assay was used to measure the phagocytic ability of RAW264.7 cells. The Griess assay was used to detect nitric oxide (NO) levels. Flow cytometry was used to measure reactive oxygen species (ROS) levels. Enzyme-linked immunosorbent assay (ELISA) was used to measure tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) levels. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) was used to detect the expression of inducible nitric oxide synthase (iNOS), TNF-α, and IL-6 messenger ribonucleic acid (mRNA). Western blot (WB) was used to detect the expression of phosphorylated p38MARK (p-p38). Results: VCN-2 activates RAW264.7 cells and promotes cell proliferation. As the concentration of VCN-2 increases, its proliferative effect increases. In the resting state, VCN-2 improves cell proliferation by increasing cell number and cell volume. In the inflammation model, VCN-2 repairs cell morphology and restores cell proliferation. VCN-2 promotes phagocytosis of resting cells while attenuating phagocytic activity in inflammatory models. VCN-2 increases ROS levels in resting cells and reduces ROS levels in LPS-induced cells. VCN-2 increases NO, IL-6, and TNF-α content by promoting iNOS, IL-6, and TNF-α gene transcription levels, regulating macrophage immune function. In the inflammation model, VCN-2 significantly downregulates the transcription levels of iNOS, IL-6, and TNF-α genes, reduces NO, IL-6, and TNF-α content, and alleviates the inflammatory response. VCN-2 upregulates macrophage p38 in the resting state. In terms of phosphorylation expression, VCN-2 downregulates the phosphorylation expression of p38 in inflammatory model cells. Conclusion: VCN-2 enhances RAW264.7 cell proliferation, improves cell morphology, and amplifies phagocytic capacity. VCN-2 modulates the transcription of iNOS, IL-6, and TNF-α mRNA through the dual regulation of p38 phosphorylation, influencing the secretion of NO, IL-6, TNF-α, and ROS in RAW264.7 cells under varying conditions. This study demonstrates that VCN-2 exerts a bidirectional regulatory effect on macrophage immune function, supporting its potential as a therapeutic agent for inflammatory and immune-related diseases.

KEYWORDS

• Vicenin-2
• macrophage RAW264.7
• cytokines
• immune function
• p38MAPK

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• DOI10.1177/09731296251360569Copy

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