Mechanisms of RAW264.7 macrophages immunomodulation mediated by polysaccharide from mung bean skin based on RNA-seq analysis

In this study, the immunomodulation properties and the possible molecular mechanisms of mung bean skin polysaccharide (MBP) on RAW264.7 were investigated. The results showed that MBP could enhance the phagocytic activity, promote intracellular reactive oxygen species (ROS) production as well as the release of nitric oxide (NO) and cytokines in macrophages. The results of global screening of all transcripts by RNA-seq revealed 927 differentially expressed genes between the control and MBP-treated groups, including 196 up-regulated genes and 731 down-regulated genes. From GO analysis, there were 5 cellular components terms (CC), 5 biological processes terms (BP) and 10 molecular functions terms (MF). Toll-like receptor 4 (TLR4 receptor), mitogen-activated protein kinases (MAPKs) and nuclear factor-κB (NF-κB) signaling pathways were selected by KEGG analysis to elucidate the mechanism of MBP action on macrophages. Western blot analysis showed that MBP could increase the expression of TLR4, MAPKs and NF-κB signaling pathway-related proteins, which confirmed that TLR4, MAPKs and NF-κB signaling pathways were essential to MBP-mediated mocrophage activation. Moreover, the production of both NO and cytokine TNF-α was found to be significantly reduced after the addition of TLR4, MAPKs and NF-κB inhibitors. These results suggest that MBP can control the immune response of macrophages through TLR4-mediated MAPKs and NF-κB signaling pathways. This work concluded MBP could serve as a potential natural immunomodulator, providing a molecular basis for its application in functional foods.