Effect of Platycodon grandiflorus Polysaccharide on M1 Polarization Induced by Autophagy Degradation of SOCS1/2 Proteins in 3D4/21 Cells

M1-polarized macrophages can improve the body's immune function. This study aimed to explore the mechanism of Platycodon grandiflorus polysaccharide (PGPS(t)) degrading SOCS1/2 protein through autophagy and promoting M1 polarization in 3D4/21 cells. Immunoprecipitation, confocal laser scanning microscopy, flow cytometry, and intracellular co-localization were used to detect the expression of related phenotypic proteins and cytokines in M1-polarized cells. The results showed that PGPS(t) significantly promoted the mRNA expression of IL-6, IL-12, and TNF-alpha and enhanced the protein expression of IL-6, IL-12, TNF-alpha, IL-1 beta, iNOS, CD80, and CD86, indicating that PGPS(t) promoted M1 polarization in 3D4/21 cells. Next, the effect of the PGPS(t) autophagy degradation of SOCS1/2 on the M1 polarization of 3D4/21 cells was detected. The results showed that PGPS(t) significantly downregulated the expression level of SOCS1/2 protein, but had no obvious effect on the mRNA expression level of SOCS1/2, indicating that PGPS(t) degraded SOCS1/2 protein by activating the lysosome system. Further research found that under the action of 3-MA and BafA1, PGPS(t) upregulated LC3B II and downregulated SOCS1/2 protein expression, which increased the possibility of LC3B, the key component of autophagy, bridging this connection and degrading SOCS1/2. The interaction between SOCS1/2 and LC3 was identified by indirect immunofluorescence and Co-IP. The results showed that the co-localization percentage of the two proteins increased significantly after PGPS(t) treatment, and LC3 interacted with SOCS1 and SOCS2. This provides a theoretical basis for the application of PGPS(t) in the treatment or improvement of diseases related to macrophage polarization by regulating the autophagy level.