Abstract:Objective To study the polarization regulation of compound Wufengcao liquid and one of its components, protocatechuic acid, on human macrophage cell line (classical activated U937 cell M1) induced by lipopolysaccharide (LPS). Methods U937 cells were induced with phorbol myristate acetate (PMA) for 24 hours to establish unactivated M0 cells; LPS was used to induce classical activated M1 cells for 48 hours. The control group (M0), model group (M1), compound Wufengcao liquid intervention group (CO-WFC-M1), and protocatechuic acid intervention group (PA-M1) were set up. Different concentrations of protocatechuic acid were used to intervene LPS-induced M1 macrophages, and CCK8 was used to detect the proliferation ability of M1 cells. Neutral red, nitrate reductase, enzyme-linked immunosorbent assay (ELISA), and RT-qPCR were used to detect the phagocytic ability of macrophages in each group and the levels of nitric oxide (NO), as well as the protein and mRNA expressions of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), IL-6, and inducible nitric oxide synthase (iNOS). Flow cytometry was used to detect the expression of CD80 and CD206 on the membrane surface of LPS-induced macrophages and protocatechuic acid-treated cells. Results Compared with the blank group (protocatechuic acid concentration of 0), protocatechuic acid at concentrations of 0.312-5 mmol/L had a significant inhibitory effect on U937-M1 cells (P＜0.05). The phagocytic rates of macrophages in the M0, M1, CO-WFC-M1, and PA-M1 groups were 0.48±0.28, 1.25±0.26, 2.01±0.18, and 1.80±0.32, respectively, with statistically significant differences between groups (F=166.029, P＜0.01). Compared with M0, the NO level in M1 significantly increased, and compared with M1, the NO level significantly decreased after intervention with compound Wufengcao liquid or protocatechuic acid (P＜0.05). In terms of protein and mRNA expression, compared with M0, the expression of inflammatory factors TNF-α, IL-1β, IL-6, iNOS in M1 significantly increased, and compared with M1, the expression of iNOS significantly decreased, arginase-1 (Arg-1) expression significantly increased, and the above inflammatory indicators significantly decreased after intervention with compound Wufengcao liquid or protocatechuic acid (P＜0.05). The positive rate of CD80 on the surface of M1 was 97.50%, and after intervention with protocatechuic acid, the positive rate of CD80 on the surface of M1 was 1.33%. Conclusion Both compound Wufengcao liquid and protocatechuic acid can regulate macrophage polarization, that is, inhibiting the MI macrophages that secretes pro-inflammatory factors and M2 macrophages that promote the secretion of growth factors, which may be the mechanism by which the compound Wufengcao liquid can promote wound healing, and this effect is related to one of its components, protocatechuic acid.