The M3 macrophage phenotype increases the efficiency of phagocytosis, the first stage of antigen cross-presentation
Abstract
Background. The M3 phenotype of macrophages, which, in contrast to the M1 and M2 phenotypes, produces proinflammatory cytokines (PI-M3) in response to the anti-inflammatory factors and exerts a pronounced in vivo anti-tumor effect. This study hypothesized that the more pronounced anti-tumor effect of PI-M3 macrophages compared to other phenotypes is based on a greater ability of the PI-M3 phenotype to phagocytize pathogenic cells and, thus, to perform better antigen cross-presentation. This makes the PI-M3 phenotype a desirable candidate for development of new macrophage-based cancer immunotherapy.
The aim of the study was to test this hypothesis.
Methods. Mouse macrophages isolated from peritoneal lavage were used in this study. The M1 phenotype was obtained by culturing macrophages in the DMEM/F12 medium with IFN-γ. The M2 phenotype was obtained by culturing macrophages in the DMEM/F12 medium supplemented with 20-40% FBS. The PI-M3 macrophage phenotype was obtained by adding IFN-γ and STAT3, STAT6, and SMAD3 inhibitors to the culture medium followed by stimulation with lipopolysaccharide (LPS). After the end of macrophage stimulation with LPS, the phagocytic ability of various phenotypes was assessed. The assessment was performed using the pHrodo® Red Staphylococcus aureus Bioparticles stained bacteria kit (Invitrogen).
Results. The phagocytic activity of the PI-M3 phenotype assessed as the number of phagocytic cells and the pattern and index of phagocytic activity, practically did not differ from the activity of the M1 phenotype but was significantly higher than that of the M2 phenotype under normal conditions. Culturing macrophages in a tumor environment reduced the phagocytic activity of all phenotypes but, at the same time, revealed significant stability of the phagocytic activity of the PI-M3 phenotype compared to other phenotypes.
Conclusion. This study demonstrated a high capability of PI-M3 macrophages for phagocytosis. This result is consistent with the hypothesis that the more pronounced anti-tumor effect of PI-M3 macrophages may be based on the greater functional ability of the PI-M3 phenotype to phagocytize the antigen.
