Fig. 3

SMC18 increases macrophage phagocytosis of MC38 cells in vitro. A Representative Western blotting of SIRPα phosphorylation in the co-culture system of bone marrow-derived macrophages (BMDMs) with MC38 cells treated with PBS, SMC18 (50 μM), and anti-mouse CD47 (clone miap301, 20 µg/mL). Miap301 was set as a positive control and PBS as a negative control. IP (immunoprecipitation), IB (immunoblotting), and anti-pY (anti-phosphotyrosine). B Confocal images of bone marrow-derived macrophages from mice were labeled with DiR red dye and cultured with EGFP-labeled MC38 tumor cells. The cells were incubated with PBS, SMC18(50 μM) or miap301 (20 µg/mL). Representative images are shown. Arrows pointed to phagocytosed tumor cells. C Effects of SMC18 on phagocytosis of macrophages. Phagocytosis assays were conducted using EGFP-labeled tumor cells and mouse bone marrow-derived macrophages at a ratio of 1:4. SMC18 was used at a concentration of 50 μM. Co-cultured GFP⁺MC38 cells were assessed for the percentage of GFP⁺ macrophages using flow cytometry. The phagocytosis rate was calculated using the formula (GFP⁺ and F4/80⁺ cells)/F4/80⁺ cells. n = 3. The data are presented as means ± S.E.M. Statistical significance was determined using the unpaired Student's t-test (*P < 0.05, **P < 0.01, ***P < 0.001)