Fig. 6
p-FoxM1.S25 functions in recruitment of macrophages and triggers polarization of M2-like TAM. a, b Monocyte THP-1 cells were co-cultured with A549 cells expressing mock, WT, S25A, and S25E FoxM1 for 48 h. In THP-1 cells, qRT-PCR was performed for markers of M1 (INOS, IL12B), M2 (IL10, CD163, CD206), and TAM (TGFB1, VEGFA) (a). In A549 cells, qRT-PCR was performed for IL4, IL6, IL10, VEGFA, and CD274 (b). *p < 0.05; **p < 0.01; ***p < 0.001; (n = 3). Data are presented as mean ± SD. c, THP-1 cells were co-cultured with A549 cells expressing mock, WT, S25A, and S25E FoxM1. The secreted levels of TGF-β1 and VEGFA from THP-1 cells co-cultured with A549 cells were detected using ELISA. d Monocyte THP-1 cells were co-cultured with A549 cells expressing mock or S25E depleted FoxM1 using shRNA for 48 h. Using THP-1 cells, qRT-PCR was performed for CD163, CD206, and VEGFA. *p < 0.05; **p < 0.01; ***p < 0.001; (n = 3). e, f Representative CD68 (pan-macrophage marker) staining (e, upper panel) and CD163 (TAM marker) staining (e, lower panel) were performed using lung tissue from mice. The relative density of CD68 staining (f, left panel) and CD163 staining (f, right panel) was analyzed and plotted. *p < 0.05; **p < 0.01; ***p < 0.001. Data are presented as mean ± SD. g Immunoblotting was performed using lung tissue lysates from each mouse model. FoxM1, CD68, CD163 and β-actin were detected using specific antibodies (upper panel). The relative protein intensities were analyzed and plotted (lower panel). h The viability of A549 cells expressing mock, WT, S25A, and S25E FoxM1 was measured when the cells were co-cultured with monocyte THP-1 cells. The ratio between A549 cells and THP-1 cells was 1:0, 1:2, 1:4, and 1:6, as indicated. i Monocyte THP-1 cells were co-cultured with A549 cells expressing mock, WT, S25A, and S25E FoxM1. qRT-PCR was performed for CD279 mRNA level in THP-1 cells. Data are presented as mean ± SD of at least three independent experiments (significantly different from the experimental control). **p < 0.01; ***p < 0.001; (n = 3)