Fig. 5. Wnt ligands are secreted by tumor cells cause the Wnt/β-catenin activation in macrophages leading to M2 macrophage polarization.

a BMDMs (M0) were stimulated with LPS + IFN-γ (M1) or IL-4 (M2). The expression of Wnt ligands was determined by qRT-PCR, using β-actin as an internal control. A quantitative comparison among M0, M1, and M2 macrophages was presented (n = 5). b The expression of the indicated Wnt ligands in KCs, TAMs, and Hepa1-6 cells was determined by qRT-PCR, and compared after calibration of their β-actin levels (n = 3). c BMDMs were cultured with the conditional medium (CM) that was collected from cultured Hepa1-6 cells for 24 h. Meanwhile BMDMs were treated with or without ICG001. The M2-related markers and downstream genes of Wnt signaling were detected by western blotting (left panel). The relative protein levels were quantitatively compared (right panel) (n = 4). d BMDMs were co-cultured with the conditional medium from Hepa1-6 cells that were infected with shRNA targeting the Wntless by lentivirus delivery system for 24 h. The M2-related markers and downstream genes of Wnt signaling were detected by western blotting (left panel). The relative protein levels were quantitatively compared (right panel) (n = 3). Bars, mean ± SD; *P < 0.05; **P < 0.01; ***P < 0.001