Fig. 3
M2 TAMs increase CSC properties in HCC cells. A After monoculture or coculture with TAMsCM or M2 MφsCyto for 72 h, HepG2 cells were collected and subjected to flow cytometry with double staining for CD44 and CD133. The bar graph depicts the percentages of CD44+/CD133+ HepG2 cells. *P < 0.05; ***P < 0.001 vs. monoculture. B Gene expression of the stem cell markers NANOG, SOX2, and OCT4 in HepG2 cells at 72 h after monoculture or coculture was determined by qPCR. The bar graphs depict the relative mRNA expression. **P < 0.01; ***P < 0.001 vs. monoculture. C After monoculture or coculture with TAMsCM or M2 MφsCyto for 72 h, HepG2 cells were cultured in ultra-low attachment surface dishes for 14 days to form spheres. The morphology of HepG2 spheres was observed with bright field microscopy (left lower). Magnification, 200 × , scale bar, 100 μm. The bar graph depicts the numbers of HepG2 spheres with a diameter greater than 100 μm in each field. D After coculture with TAMsCM or M2 MφsCyto for 72 h, HepG2 cells with shControl, shNANOG, shSOX2, or shOCT4 were treated with SOR (10 μM) for 48 h. Cell viability was assessed by MTT assay. The bar graphs depict the relative viability of HepG2 cells. **P < 0.01; ***P < 0.001 vs. shControl