M2 Macrophages Mediate the Resistance of Gastric Adenocarcinoma Cells to 5-Fluorouracil through the Expression of Integrin β3, Focal Adhesion Kinase, and Cofilin

Figure 3

(a) Cell viability of AGS cells treated with 5FU with conditioned media from polarized macrophages. AGS cells were plated in 96-well plates and incubated for 24 h at 37°C. After 24 h of incubation, the media were replaced with a mixture of fresh media and conditioned media (CM) from macrophages to a final ratio of 60 : 40, respectively (control = no CM, M0 = CM from PMA-differentiated THP-1 cells, M1 = CM from THP-1 cells activated with LPS, and M2 = CM of THP-1 cells activated with IL-4). 5FU was simultaneously added to the cell to a final concentration of 10 μg/ml and the plates were incubated for 24 h. After treatment, the media in the plate was removed and replaced with fresh media, and then 10 μl of WST-1 solution was added in each well, and the plate was incubated for further 4 h at 37°C. Absorbance was obtained using the ELISA microplate reader at 490 nm wavelength. (b) Proliferation of AGS cells cocultured with M2 macrophages. AGS cells were plated in 6-well plates and incubated for 24 h at 37°C. After 24 h of incubation, AGS cells were treated with 10 μg/ml of 5FU, and then polarized THP-1-M2 macrophages cultured in 6-well plate inserts of 0.4 μm pore size were added to the AGS wells and incubated for additional 24 h. After treatment, proteins were extracted from AGS cells and separated by Western blot. (c) Quantification of western blot bands. Statistically significant difference was set at ^#, , .

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