Fig. 1.

Carcinoma cells harboring mutp53 exert a non-cell-autonomous effect over macrophages. a Primary human monocytes were grown and differentiated towards three different lineages of macrophages (M0, M1, and M2) and co-cultured with an isogenic set of HCT116 cells differing by their p53 status (+/+ = WT p53, −/− = p53 null, mut = mutp53, p.R248W). RNA was extracted and subjected to qPCR analysis with primers specific to TNF-α and IL-10. Values were normalized for GAPDH mRNA in the same sample. b Primary monocytes were grown as in a and co-cultured with HT29 (mutp53- R273H) cells that underwent stable shRNA knock-down using scrambled oligo (ShCon) or specific to p53 (Shp53). TNF-α and IL-10 were analyzed as in a. c Co-cultured macrophages were seeded onto a cy-3-gelatin covered glass slide for 48 h and gelatin degradation rates were measured. d Co-cultured macrophages were harvested, stained with fluorescent antibodies against CD163 and CD206 and analyzed by flow cytometry. Relative intensities were compared with isotype controls. e Co-cultured macrophages were seeded in an eight-well chamber slide and incubated with fluorescent zymosan particles for 24 h, after which zymosan phagocytosis was evaluated. f,g Co-cultured macrophages were harvested and reseeded in an electrical-impedance monitoring chamber for 5 days to measure either migration (f) or invasion (g) properties. All experiments in this figure were repeated three times, error bars represent standard errors