Fig 2: (A) Representative western blot of phenotypic changes observed in NCF stimulated with IL1ß. After 72 h IL1ß-stimulation (10 ng/mL) NCF myofibroblasts lose myofibroblastic markers (aSMA, Calponin, Synpo2) and overexpress activated CAF marker fibroblast activating protein (FAP). The bar graph below depicted mean (plus standard deviation) normalized densities (using ß-Actin as loading charge) for three independent western blot cell extracts, illustrating the decrease in myofibroblastic markers aSMA, Calponin, and Synaptopodin 2. (B) IL1ß (10 ng/mL) induced tumor cell proliferation in only two of the seven colorectal cancer cell lines. Bars depicted mean + sd of four independent experiments of three replicates each. (C) Conversely, IL1ß induced proliferation of NCFs in a 5-day WST-1 assay. These values could be restored by the addition of a P38 inhibitor (VX-702, 400 nM) and a neutralizing polyclonal antibody against IL1ß (2 µg/mL) (Kruskal–Wallis, Dunn’s multiple comparison test; bars depicted mean + sd of four independent experiments of three technical replicates each). Selecting the cell lines that responded to (10 ng/mL) IL1ß, we checked the dose–response effect of IL1ß on proliferation and survival against IC50 values for oxaliplatin (L-OHP), observing a dose–response trend only in HCT116 cells (D). IL1ß did not induce any protection against L-OHP. No effect was observed in HT29 cells (E). Both D and E represent mean values + sd of three independent experiments of six technical replicates each. Assaying the effect of IL1ß on fibroblast migration revealed a statistically significant increase in migration induced by the interleukin (F). We reported the same observation when analyzing directional migration of fibroblasts (H). This effect could be counterbalanced by the addition of a neutralizing polyclonal antibody against IL1ß (2 µg/mL). Bars displayed mean values + sd of three independent experiments. Conditioned media from IL1ß-stimulated NCFs induced the migration of tumor cells, both in a wound healing assay (G); white bar; bars displayed mean values + sd of three independent experiments; adj p value = 0.051) and directional migration in transwell, seeding NCFs in the bottom chamber (I); p = 0.0006, U Mann–Whitney test). (J) Representative western blot displaying that the blocking of IL1ß with a polyclonal neutralizing antibody anti-IL1ß (2 µg/mL) maintains the myofibroblastic phenotype in NCFs determined as the expression of aSMA and the decrease of FAP. The bar graph shows the mean + sd for three independent experiments not reaching statistical significance for FAP (p = 0.06 after adjusting for multiple comparison), but significant for aSMA (p = 0.021, after adjusting for multiple comparison; Kruskal–Wallis plus Dunn’s multiple comparison test).