Fig. 1

GrB production and invasion in CRC cells. a (upper panel) GrB expression and EMT biomarkers (Snail 1, and E- and N-cadherin) were analyzed in CRC cell lines by WB on cell lysates; β-actin was used as loading control; a (lower panel) GrB secretion was investigated in the indicated cells pretreated with (+) or without (−) brefeldin A (BrfA), their supernatants were collected and analyzed by WB; β-actin was used as intracellular protein control and Ponceau staining as loading control. YT-S (NK leukemia) and T24 (bladder cancer) cells were used as positive and negative controls, respectively. b (left panel) The expression of colon stem cell markers such as Lgr5, Cd133 and CD166 and Bmi1 were analyzed in CSCs vs HT-29 cell lines by RT-PCR; GAPDH was used as loading control; b (right panel) GrB expression and secretion were analyzed in the indicated CSCs as in (a); c Invasion of CRC cells using the BioCoat Matrigel Invasion Chamber test; d HCT 116 cells were transfected with GrB-specific Stealth RNAi (siGrB) or Control Stealth RNAi (siCtr); GrB expression was verified by WB; β-actin was used as loading control; numbers indicate band intensities (b.i.) = band volume/area x mean pixel intensity, normalized for β-actin and quantified using Quantity One 1-D analysis software; invasion assay was performed using the BioCoat Matrigel Invasion Chambers test; bladder RT112 and pancreatic PT45 cancer cells served as positive controls; * p < 0.0001; **p < 0.001. Representative experiments out of at least three