Involvement of Met receptor pathway in aggressive behavior of colorectal cancer cells induced by parathyroid hormone-related peptide

Figure 2 Parathyroid hormone-related peptide increases Met phosphorylation and activation through Src kinase in HCT116 cells. A: Cells were exposed to 10^-8 mol/L parathyroid hormone-related peptide (PTHrP) 10^-8 mol/L for 3 min, 5 min, and 10 min. The protein levels of Met and p-Met (phosphorylated in the residues Tyr1234 and Tyr1235) were assessed by Western blot to investigate whether PTHrP is capable of modulating Met phosphorylation and activation in HCT116 cells. Graph bars represent the average of the results obtained from three independent experiments; B: Cells were pre-treated with PP1, a selective Src inhibitor, for 30 min and then exposed to 10^-8 mol/L PTHrP for 3 min to evaluate whether Src kinase mediates the effect of PTHrP on Met activation in HCT116 cells. Controls were run by adding an equivalent volume of DMSO, the vehicle of the inhibitor. The protein levels of p-Met and Met were evaluated by Western blot. Graph bars represent the average of the results obtained from three independent experiments. In all experiments, GAPDH protein levels were determined as a control of the amount of proteins present in the membrane, since this protein is not substantially modified with the treatment by the cytokine. DMSO: Dimethylsulfoxide; GAPDH: Glyceraldehyde 3-phosphate dehydrogenase; Met: Receptor tyrosine kinase Met; p-Me: Phospho-Met (Tyr1234/1235); PTHrP: Parathyroid hormone-related peptide. ^aP < 0.05; ^bP < 0.01.