Knockdown of E-cad increased EGFR expression by enhancing its mRNA stability. (a) 24 hours after transfection of PCI-37A and 686LN cells with siRNA, cells were treated with actinomycin-D (Sigma-Aldrich) at 5 μg/ml for 0, 4, 8 and 24 hours. RT-PCR showed that the EGFR mRNA level remained higher in E-cad knockdown cells than in the control cells even after mRNA synthesis was stopped by actinomycin-D for 24 hours. (b) EGFR and GAPDH mRNA level as PCR products were measured using UVP BioImaging System Representative. The ratios of EGFR mRNA levels in E-cad knockdown cells (PCI-37A) against that in control cells at different time points were determined after being normalized to GAPDH. (c) Real-time RT-PCR on PCI-37A cells confirmed the ratios of EGFR mRNA level in E-cad knockdown cells against the control cells at different time points. These results indicate that EGFR mRNA stability was enhanced in E-cad knockdown cells. The experiments were repeated 3 times. Error bars represent standard deviation.