Figure 4. Efficiency of MKP-1 protein knock-down in HMCs using siRNA and antisense oligonucleotide approaches.

At 24 h after transfection with MKP-1 siRNA, or MKP-1 antisense oligonucleotides, cells were incubated with or without 20 ng/ml r-CTGF for 1 h. To control for non-specific effects of siRNA delivery, controls were included in which cells were either mock-transfected (transfection reagent only) or transfected with luciferase siRNA, whereas a CG-matched randomized sequence oligonucleotide was used as a control for the specificity of the antisense delivery. At the end of the incubation time, total RNA was extracted and used for semi-quantitative RT–PCR (A). In parallel experiments, cells were lysed and protein was subjected to SDS/PAGE and analysed by Western blotting using anti-MKP-1 antibody (B, C). Membranes were stripped and reprobed with an anti-β-actin antibody to control for sample loading. Results are representative of three separate experiments.