Figure 4

Effect of endothelial protein C receptor (EPCR) blockage on the expression/activation of mitogen-activated protein (MAP) kinase ERK, p38, and JNK in rheumatoid synovial fibroblasts (RASFs). RASFs were transfected with control (Con) or EPCR small interfering RNA (siRNA) for 24 hours. Medium was then replaced with Dulbecco’s modified Eagle’s medium (DMEM) without fetal bovine serum (FBS) and treated with tumor necrosis factor-alpha (TNF-α) (100 ng/mL) for a further 24 hours. Cells were collected and lysed for Western blot analysis and semi-quantified by image analysis software to measure ERK and P-ERK (A), p38 and P-p38 (B), and JNK and P-JNK (C); β-actin was included as an internal standard. The gel images represent one of three experiments. Data on graphs are shown as mean ± standard deviation (SD) of three different RASF cell lines and were analyzed by analysis of variance followed by Tukey’s honestly significant difference (HSD) post hoc test. **P <0.01. Relative comparisons are indicated by horizontal lines.