Figure 3

PAR-1 knockdown attenuates laminar flow-induced actin stress fiber formation in ECs. (A) HUVECs transfected with control RNA or PAR-1 siRNA were exposed to laminar flow. Representative photomicrographs showing F-actin with Texas Red-X Phalloidin and nucleus with DAPI (original magnification ×200). (B) Quantification of actin stress fiber (ASF) is presented by bar graph. (C) Cells transfected with control RNA or PAR-1 siRNA (100 pmol/L) for 2 days were exposed to laminar flow for 30 or 60 minutes. Protein levels of pFAK, PAR-1, and tubulin were determined by Western blotting against specific antibodies. (D) Cells transfected with control or PAR-1 siRNA (100 pmol/L) for 2 days were exposed to laminar flow for 10, 30 and 60 minutes. Protein levels of pSrc, pErk1/2, Erk1/2, PAR-1, and tubulin were assayed by Western blotting against specific antibodies. The results shown are representative of three independent experiments. Cells were used for experiments at passage 4, 5 and 6.