Plasmid Construct, RNA Interface and Transfection

ROCK1 plasmid (pCAG-myc-ROCK1^myc-727 Δ3, ROCK1 CA) was constructed as previously reported (Ishizaki et al., 1997). The siRNA sequence against ROCK1 (sense 5′-GGCAGAGGAAGAAUAUAAATT-3′; antisense 5′-UUUAUAUUCUUCCUCUGCCTT-3′) and scrambled siRNA was synthesized by GenePharma (Shanghai, China). Cells were transfected with ROCK1 plasmid, ROCK1 siRNA or scrambled siRNA using Lipofectamine 2000 reagent according to the manufacturer’s instructions. At 72 h post-transfection, cells and supernatant were harvested and then processed for western blot analysis.