Hydroxyfasudil hydrochloride is a ROCK inhibitor, with IC50s of 0.73 and 0.72 μM for ROCK1 and ROCK2, respectively. Hydroxyfasudil also less potently inhibits PKA, with an IC50 of 37 μM, 50-fold higher than those of the ROCKs. Hydroxyfasudil increases eNOS mRNA levels, with an EC50 value of 0.8 ± 0.3 μM. Hydroxyfasudil (0-100 μM) concentration-dependently increases eNOS activity and stimulates NO production in human aortic endothelial cells (HAEC). Hydroxyfasudil (10 μM) increases the half-life of eNOS mRNA from 13 to 16 hours, but does not affect eNOS promoter activity at concentrations from 0.1 to 100 μM.