Protease-activated receptor 1 (PAR1) that can be activated by
serine proteinases such as
thrombin has been demonstrated to contribute to the development of cardiac remodeling and
hypertrophy after myocardial injury. Here, we investigated the mechanisms by which PAR1 leads to hypertrophic cardiomyocyte growth using cultured rat neonatal ventricular myocytes. PAR1 stimulation with
thrombin (1 U/ml) or a synthetic agonist
peptide (TFLLR-NH2, 50 μM) for 48 h induced an increase in cell size and myofibril formation associated with BNP (
brain natriuretic peptide) production. This actin reorganization assessed by
fluorescein isothiocyanate (
FITC)-conjugated
phalloidin staining appeared at 1 h after PAR1 stimulation, and this response was reduced by a
protein kinase C (PKC) inhibitor,
chelerythrine, inhibitors of Rho (
simvastatin) and
Rho-associated kinase (ROCK) (Y-27632), but not by
pertussis toxin (PTX). By Western blot analysis, translocation of PKCα or PKCε from the cytosol to membrane fractions was observed in cells stimulated with
thrombin or
TFLLR-NH2 for 2 - 5 min. In addition, PAR1 stimulation for 3 - 5 min increased the level of active RhoA. Furthermore, inhibitors of PKC and ROCK and Rho abrogated PAR1-mediated increase in cell size. Depletion of PKCα or PKCε by specific
small interfering RNA also suppressed both actin reorganization and cell growth. These results suggest that PAR1 stimulation of cardiomyocytes induces cell
hypertrophy with actin cytoskeletal reorganization through activation of PKCα and PKCε
isoforms and RhoA via PTX-insensitive
G proteins.