Canstatin inhibits isoproterenol-induced apoptosis through preserving mitochondrial morphology in differentiated H9c2 cardiomyoblasts

Canstatin, a non-collagenous fragment, is cleaved from type IV collagen alpha 2 chain, an essential component of basement membrane surrounding cardiomyocytes. Although canstatin is known as an endogenous anti-angiogenic factor, its effects on cardiomyocytes have not been clarified. This study examined the effects of canstatin on isoproterenol-induced apoptosis in differentiated H9c2 cardiomyoblasts. Retinoic acid was used to differentiate H9c2 myoblast to cardiomyocyte-like phenotype. Cell viability was determined by a cell counting assay. Western blotting was performed to detect expression of cleaved casepase-3 and phosphorylation of dynamin related protein (Drp) 1 at Ser637 which regulates mitochondrial fission. Mito Sox Red staining was performed to examine a mitochondria-dependent production of reactive oxygen species (ROS). Mitochondrial morphology was detected by Mito Tracker Red staining. Isoproterenol (100 mu M, 48 h) significantly decreased cell viability and increased cleaved caspase-3 expression, which were inhibited by canstatin (10-250 ng/ml) in a concentration-dependent manner. Canstatin suppressed the isoproterenol-induced mitochondrial fission but not ROS. Canstatin also inhibited the isoproterenol-induced dephosphorylation of Drp1 at Ser637. In conclusion, canstatin inhibits isoproterenol-induced apoptosis through the inhibition of mitochondrial fission via the suppression of dephosphorylation of Drp1 at Ser637 in differentiated H9c2 cardiomyoblasts.