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

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Abstract

Canstatin, a non-collagenous fragment, is cleaved from type IV collagen α2 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 μM, 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.

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Abbreviations

DMEM:

Dulbecco’s modified Eagle’s Medium

Drp1:

Dynamin related protein 1

NFAT:

Nuclear factor of activated T-cells

PI3K:

Phosphatidylinositol 3-kinase

ROS:

Reactive oxygen species

S.E.M.:

Standard error of the mean

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Acknowledgments

This research was supported by Kitasato University Research Grant for Young Researchers and JSPS KAKENHI Grant Number 24780289 (Grant-in-Aid for Young Scientists B).

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Authors and Affiliations

  1. Laboratory of Veterinary Pharmacology, School of Veterinary Medicine, Kitasato University, Higashi 23 Bancho 35-1, Towada, Aomori, 034-8628, Japan

    Muneyoshi Okada, Suiri Morioka, Hiroki Kanazawa & Hideyuki Yamawaki

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  1. Muneyoshi Okada
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  2. Suiri Morioka
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Correspondence to Muneyoshi Okada.

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Okada, M., Morioka, S., Kanazawa, H. et al. Canstatin inhibits isoproterenol-induced apoptosis through preserving mitochondrial morphology in differentiated H9c2 cardiomyoblasts. Apoptosis 21, 887–895 (2016). https://doi.org/10.1007/s10495-016-1262-1

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