title: Canstatin inhibits hypoxia-induced apoptosis through activation of integrin/focal adhesion kinase/Akt signaling pathway in H9c2 cardiomyoblasts.
abstract: A hypoxic stress which causes apoptosis of cardiomyocytes is the main problem in the ischemic heart disease. Canstatin, a non-collagenous fragment of type IV collagen α2 chain, is an endogenous anti-angiogenic factor. We have previously reported that canstatin has a cytoprotective effect on cardiomyoblasts. In the present study, we examined the effects of canstatin on hypoxia-induced apoptosis in H9c2 cardiomyoblasts. Cell counting assay was performed to determine a cell viability. Western blotting was performed to detect expression of cleaved casepase-3 and phosphorylation of focal adhesion kinase (FAK) and Akt. Immunocytochemical staining was performed to observe a distribution of αv integrin. Hypoxia (1% O2, 48 h) significantly decreased cell viability and increased cleaved caspase-3 expression. Canstatin (10-250 ng/ml) significantly inhibited these changes in a concentration-dependent manner. Cilengitide (1 μM), an αvβ3 and αvβ5 integrin inhibitor, significantly prevented the protective effects of canstatin on cell viability. Canstatin significantly increased phosphorylation of FAK and Akt under hypoxic condition, which were inhibited by cilengitide. LY294002, an inhibitor of phosphatidylinositol-3 kinase/Akt pathway, suppressed the canstatin-induced Akt phosphorylation and reversed the protective effects of canstatin. It was observed that hypoxia caused a localization of αv integrin to focal adhesion. In summary, we for the first time clarified that canstatin inhibits hypoxia-induced apoptosis via FAK and Akt pathways through activating integrins in H9c2 cardiomyoblasts.
authors.affiliation: Laboratory of Veterinary Pharmacology, School of Veterinary Medicine, Kitasato University, Towada, Aomori, Japan.
authors.forename: Hiroki
authors.initials: H
authors.lastname: Kanazawa
authors.affiliation: Laboratory of Veterinary Pharmacology, School of Veterinary Medicine, Kitasato University, Towada, Aomori, Japan.
authors.forename: Keisuke
authors.initials: K
authors.lastname: Imoto
authors.affiliation: Laboratory of Veterinary Pharmacology, School of Veterinary Medicine, Kitasato University, Towada, Aomori, Japan.
authors.forename: Muneyoshi
authors.identifier: http://orcid.org/0000-0002-0746-206X
authors.initials: M
authors.lastname: Okada
authors.affiliation: Laboratory of Veterinary Pharmacology, School of Veterinary Medicine, Kitasato University, Towada, Aomori, Japan.
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Yamawaki
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D015687:Cell Hypoxia; D002460:Cell Line; D024141:Collagen Type IV / Q000494:pharmacology*; D004353:Drug Evaluation, Preclinical; D051417:Focal Adhesion Kinase 1 / Q000378:metabolism; D016023:Integrins / Q000378:metabolism; D032386:Myoblasts, Cardiac / Q000187:drug effects / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D051381:Rats; D015398:Signal Transduction / Q000187:drug effects*
chemical_list: C502625:COL4A2 protein, human; D024141:Collagen Type IV; D016023:Integrins; D019869:Phosphatidylinositol 3-Kinases; D051417:Focal Adhesion Kinase 1; C496052:Ptk2 protein, rat; D051057:Proto-Oncogene Proteins c-akt