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.
Highlights
Ischemic heart disease such as myocardial infarction is one of the leading causes for death throughout the world [1,2,3]
We examined the effects of canstatin on hypoxia-induced morphological changes in H9c2 cardiomyoblasts
We found that hypoxia (48 h) significantly decreased cell viability (p
Summary
Ischemic heart disease such as myocardial infarction is one of the leading causes for death throughout the world [1,2,3]. Canstatin inhibits hypoxia-induced apoptosis apoptosis in cardiomyocytes is thought to be an important therapeutic strategy in the treatment of ischemic heart disease. It has been reported that cell surface expression of αvβ and αvβ integrins is increased by hypoxia stimulation in tumor cells [22,23]. Β3 integrin prevents oxidative stress-induced apoptosis in HL-1 mouse cardiomyocyte cell line [24]. It has been reported that αvβ and αvβ integrins activated the survival signaling pathway through the activation of Akt [25,26,27] which was responsible for the protection against ischemia-reperfusion injury in mouse cardiomyocytes [28]. We tested the hypothesis that canstatin affects hypoxia-induced apoptosis of cardiomyoblasts through the integrins/Akt pathways
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