Previous studies have indicated that 14-3-3γ is upregulated by stress in LPS-induced cardiovascular injury. In this study, we investigated the interaction of 14-3-3γ and Bcl-2 family members in the control of the mitochondrial permeability transition (MPT) to test the hypothesis that abundant levels of 14-3-3γ can protect against LPS-induced injury via a Bcl-2 family/mitochondria pathway. The cardiomyocytes were treated with LPS (1mg l(-1)) for 6h; the interaction between 14-3-3γ and phospho-Bad(S112) was detected by co-immunoprecipitation (co-IP); the levels of Bcl-2 family members in the cytosolic and mitochondrial fractions were examined by Western blot; the apoptosis and mitochondrial membrane potential (ΔΨm) were detected by flow cytometry; and the mitochondrial permeability transition pore (mPTP) opening was tested by mitochondrial swelling. Our results revealed that LPS treatment results in cardiomyocyte injury, and these effects were significantly attenuated by pFLAG-14-3-3γ. Moreover, LPS treatment induced Bax translocation to the mitochondria, ΔΨm loss, mitochondrial swelling, and cytochrome c release, and pFLAG-14-3-3γ reversed these effects induced by LPS. Moreover, overexpressed 14-3-3γ protein could assist Bad(S112) phosphorylation and interact with it to form a complex, which might result in the disassociation of Bcl-2 from the Bad/Bcl-2 complex and its translocation from the cytosol to the mitochondria. Our data firstly confirmed that a high level of 14-3-3γ protects against LPS-induced cardiomyocyte injury likely through a pathway associated with the regulation of the subcellular localizations of Bcl-2 and Bad that results in the prevention of mPTP opening, the maintenance of ΔΨm, and ultimately the inhibition of apoptosis.
Keywords: 14-3-3γ; Apoptosis; Bcl-2 family member; Cardiomyocyte; Lipopolysaccharide; Mitochondrial permeability transition pore.
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