Abstract
Background and objective: Nitric oxide (NO) is known to exert cardioprotective effects against heart ischemic damage and may be involved in ischemic pre- and postconditioning. NO-triggered cardioprotective mechanisms are not well understood but may involve regulation of mitochondrial permeability transition pore (mPTP). In this study, we aimed to identify differentially phosphorylated mitochondrial proteins possibly involved in the NO/protein kinase G (PKG)/mPTP signaling pathway that can increase the resistance of cardiomyocytes to ischemic damage. Materials and methods: Isolated hearts from Wistar rats were perfused with NO donor NOC-18 prior to induction of stop–flow ischemia. To quantify and characterize the phosphoproteins, mitochondrial proteins were resolved and analyzed by two-dimensional gel electrophoresis followed by Pro-Q Diamond phosphoprotein gel staining, excision, trypsin digestions, and mass spectrometry. Quantitative proteomic analysis coupled with liquid chromatography–tandem mass spectrometry was also performed. Results: Mitochondrial protein phosphorylation patterns in NOC-18-pretreated ischemic hearts versus ischemic hearts were compared. Pretreatment of hearts with NOC-18 caused changes in mitochondrial phosphoproteome after ischemia which involved modifications of 10 mitochondrial membrane-bound and 10 matrix proteins. Among them, α-subunit of ATP synthase and adenine nucleotide (ADP/ATP) translocase 1, both of which are considered as potential structural components of mPTP, were identified. We also found that treatment of isolated non-ischemic mitochondria with recombinant PKG did not cause the same protein phosphorylation as pretreatment of hearts with NOC-18. Conclusions: Our study suggests that pretreatment of hearts with NOC-18 causes changes in mitochondrial phosphoproteome after ischemia which involves modifications of certain proteins thought to be involved in the regulation of mPTP opening and intracellular redox state. These proteins may be potential targets for pharmacological preconditioning of the heart.
Highlights
Ischemic heart pathologies including acute myocardial infarction which can result in heart failure are the leading cause of mortality and disability worldwide
To determine which mitochondrial proteins are phosphorylated after ischemia and pretreatment with Nitric oxide (NO) donor NOC-18, 2-DE gels were stained with Pro-Q Diamond for phosphoproteins and with
Further densitometric analysis of these spots revealed that only six protein spots from two experiments showed higher than 1.5-fold differences in relative spot volume between ischemic and ischemic plus NOC-18 mitochondrial samples
Summary
Ischemic heart pathologies including acute myocardial infarction which can result in heart failure are the leading cause of mortality and disability worldwide. It is generally accepted that mitochondria play a crucial role in the pathogenesis of multiple cardiac diseases including ischemic heart disease and myocardial infarction, mainly due to the imbalance of cellular bioenergetics (loss of ATP synthesis and increase of ATP hydrolysis), impairment in ionic homeostasis (intracellular Ca2+ in particular), formation of reactive oxygen species (ROS) and release of proapoptotic proteins—all being considered as key factors in the process of irreversible heart damage [1,2,3] Mitochondrial dysfunction impairs their communication with other cellular organelles leading to alteration in energy homeostasis that affects whole cellular metabolism and cause cell death. Conclusions: Our study suggests that pretreatment of hearts with NOC-18 causes changes in mitochondrial phosphoproteome after ischemia which involves modifications of certain proteins thought to be involved in the regulation of mPTP opening and intracellular redox state These proteins may be potential targets for pharmacological preconditioning of the heart
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
