Inhibition of MMP-2 expression affects metabolic enzyme expression levels: Proteomic analysis of rat cardiomyocytes Han-Bin Lin a , Keshav Sharma a , Dariusz Bialy b , Magdalena Wawrzynska c , Randy Purves d , Francisco S. Cayabyab e , Mieczyslaw Wozniak a, f , Grzegorz Sawicki a, f , ⁎ a Department of Pharmacology, University of Saskatchewan, Saskatoon, Saskatchewan, Canada b Department and Clinic of Cardiology, Medical University of Wroclaw, Wroclaw, Poland c Department of Medical Emergency, Medical University of Wroclaw, Wroclaw, Poland d National Research Council of Canada, Saskatoon, Saskatchewan, Canada e Department of Physiology, University of Saskatchewan, Saskatoon, Saskatchewan, Canada f Department of Clinical Chemistry, Medical University of Wroclaw, Wroclaw, Poland ARTICLE INFO ABSTRACT Article history: Received 6 January 2014 Accepted 13 April 2014 In this study we examined the effect of inhibition of MMP-2 expression, using siRNA, on the cardiomyocyte proteome. Isolated cardiomyocytes were transfected with MMP-2 siRNA and incubated for 24 h. Control cardiomyocytes from the same heart were transfected with scrambled siRNA following the same protocol. Comparison of control cardiomyocyte proteomes with proteomes from MMP-2 suppressed cardiomyocytes revealed 13 protein spots of interest (9 protein spots increased; 4 decreased). Seven protein spots were identified as mitochondrial enzymes involved in energy production and represent: ATP synthase beta subunit, dihydrolipoyllysine-residue succinyltransferase component of 2-oxoglutarate dehydrogenase complex, cytochrome c oxidase subunit 5A, electron transfer flavoprotein subunit beta, NADH dehydrogenase (ubiquinone) 1 alpha subcomplex subunit 5 and a fragment of mitochondrial precursor of long-chain specific acyl-CoA dehydrogenase. Furthermore, precursor of heat shock protein 60 and Cu–Zn superoxide dismutase were identified. Two protein spots corresponding to MLC1 were also detected. In addition, ATP synthase activity was measured and was increased by approximately 30%. Together, these results indicate that MMP-2 inhibition represents a novel cardioprotective therapy by promoting alterations in the levels of mitochondrial enzymes for improved energy metabolism and by preventing degradation of contractile proteins needed for normal excitation–contraction coupling. Biological significance During ischemia and reperfusion of cardiomyocytes, abnormality in excitation–contraction coupling and decreased energy metabolism often lead to myocardial infarction, but the cellular mechanisms are not fully elucidated. We show for the first time that intracellular Keywords: Matrix metalloproteinase-2 ATP synthase Cardiomyocytes siRNA Metabolic enzymes Contractility JOURNAL OF PROTEOMICS 106 (2014) 74 – 85 ⁎ Corresponding author at: Department of Pharmacology, College of Medicine, University of Saskatchewan, 107 Wiggins Road, Saskatoon, Saskatchewan S7N 5E5, Canada. Tel.: +1 306 966 6997. E-mail address: greg.sawicki@usask.ca (G. Sawicki). http://dx.doi.org/10.1016/j.jprot.2014.04.026 1874-3919/© 2014 Elsevier B.V. All rights reserved. Available online at www.sciencedirect.com ScienceDirect www.elsevier.com/locate/jprot