Identification of Cardiac Myosin-binding Protein C as a Candidate Biomarker of Myocardial Infarction by Proteomics Analysis

Abstract

Acute myocardial infarction (AMI) is a common cause of death for which effective treatments are available provided that diagnosis is rapid. The current diagnostic gold standards are circulating cardiac troponins I and T. However, their slow release delays diagnosis, and their persistence limits their utility in the identification of reinfarction. The aim was to identify candidate biomarkers of AMI. Isolated mouse hearts were perfused with oxygenated protein-free buffer, and coronary effluent was collected after ischemia or during matched normoxic perfusion. Effluents were analyzed using proteomics approaches based on one-or two-dimensional initial separation. Of the 459 proteins identified after ischemia with one-dimensional separation, 320 were not detected in the control coronary effluent. Among these were all classic existing biomarkers of AMI. We also identified the cardiac isoform of myosin-binding protein C in its full-length form and as a 40-kDa degradation product. This protein was not detected in the other murine organs examined, increased markedly with even trivial myocardial infarction, and could be detected in the plasma after myocardial infarction in vivo, a profile compatible with a biomarker of AMI. Two-dimensional fluorescence DIGE of ischemic and control coronary effluents identified more than 200 asymmetric spots verified by swapping dyes. Once again existing biomarkers of injury were confirmed as well as posttranslational modifications of antioxidant proteins such as peroxiredoxins. Perfusing hearts with protein-free buffers provides a platform of graded ischemic injury that allows detailed analysis of protein release and identification of candidate cardiac biomarkers like myosin-binding protein C.

Proteomic mapping of proteins released during necrosis and apoptosis from cultured neonatal cardiac myocytes

“…Our data are in line with prior studies examining proteins released into the coronary effluent following ischemia-reperfusion in isolated mouse and rabbit hearts (8,14). In addition to classical necrotic markers such as LDH, creatine kinase, and troponins, the rabbit heart study also identified Crsp3, a LIM domain-containing protein, as being released following ischemia-reperfusion injury (8).…”

“…We also found Crsp3 in our proteomic analyses, and it appeared to increase, albeit modestly, during necrosis but not apoptosis. The mouse study identified 320 proteins as being upregulated in coronary effluent following 5 min of ischemia (14). Analysis of the data indicated a considerable degree of overlap in the proteins identified.…”

“…Proteins included the classical markers LDH, creatine kinase, FABP3, and troponin proteins but also novel candidates such as cysteine and glycine-rich protein 3 (Crsp3) and myosin-binding protein C (MyBP-C). A similar study identified 320 proteins released following myocardial infarction, especially MyBP-C and oxidative stressrelated proteins such as peroxiredoxins (14).…”

Quantitative Mass Spectrometry-Based Approaches in Cardiovascular Research

“…Jacquet et al identified biomarkers of myocardial infarction by DIGE approach. 12 Meng et al applied 2D-DIGE in combination with matrix-assisted laser desorption/ionization (MALDI)-time of flight/time of flight MS to identify alterations in mitochondrial enzymes that contribute to cardiac hypertrophy in spontaneously hypertensive rats. 13 A combined proteomic and metabolomic analysis has been carried out in murine hearts to obtain mechanistic insights of protein kinase C epsilon (PKCϵ)-mediated cardioprotection.…”

Introducing a series of topical special issues of the Journal of Muscle Research and Cell Motility

“…The potential mis-regulation of MyBP-C-linked kinase signalling in heart disease is likely an underestimated factor, as discussed here (Bardswell et al 2012;Knöll 2012;Kuster et al 2012). While N-terminal fragments due to premature stop-codons do not accumulate detectably in the myocardium of HCM patients, ischaemia-reperfusion injury leads to the proteolytic cleavage of cardiac MyBP-C and the liberation of full-length protein and at least two proteolytic fragments (Jacquet et al 2009;Govindan et al 2012a) that show, in agreement with other studies on N-terminal fragments, tight localisation to the A-band (Govindan et al 2012b). This suggests that such proteolytic MyBP-C fragments might exert ectopic effects on myosin regulation that could contribute to contractile failure in the reperfused heart.…”