We discovered new biomarker candidates for acute myocardial infarction (AMI) using micro LC and accurate mass spectrometry. Two serum samples were pooled and used for biomarker screening. UHPLC‐LTQ‐Orbitrap analysis was used with data‐dependent acquisition followed by Sequest search. Six biomarker candidates were selected after fold change analysis of peptides in AMI patient samples compared to healthy controls. Literature search results were reflected. The six biomarker candidates were characterized and quantified using multi reaction monitoring using synthesized peptides or trypsin digested protein standards. Levels of obscurin, alpha‐1‐acid glycoprotein 1, and clusterin were significantly increased (p < 0.001) in AMI patient samples compared to levels in healthy controls, while those of antithrombin III and fibrinogen beta were not. We confirmed the diagnostic performance of obscurin and clusterin as biomarker candidates for AMI using commercial ELISA kits. Receiver operating characteristic results showed that obscurin and clusterin showed highest sensitivity and specificity for AMI diagnosis together with troponins by both of multi‐reaction monitoring analysis and ELISA.

Acute myocardial infarction (AMI) is a multifaceted syndrome influenced by the functions of various extrinsic and intrinsic pathways and pathological processes, which can be detected in circulation using biomarkers. In this study, we investigated the secretome protein profile of induced-hypertrophy cardiomyocytes to identify next-generation biomarkers for AMI diagnosis and management. Hypertrophy was successfully induced in immortalized human cardiomyocytes (T0445) by 200 nM ET-1 and 1 µM Ang II. The protein profiles of hypertrophied cardiomyocyte secretomes were analyzed by nano-liquid chromatography with tandem mass spectrometry and differentially expressed proteins that have been identified by Ingenuity Pathway Analysis. The levels of 32 proteins increased significantly (>1.4 fold), whereas 17 proteins (<0.5 fold) showed a rapid decrease in expression. Proteomic analysis showed significant upregulation of six 14-3-3 protein isoforms in hypertrophied cardiomyocytes compared to those in control cells. Multi-reaction monitoring results of human plasma samples showed that 14-3-3 protein-zeta levels were significantly elevated in patients with AMI compared to those of healthy controls. These findings elucidated the role of 14-3-3 protein-zeta in cardiac hypertrophy and cardiovascular disorders and demonstrated its potential as a novel biomarker and therapeutic strategy.