title: Altered expression of calcineurin, calpain, calpastatin and HMWCaMBP in cardiac cells following ischemia and reperfusion.
abstract: A rise in intracellular myocardial Ca(2+) during cardiac ischemia activates calpain (Calpn) thereby causing damage to myocardial proteins, which leads to myocyte death and consequently to loss of myocardial structure and function. Calcineurin (CaN) interacts with Calpn and causes cellular damage eventually leading to cell death. Calpastatin (Calp) and high molecular weight calmodulin-binding protein (HMWCaMBP) (homolog of Calp), inhibit Calpn activity and thus prevent cell death. CaN stimulation can also result in self-repair of damaged cardiomyocytes. The present study attempts to elucidate the expression of these proteins in cells under pre-ischemic condition (control), following ischemia induction and also reperfusion subsequent to ischemia. For the first time, flow cytometric analysis (FACS) has been used for analyzing protein expression concurrently with viability. We induced ischemia and subsequently reperfusion in 80% confluent cultures of neonatal murine cardiomyocytes (NMCC). Viability following induction was assessed with 7-AAD staining and the cells were simultaneously checked for protein expression by FACS. We observed that ischemia induction results in increased expression of CaN, Calp and Calpn. HMWCaMBP expression was reduced in live cells following ischemia which suggests that there is a poor survival outcome of cells expressing HMWCaMBP thereby making it a potential biomarker for such cells. Most live cells following ischemia expressed CaN pointing towards self-repair and favorable survival outcomes.
authors: Department of Pathology and Laboratory Medicine, Cancer Research Cluster, University of Saskatchewan, Room 4D40, 107 Wiggins Road, Saskatoon, Saskatchewan S7N 5E5, Canada.
authors: Sreejit
authors: S
authors: Parameswaran
authors: Department of Pathology and Laboratory Medicine, Cancer Research Cluster, University of Saskatchewan, Room 4D40, 107 Wiggins Road, Saskatoon, Saskatchewan S7N 5E5, Canada. Electronic address: rajendra.sharma@usask.ca.
authors: Rajendra K
authors: RK
authors: Sharma
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D019703:Calcineurin / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D002148:Calmodulin-Binding Proteins / Q000378:metabolism*; D002154:Calpain / Q000378:metabolism*; D002478:Cells, Cultured; D005786:Gene Expression Regulation; D051379:Mice; D009206:Myocardium / Q000378:metabolism*; D032383:Myocytes, Cardiac / Q000378:metabolism*; D015427:Reperfusion Injury / Q000378:metabolism* / Q000473:pathology
chemical_list: D002135:Calcium-Binding Proteins; D002148:Calmodulin-Binding Proteins; C033668:calpastatin; D019703:Calcineurin; D002154:Calpain
keywords: CaN; Calcineurin; Calp; Calpain; Calpastatin; Calpn; DPBS; FACS; FITC; HMWCaMBP; I/R; Ischemia; NDB; NMCC; PE; Primary cardiomyocyte culture; R-phycoerythrin; Reperfusion; calcineurin; calpain; calpastatin; dulbecco’s phosphate buffered saline; flow cytometry; fluorescein isothiocyanate; high molecular weight calmodulin-binding protein; ischemia and reperfusion; nutrient deficient buffer; primary neonatal mouse cardiomyocyte culture