title: Ischemia and reperfusion induce differential expression of calpastatin and its homologue high molecular weight calmodulin-binding protein in murine cardiomyocytes.
abstract: In the heart, calpastatin (Calp) and its homologue high molecular weight calmodulin-binding protein (HMWCaMBP) regulate calpains (Calpn) by inhibition. A rise in intracellular myocardial Ca2+ during cardiac ischemia activates Calpn thereby causing damage to myocardial proteins, which leads to myocyte death and consequently to loss of myocardial structure and function. The present study aims to elucidate expression of Calp and HMWCaMBP with respect to Calpn during induced ischemia and reperfusion in primary murine cardiomyocyte cultures. Ischemia and subsequently reperfusion was induced in ∼ 80% confluent cultures of neonatal murine cardiomyocytes (NMCC). Flow cytometric analysis (FACS) has been used for analyzing protein expression concurrently with viability. Confocal fluorescent microscopy was used to observe protein localization. We observed that ischemia induces increased expression of Calp, HMWCaMBP and Calpn. Calpn expressing NMCC on co-expressing Calp survived ischemic induction compared to NMCC co-expressing HMWCaMBP. Similarly, living cells expressed Calp in contrast to dead cells which expressed HMWCaMBP following reperfusion. A significant difference in the expression of Calp and its homologue HMWCaMBP was observed in localization studies during ischemia. The current study adds to the existing knowledge that HMWCaMBP could be a putative isoform of Calp. NMCC on co-expressing Calp and Calpn-1 survived ischemic and reperfusion inductions compared to NMCC co-expressing HMWCaMBP and Calpn-1. A significant difference in expression of Calp and HMWCaMBP was observed in localization studies during ischemia.
authors.affiliation: Department of Pathology and Laboratory Medicine, College of Medicine, University of Saskatchewan, Saskatoon, Saskatchewan, Canada.
authors.forename: Sreejit
authors.initials: S
authors.lastname: Parameswaran
authors.affiliation: Department of Pathology and Laboratory Medicine, College of Medicine, University of Saskatchewan, Saskatoon, Saskatchewan, Canada.
authors.forename: Rajendra K
authors.initials: RK
authors.lastname: Sharma
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D015153:Blotting, Western; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D002148:Calmodulin-Binding Proteins / Q000378:metabolism*; D002154:Calpain / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002478:Cells, Cultured; D005260:Female; D005434:Flow Cytometry; D051379:Mice; D018613:Microscopy, Confocal; D008970:Molecular Weight; D017202:Myocardial Ischemia / Q000503:physiopathology*; D032383:Myocytes, Cardiac / Q000166:cytology / Q000378:metabolism*; D011485:Protein Binding; D020033:Protein Isoforms; D015424:Reperfusion
chemical_list: D002135:Calcium-Binding Proteins; D002148:Calmodulin-Binding Proteins; D020033:Protein Isoforms; C033668:calpastatin; D002154:Calpain