title: Metformin influences cardiomyocyte cell death by pathways that are dependent and independent of caspase-3.
abstract: OBJECTIVE
  Metformin has been shown to increase fatty acid oxidation, an effect mediated by AMP activated protein kinase (AMPK). We hypothesised that metformin could prevent both caspase-3 activation and apoptosis when induced by palmitic acid.
  
  
  METHODS
  Cardiomyocytes were incubated with 1 mmol/l palmitic acid, in the absence or presence of metformin (1-5 mmol/l). Following 1 to 16 h, cell damage was evaluated by measuring lactate dehydrogenase released into the incubation medium, and Hoechst staining. To investigate the mechanism of metformin's effect on cardiomyocytes, substrate utilisation and phosphorylation of AMPK and acetyl-CoA carboxylase were measured. Intracellular mediators of apoptosis were also evaluated.
  
  
  RESULTS
  Incubation of myocytes with palmitic acid for 16 h increased apoptosis, an effect that was partly blunted by 1 and 2 mmol/l metformin. This beneficial effect of metformin was associated with increased AMPK phosphorylation, palmitic acid oxidation and suppression of high-fat-induced increases in (1) long chain base biosynthesis protein 1 levels, (2) ceramide levels, and (3) caspase-3 activity. Unexpectedly, 5 mmol/l metformin dramatically increased apoptosis in myocytes incubated with high fat. This effect was associated with a robust increase in glycolysis, lactate accumulation, and a significant drop of pH in the myocyte incubation medium.
  
  
  CONCLUSIONS
  Our study demonstrates that metformin reduces high-fat-induced cardiac cell death, probably through inhibition of ceramide synthesis. However, at high concentrations, metformin causes proton and lactate accumulation, leading to cell damage that is independent of caspase-3.
authors.affiliation: Division of Pharmacology and Toxicology, Faculty of Pharmaceutical Sciences, The University of British Columbia, 2146 East Mall, Vancouver, BC, Canada, V6T 1Z3.
authors.forename: D
authors.initials: D
authors.lastname: An
authors.forename: G
authors.initials: G
authors.lastname: Kewalramani
authors.forename: J K Y
authors.initials: JK
authors.lastname: Chan
authors.forename: D
authors.initials: D
authors.lastname: Qi
authors.forename: S
authors.initials: S
authors.lastname: Ghosh
authors.forename: T
authors.initials: T
authors.lastname: Pulinilkunnil
authors.forename: A
authors.initials: A
authors.lastname: Abrahani
authors.forename: S M
authors.initials: SM
authors.lastname: Innis
authors.forename: B
authors.initials: B
authors.lastname: Rodrigues
mesh_terms: D000103:Acetyl-CoA Carboxylase / Q000378:metabolism; D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D053148:Caspase 3 / Q000378:metabolism*; D002478:Cells, Cultured; D002518:Ceramides / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005227:Fatty Acids / Q000378:metabolism; D006019:Glycolysis / Q000187:drug effects; D006863:Hydrogen-Ion Concentration / Q000187:drug effects; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D019344:Lactic Acid / Q000378:metabolism; D008297:Male; D008687:Metformin / Q000494:pharmacology*; D008954:Models, Biological; D032383:Myocytes, Cardiac / Q000166:cytology / Q000187:drug effects*; D019308:Palmitic Acid / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D017382:Reactive Oxygen Species / Q000378:metabolism; D013997:Time Factors
chemical_list: D002518:Ceramides; D005227:Fatty Acids; D017382:Reactive Oxygen Species; D019308:Palmitic Acid; D019344:Lactic Acid; D008687:Metformin; D007770:L-Lactate Dehydrogenase; D017346:Protein Serine-Threonine Kinases; D053148:Caspase 3; D000103:Acetyl-CoA Carboxylase