title: Investigating mitochondrial metabolism in contracting HL-1 cardiomyocytes following hypoxia and pharmacological HIF activation identifies HIF-dependent and independent mechanisms of regulation.
abstract: Hypoxia is a consequence of cardiac disease and downregulates mitochondrial metabolism, yet the molecular mechanisms through which this occurs in the heart are incompletely characterized. Therefore, we aimed to use a contracting HL-1 cardiomyocyte model to investigate the effects of hypoxia on mitochondrial metabolism. Cells were exposed to hypoxia (2% O2) for 6, 12, 24, and 48 hours to characterize the metabolic response. Cells were subsequently treated with the hypoxia inducible factor (HIF)-activating compound, dimethyloxalylglycine (DMOG), to determine whether hypoxia-induced mitochondrial changes were HIF dependent or independent, and to assess the suitability of this cultured cardiac cell line for cardiovascular pharmacological studies. Hypoxic cells had increased glycolysis after 24 hours, with glucose transporter 1 and lactate levels increased 5-fold and 15-fold, respectively. After 24 hours of hypoxia, mitochondrial networks were more fragmented but there was no change in citrate synthase activity, indicating that mitochondrial content was unchanged. Cellular oxygen consumption was 30% lower, accompanied by decreases in the enzymatic activities of electron transport chain (ETC) complexes I and IV, and aconitase by 81%, 96%, and 72%, relative to controls. Pharmacological HIF activation with DMOG decreased cellular oxygen consumption by 43%, coincident with decreases in the activities of aconitase and complex I by 26% and 30%, indicating that these adaptations were HIF mediated. In contrast, the hypoxia-mediated decrease in complex IV activity was not replicated by DMOG treatment, suggesting HIF-independent regulation of this complex. In conclusion, 24 hours of hypoxia increased anaerobic glycolysis and decreased mitochondrial respiration, which was associated with changes in ETC and tricarboxylic acid cycle enzyme activities in contracting HL-1 cells. Pharmacological HIF activation in this cardiac cell line allowed both HIF-dependent and independent mitochondrial metabolic changes to be identified.
authors.affiliation: Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, England, United Kingdom.
authors.forename: Lucy J A
authors.initials: LJ
authors.lastname: Ambrose
authors.affiliation: Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, England, United Kingdom.
authors.forename: Amira H
authors.initials: AH
authors.lastname: Abd-Jamil
authors.affiliation: Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, England, United Kingdom.
authors.forename: Renata S M
authors.initials: RS
authors.lastname: Gomes
authors.affiliation: Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, England, United Kingdom.
authors.forename: Emma E
authors.initials: EE
authors.lastname: Carter
authors.affiliation: Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, England, United Kingdom.
authors.forename: Carolyn A
authors.initials: CA
authors.lastname: Carr
authors.affiliation: Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, England, United Kingdom.
authors.forename: Kieran
authors.initials: K
authors.lastname: Clarke
authors.affiliation: Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, England, United Kingdom lisa.heather@dpag.ox.ac.uk.
authors.forename: Lisa C
authors.initials: LC
authors.lastname: Heather
mesh_terms: D000154:Aconitate Hydratase / Q000378:metabolism; D000600:Amino Acids, Dicarboxylic / Q000494:pharmacology*; D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics / Q000378:metabolism*; D015687:Cell Hypoxia; D002460:Cell Line; D019069:Cell Respiration / Q000187:drug effects; D042967:Electron Transport Complex I / Q000378:metabolism; D003576:Electron Transport Complex IV / Q000378:metabolism; D051272:Glucose Transporter Type 1 / Q000819:agonists / Q000378:metabolism; D006019:Glycolysis / Q000187:drug effects; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000235:genetics / Q000378:metabolism*; D019344:Lactic Acid / Q000378:metabolism; D051379:Mice; D008929:Mitochondria, Heart / Q000187:drug effects* / Q000378:metabolism; D009200:Myocardial Contraction / Q000187:drug effects*; D032383:Myocytes, Cardiac / Q000187:drug effects* / Q000378:metabolism; D010101:Oxygen Consumption / Q000187:drug effects; D012333:RNA, Messenger / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013997:Time Factors; D015854:Up-Regulation; D000099162:Endothelial PAS Domain-Containing Protein 1
chemical_list: D000154:Aconitate Hydratase; D000600:Amino Acids, Dicarboxylic; D051792:Basic Helix-Loop-Helix Proteins; D042967:Electron Transport Complex I; D003576:Electron Transport Complex IV; D051272:Glucose Transporter Type 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D019344:Lactic Acid; D012333:RNA, Messenger; D000099162:Endothelial PAS Domain-Containing Protein 1; C497443:Hif1a protein, mouse; C495678:Slc2a1 protein, mouse; C040947:oxalylglycine
keywords: cardiac pharmacology; high altitude medicine and physiology; molecular biology