Mitochondrial metabolism and bioenergetic function in an anoxic isolated adult mouse cardiomyocyte model of in vivo cardiac ischemia-reperfusion injury
Anja V. Gruszczyk,
Alva M. Casey,
Andrew M. James,
Hiran A. Prag,
Nils Burger,
Georgina R. Bates,
Andrew R. Hall,
Fay M. Allen,
Thomas Krieg,
Kourosh Saeb-Parsy,
Michael P. Murphy
Affiliations
Anja V. Gruszczyk
MRC Mitochondrial Biology Unit, Biomedical Campus, University of Cambridge, Cambridge, CB2 0XY, UK; Department of Surgery and Cambridge NIHR Biomedical Research Centre, Biomedical Campus, University of Cambridge, Cambridge, CB2 2QQ, UK; NIHR Biomedical Research Centre and NIHR Blood and Transplant Research Unit in Organ Donation and Transplantation, Cambridge Biomedical Campus, Cambridge, UK
Alva M. Casey
MRC Mitochondrial Biology Unit, Biomedical Campus, University of Cambridge, Cambridge, CB2 0XY, UK
Andrew M. James
MRC Mitochondrial Biology Unit, Biomedical Campus, University of Cambridge, Cambridge, CB2 0XY, UK
Hiran A. Prag
MRC Mitochondrial Biology Unit, Biomedical Campus, University of Cambridge, Cambridge, CB2 0XY, UK; Department of Medicine, University of Cambridge, Cambridge, CB2 0QQ, UK
Nils Burger
MRC Mitochondrial Biology Unit, Biomedical Campus, University of Cambridge, Cambridge, CB2 0XY, UK
Georgina R. Bates
MRC Mitochondrial Biology Unit, Biomedical Campus, University of Cambridge, Cambridge, CB2 0XY, UK
Andrew R. Hall
MRC Mitochondrial Biology Unit, Biomedical Campus, University of Cambridge, Cambridge, CB2 0XY, UK
Fay M. Allen
MRC Mitochondrial Biology Unit, Biomedical Campus, University of Cambridge, Cambridge, CB2 0XY, UK
Thomas Krieg
Department of Medicine, University of Cambridge, Cambridge, CB2 0QQ, UK
Kourosh Saeb-Parsy
Department of Surgery and Cambridge NIHR Biomedical Research Centre, Biomedical Campus, University of Cambridge, Cambridge, CB2 2QQ, UK; NIHR Biomedical Research Centre and NIHR Blood and Transplant Research Unit in Organ Donation and Transplantation, Cambridge Biomedical Campus, Cambridge, UK
Michael P. Murphy
MRC Mitochondrial Biology Unit, Biomedical Campus, University of Cambridge, Cambridge, CB2 0XY, UK; Department of Medicine, University of Cambridge, Cambridge, CB2 0QQ, UK; Corresponding author. MRC Mitochondrial Biology Unit, Biomedical Campus, University of Cambridge, Cambridge, CB2 0XY, UK.
Cell models of cardiac ischemia-reperfusion (IR) injury are essential to facilitate understanding, but current monolayer cell models poorly replicate the in vivo IR injury that occurs within a three-dimensional tissue. Here we show that this is for two reasons: the residual oxygen present in many cellular hypoxia models sustains mitochondrial oxidative phosphorylation; and the loss of lactate from cells into the incubation medium during ischemia enables cells to sustain glycolysis. To overcome these limitations, we incubated isolated adult mouse cardiomyocytes anoxically while inhibiting lactate efflux. These interventions recapitulated key markers of in vivo ischemia, notably the accumulation of succinate and the loss of adenine nucleotides. Upon reoxygenation after anoxia the succinate that had accumulated during anoxia was rapidly oxidized in association with extensive mitochondrial superoxide/hydrogen peroxide production and cell injury, mimicking reperfusion injury. This cell model will enable key aspects of cardiac IR injury to be assessed in vitro.
