Ischaemia-reperfusion injury occurs when the blood supply to an organ is disrupted and then restored, and underlies many disorders, notably heart attack and stroke. While reperfusion of ischaemic tissue is essential for survival, it also initiates oxidative damage, cell death and aberrant immune responses through the generation of mitochondrial reactive oxygen species (ROS)1,2,3,4,5. Although mitochondrial ROS production in ischaemia reperfusion is established, it has generally been considered a nonspecific response to reperfusion1,3. Here we develop a comparative in vivo metabolomic analysis, and unexpectedly identify widely conserved metabolic pathways responsible for mitochondrial ROS production during ischaemia reperfusion. We show that selective accumulation of the citric acid cycle intermediate succinate is a universal metabolic signature of ischaemia in a range of tissues and is responsible for mitochondrial ROS production during reperfusion. Ischaemic succinate accumulation arises from reversal of succinate dehydrogenase, which in turn is driven by fumarate overflow from purine nucleotide breakdown and partial reversal of the malate/aspartate shuttle. After reperfusion, the accumulated succinate is rapidly re-oxidized by succinate dehydrogenase, driving extensive ROS generation by reverse electron transport at mitochondrial complex I. Decreasing ischaemic succinate accumulation by pharmacological inhibition is sufficient to ameliorate in vivo ischaemia-reperfusion injury in murine models of heart attack and stroke. Thus, we have identified a conserved metabolic response of tissues to ischaemia and reperfusion that unifies many hitherto unconnected aspects of ischaemia-reperfusion injury. Furthermore, these findings reveal a new pathway for metabolic control of ROS production in vivo, while demonstrating that inhibition of ischaemic succinate accumulation and its oxidation after subsequent reperfusion is a potential therapeutic target to decrease ischaemia-reperfusion injury in a range of pathologies.

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Supported by the Medical Research Council (UK) and by grants from Canadian Institutes of Health Research and the Gates Cambridge Trust (E.T.C.) and the British Heart Foundation (T.K., V.R.P., L.M.W.). We thank J. Hirst and G. C. Brown for discussions.

Author information

Author notes

    • Edward T. Chouchani
    •  & Victoria R. Pell

    These authors contributed equally to this work.


  1. MRC Mitochondrial Biology Unit, Hills Road, Cambridge CB2 0XY, UK

    • Edward T. Chouchani
    • , Ellen L. Robb
    • , Angela Logan
    • , Anthony C. Smith
    • , Filmon Eyassu
    • , Anna J. Dare
    • , Andrew M. James
    • , Sebastian Rogatti
    • , Alan J. Robinson
    •  & Michael P. Murphy
  2. Department of Medicine, University of Cambridge, Addenbrooke’s Hospital, Hills Road, Cambridge CB2 0QQ, UK

    • Edward T. Chouchani
    • , Victoria R. Pell
    • , Chou-Hui Hu
    •  & Thomas Krieg
  3. MRC Cancer Unit, University of Cambridge, Hutchison/MRC Research Centre, Box 197, Cambridge Biomedical Campus, Cambridge CB2 0XZ, UK

    • Edoardo Gaude
    • , Ana S. H. Costa
    •  & Christian Frezza
  4. King’s College London, British Heart Foundation Centre of Research Excellence, The Rayne Institute, St Thomas’ Hospital, London SE1 7EH, UK

    • Dunja Aksentijević
    •  & Michael J. Shattock
  5. Department of Cell and Developmental Biology and UCL Consortium for Mitochondrial Biology, University College London, Gower Street, London WC1E 6BT, UK

    • Stephanie Y. Sundier
    •  & Michael R. Duchen
  6. Department of Anesthesiology, University of Rochester Medical Center, 601 Elmwood Avenue, Rochester, New York 14642, USA

    • Sergiy M. Nadtochiy
    •  & Paul S. Brookes
  7. Institute of Cardiovascular & Medical Sciences, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow G12 8TA, UK

    • Emily N. J. Ord
    • , Rachel Shirley
    •  & Lorraine M. Work
  8. School of Chemistry, University of Glasgow, Glasgow G12 8QQ, UK

    • Richard C. Hartley
  9. Unit of Paediatric Surgery, UCL Institute of Child Health, London WC1N 1EH, UK

    • Simon Eaton
  10. Hatter Cardiovascular Institute, University College London, 67 Chenies Mews, London WC1E 6HX, UK

    • Sean M. Davidson
  11. University Department of Surgery and Cambridge NIHR Biomedical Research Centre, Addenbrooke’s Hospital, Cambridge CB2 0QQ, UK

    • Kourosh Saeb-Parsy


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E.T.C. designed research, carried out biochemical experiments, analysed data from in vivo experiments and co-wrote the paper. T.K., V.R.P. and C.-H.H. designed and carried out the ex vivo and in vivo experiments. C.F. and E.G. designed and carried out mass spectrometry and metabolomics analyses, with A.S.H.C. assisting. D.A. and M.J.S. designed and carried out ex vivo perfused heart experiments. S.Y.S., S.M.D., M.R.D., S.M.N., E.L.R. and P.S.B. designed and carried out cell experiments. L.M.W., E.N.J.O. and R.S. designed and carried out brain experiments. A.J.D., S.R. and K.S.-P. designed and carried out kidney experiments. A.L. and R.C.H. carried out ROS analyses. S.E. carried out analyses. A.M.J. helped with data interpretation. A.C.S., A.J.R. and F.E. designed and performed bioinformatic analyses. E.T.C., T.K., C.F. and M.P.M. directed the research and co-wrote the paper, with assistance from all other authors.

Competing interests

M.P.M., E.T.C., L.M.W., C.F. and T.K. have applied for a patent on some of the work described here.

Corresponding authors

Correspondence to Christian Frezza or Thomas Krieg or Michael P. Murphy.

Extended data

Extended data figures

  1. 1.

    Comparative analysis of metabolites significantly accumulated in ischaemic conditions.

  2. 2.

    Time course of succinate levels in the in vivo heart during ischaemia and reperfusion and potential metabolic inputs for succinate.

  3. 3.

    Metabolic labelling of CAC and proximal metabolites by 13C-glucose in the ischaemic and normoxic myocardium.

  4. 4.

    Metabolic labelling of CAC and proximal metabolites by 13C-palmitate in the ischaemic and normoxic myocardium.

  5. 5.

    Metabolic labelling of CAC and proximal metabolites by 13C-glutamine in the ischaemic and normoxic myocardium, and measurement of the effect of inhibition of GABA transaminase on succinate accumulation in the ischaemic myocardium.

  6. 6.

    Unabridged metabolic model identifying pathways that can become activated by tissue ischaemia to drive succinate accumulation.

  7. 7.

    Effects of dimethyl malonate and dimethyl succinate treatment of cells and in vivo on intracellular accumulation of malonate and succinate, and respiration and comparison of 13C-labelled ischaemic metabolite fluxes to succinate relative to isotopic donor pools.

  8. 8.

    Predicted changes in pathways of succinate and oxidative phosphorylation metabolism during ischaemia and following reperfusion.

  9. 9.

    Tracking DHE oxidation, NAD(P)H reduction state, and mitochondrial membrane potential in primary cardiomyocytes during in situ IR.

  10. 10.

    Quantification of CAC intermediates in the heart following infusion of dimethyl succinate and in the brain after infusion of dimethyl malonate, and extended summary cytoprotection and neurological scores of rats subjected to tMCAO IR in vivo ± dimethyl malonate infusion.

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