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Ischaemic accumulation of succinate controls reperfusion injury through mitochondrial ROS.

ABSTRACT: 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). Although mitochondrial ROS production in ischaemia reperfusion is established, it has generally been considered a nonspecific response to reperfusion. 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.

SUBMITTER: Chouchani ET

PROVIDER: S-EPMC4255242 | biostudies-literature | 2014 Nov

REPOSITORIES: biostudies-literature

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Ischaemic accumulation of succinate controls reperfusion injury through mitochondrial ROS.

Chouchani Edward T ET Pell Victoria R VR Gaude Edoardo E Aksentijević Dunja D Sundier Stephanie Y SY Robb Ellen L EL Logan Angela A Nadtochiy Sergiy M SM Ord Emily N J ENJ Smith Anthony C AC Eyassu Filmon F Shirley Rachel R Hu Chou-Hui CH Dare Anna J AJ James Andrew M AM Rogatti Sebastian S Hartley Richard C RC Eaton Simon S Costa Ana S H ASH Brookes Paul S PS Davidson Sean M SM Duchen Michael R MR Saeb-Parsy Kourosh K Shattock Michael J MJ Robinson Alan J AJ Work Lorraine M LM Frezza Christian C Krieg Thomas T Murphy Michael P MP

Nature 20141105 7527

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). Although mitochondrial ROS production in ischaemia reperfusion is established, it has generally been considered a nonspeci ...[more]

PMID: 25383517

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Project description:AimsNo effective therapy is available in clinics to protect the heart from ischaemia/reperfusion (I/R) injury. Endothelial cells are activated after I/R, which may drive the inflammatory response by releasing ATP through pannexin1 (Panx1) channels. Here, we investigated the role of Panx1 in cardiac I/R.Methods and resultsPanx1 was found in cardiac endothelial cells, neutrophils, and cardiomyocytes. After in vivo I/R, serum Troponin-I, and infarct size were less pronounced in Panx1-/- mice, but leukocyte infiltration in the infarct area was similar between Panx1-/- and wild-type mice. Serum Troponin-I and infarct size were not different between mice with neutrophil-specific deletion of Panx1 and Panx1fl/fl mice, suggesting that cardioprotection by Panx1 deletion rather involved cardiomyocytes than the inflammatory response. Physiological cardiac function in wild-type and Panx1-/- hearts was similar. The time to onset of contracture and time to maximal contracture were delayed in Panx1-/- hearts, suggesting reduced sensitivity of these hearts to ischaemic injury. Moreover, Panx1-/- hearts showed better recovery of left ventricle developed pressure, cardiac contractility, and relaxation after I/R. Ischaemic preconditioning failed to confer further protection in Panx1-/- hearts. Panx1 was found in subsarcolemmal mitochondria (SSM). SSM in WT or Panx1-/- hearts showed no differences in morphology. The function of the mitochondrial permeability transition pore and production of reactive oxygen species in SSM was not affected, but mitochondrial respiration was reduced in Panx1-/- SSM. Finally, Panx1-/- cardiomyocytes had a decreased mitochondrial membrane potential and an increased mitochondrial ATP content.ConclusionPanx1-/- mice display decreased sensitivity to cardiac I/R injury, resulting in smaller infarcts and improved recovery of left ventricular function. This cardioprotective effect of Panx1 deletion seems to involve cardiac mitochondria rather than a reduced inflammatory response. Thus, Panx1 may represent a new target for controlling cardiac reperfusion damage.

Project description:Background The metabolite succinate accumulates during cardiac ischemia. Within 5 minutes of reperfusion, succinate returns to baseline levels via both its release from cells and oxidation by mitochondrial complex II. The latter drives reactive oxygen species (ROS) generation and subsequent opening of the mitochondrial permeability transition (PT) pore, leading to cell death. Targeting succinate dynamics (accumulation/oxidation/release) may be therapeutically beneficial in cardiac ischemia-reperfusion (IR) injury. It has been proposed that blocking MCT1 (monocarboxylate transporter 1) may be beneficial in IR injury, by preventing succinate release and subsequent engagement of downstream inflammatory signaling pathways. In contrast, herein we hypothesized that blocking MCT1 would retain succinate in cells, exacerbating ROS generation and IR injury. Methods and Results Using the mitochondrial ROS probe mitoSOX and a custom-built murine heart perfusion rig built into a spectrofluorometer, we measured ROS generation in situ during the first moments of reperfusion. We found that acute MCT1 inhibition enhanced mitochondrial ROS generation at reperfusion and worsened IR injury (recovery of function and infarct size). Both of these effects were abrogated by tandem inhibition of mitochondrial complex II, suggesting that succinate retention worsens IR because it drives more mitochondrial ROS generation. Furthermore, using the PT pore inhibitor cyclosporin A, along with monitoring of PT pore opening via the mitochondrial membrane potential indicator tetramethylrhodamine ethyl ester, we herein provide evidence that ROS generation during early reperfusion is upstream of the PT pore, not downstream as proposed by others. In addition, pore opening was exacerbated by MCT1 inhibition. Conclusions Together, these findings highlight the importance of succinate dynamics and mitochondrial ROS generation as key determinants of PT pore opening and IR injury outcomes.

Project description:BackgroundElectroacupuncture (EA) has been shown to promote functional recovery after cerebral ischemia-reperfusion (I/R) injury. However, the contribution of mitochondrial dynamics to recovery remains unclear. The aim of this study was to investigate whether mitochondrial dynamics are involved in the effects of EA on cerebral I/R injury.MethodsThe rats with cerebral I/R injury were established by the middle cerebral artery occlusion/reperfusion. Subsequently, EA was applied to Baihui (GV20) and Dazhui (GV14) acupoints, with 2 Hz/5 Hz in frequency, 1.0 mA in intensity, 20 min each time, once a day for seven consecutive days. The therapeutic outcomes were assessed by modified neurological severity score (mNSS), 2,3,5-Triphenyte-trazolium chloride (TTC) staining, and hematoxylin-eosin (HE) staining. Mitochondrial morphology was observed under transmission electron microscopy. Adenosine triphosphate (ATP) content and ATP synthases (ATPases) activity were evaluated to measure mitochondrial function using ELISA. Finally, mitochondrial dynamics-related molecules, including dynamin-related protein 1 (Drp1), fission 1 (Fis1), mitofusin 1 (Mfn1), mitofusin 2 (Mfn2), and optic atrophy 1 (OPA1), were detected by Western blot and immunofluorescence staining.ResultsCerebral I/R injury induced neurological dysfunction, cerebral infarction and neuronal injury, all of which were ameliorated by EA. And EA improved mitochondrial morphology and function. Moreover, EA altered the balance of mitochondrial dynamics. Specifically, the data showed a significant decrease in the expression of Drp1 and Fis1, leading to the inhibition of mitochondrial fission. Additionally, Mfn1, Mfn2 and Opa1, which are related to mitochondrial fusion, were effectively promoted after EA treatment. However, sham EA did not show any neuroprotective effects in rats with cerebral I/R injury.ConclusionsIn summary, our study indicates that the balance of mitochondrial dynamics is crucial for EA therapy to treat cerebral I/R injury.

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Ischaemic accumulation of succinate controls reperfusion injury through mitochondrial ROS.

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Ischaemic accumulation of succinate controls reperfusion injury through mitochondrial ROS.

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