Distinct mPTP activation mechanisms in ischaemia-reperfusion: contributions of Ca2+, ROS, pH, and inorganic polyphosphate.
OBJECTIVE
The mitochondrial permeability transition pore (mPTP) plays a central role for tissue damage and cell death during ischaemia-reperfusion (I/R). We investigated the contribution of mitochondrial inorganic polyphosphate (polyP), a potent activator of Ca(2+)-induced mPTP opening, towards mPTP activation and cardiac cell death in I/R.


RESULTS
A significant increase in mitochondrial free calcium concentration ([Ca(2+)]m), reactive oxygen species (ROS) generation, mitochondrial membrane potential depolarization (ΔΨm), and mPTP activity, but no cell death, was observed after 20 min of ischaemia. The [Ca(2+)]m increase during ischaemia was partially prevented by the mitochondrial Ca(2+) uniporter (MCU) inhibitor Ru360 and completely abolished by the combination of Ru360 and the ryanodine receptor type 1 blocker dantrolene, suggesting two complimentary Ca(2+) uptake mechanisms. In the absence of Ru360 and dantrolene, mPTP closing by polyP depletion or CSA decreased mitochondrial Ca(2+) uptake, suggesting that during ischaemia Ca(2+) can enter mitochondria through mPTP. During reperfusion, a burst of endogenous polyP production coincided with a decrease in [Ca(2+)]m, a decline in superoxide generation, and an acceleration of hydrogen peroxide (H2O2) production. An increase in H2O2 correlated with restoration of mitochondrial pHm and an increase in cell death. mPTP opening and cell death on reperfusion were prevented by antioxidants Trolox and MnTBAP [Mn (III) tetrakis (4-benzoic acid) porphyrin chloride]. Enzymatic polyP depletion did not affect mPTP opening during reperfusion, but increased ROS generation and cell death, suggesting that polyP plays a protective role in cellular stress response.


CONCLUSIONS
Transient Ca(2+)/polyP-mediated mPTP opening during ischaemia may serve to protect cells against cytosolic Ca(2+) overload, whereas ROS/pH-mediated sustained mPTP opening on reperfusion induces cell death.
Department of Molecular Biophysics and Physiology, Rush University Medical Center, 1750 W. Harrison St, Chicago, IL 60612, USA.
Lea K
LK
Seidlmayer
Department of Molecular Biophysics and Physiology, Rush University Medical Center, 1750 W. Harrison St, Chicago, IL 60612, USA.
Vanessa V
VV
Juettner
Institute of Cardiovascular and Medical Sciences, College of Veterinary Medical and Life Sciences, University of Glasgow, Glasgow, UK.
Sarah
S
Kettlewell
Dalhousie University, Halifax, NS, Canada New York University, NY, USA.
Evgeny V
EV
Pavlov
Department of Molecular Biophysics and Physiology, Rush University Medical Center, 1750 W. Harrison St, Chicago, IL 60612, USA.
Lothar A
LA
Blatter
Department of Molecular Biophysics and Physiology, Rush University Medical Center, 1750 W. Harrison St, Chicago, IL 60612, USA Elena_Dedkova@rush.edu.
Elena N
EN
Dedkova
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D016923:Cell Death / Q000187:drug effects; D006861:Hydrogen Peroxide / Q000378:metabolism; D007511:Ischemia / Q000378:metabolism*; D053078:Membrane Potential, Mitochondrial / Q000502:physiology; D008929:Mitochondria, Heart / Q000378:metabolism; D033681:Mitochondrial Membrane Transport Proteins / Q000378:metabolism*; D000083162:Mitochondrial Permeability Transition Pore; D015428:Myocardial Reperfusion Injury / Q000378:metabolism*; D010710:Phosphates / Q000378:metabolism*; D011122:Polyphosphates / Q000378:metabolism; D011817:Rabbits; D017382:Reactive Oxygen Species / Q000378:metabolism*
D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D010710:Phosphates; D011122:Polyphosphates; D017382:Reactive Oxygen Species; D006861:Hydrogen Peroxide; D002118:Calcium
Inorganic polyphosphate; Ischaemia–reperfusion injury; Mitochondrial permeability transition pore; Mitochondrial ryanodine receptor; Oxidative stress