The role of succinate and ROS in reperfusion injury – A critical appraisal

Abstract

We critically assess the proposal that succinate-fuelled reverse electron flow (REF) drives mitochondrial matrix superoxide production from Complex I early in reperfusion, thus acting as a key mediator of ischemia/reperfusion (IR) injury. Real-time surface fluorescence measurements of NAD(P)H and flavoprotein redox state suggest that conditions are unfavourable for REF during early reperfusion. Furthermore, rapid loss of succinate accumulated during ischemia can be explained by its efflux rather than oxidation. Moreover, succinate accumulation during ischemia is not attenuated by ischemic preconditioning (IP) despite powerful cardioprotection. In addition, measurement of intracellular reactive oxygen species (ROS) during reperfusion using surface fluorescence and mitochondrial aconitase activity detected major increases in ROS only after mitochondrial permeability transition pore (mPTP) opening was first detected. We conclude that mPTP opening is probably triggered initially by factors other than ROS, including increased mitochondrial [Ca2+]. However, IP only attenuates [Ca2+] increases later in reperfusion, again after initial mPTP opening, implying that IP regulates mPTP opening through additional mechanisms. One such is mitochondria-bound hexokinase 2 (HK2) which dissociates from mitochondria during ischemia in control hearts but not those subject to IP. Indeed, there is a strong correlation between the extent of HK2 loss from mitochondria during ischemia and infarct size on subsequent reperfusion. Mechanisms linking HK2 dissociation to mPTP sensitisation remain to be fully established but several related processes have been implicated including VDAC1 oligomerisation, the stability of contact sites between the inner and outer membranes, cristae morphology, Bcl-2 family members and mitochondrial fission proteins such as Drp1.

Repurposing drugs as pro‐oxidant redox modifiers to eliminate cancer stem cells and improve the treatment of advanced stage cancers

“…However, TRAP1 also limits ROS generation from the respiratory chain SDH/complex II, thereby increasing the Ca 2+ threshold required for mPTP opening in cancer cells, protecting them from death stimuli . High Ca 2+ loads (approximately 50 nmol Ca 2+ /mg protein or higher concentrations, depending on the oxidizable substrate) into mitochondria are critically involved along with ROS as triggers for activation of the mPTP and cell death induction (Figure ) . However, it has not been elucidated exactly how the mitochondrial Ca 2+ (mCa 2+ ) overload triggers cell death.…”

“…Together, these data show that the mitochondrial respiratory response and the kinetics of ΔΨ m loss from mPTP opening will vary depending on metabolic substrate availability. In marked contrast, in another study with isolated heart mitochondria, it was shown that the presence of succinate (with no addition of rotenone) provided resistance to the mPTP opening induced by Ca 2+ loads and despite promoting increased rates of ROS production (as compared with glutamate + malate as the oxidizable substrates). Further, once mPTP opening occurred, the mitochondria produced much more ROS in the presence of succinate.…”

“…Adding the CypD inhibitor, cyclosporin A or using the Ca 2+ uptake inhibitor, ruthenium red prevented the Ca 2+ activation of mitochondria, which then tolerated greater Ca 2+ loads, as well as the associated decline in respiration or collapse of ΔΨ m , confirming the dependence of mPTP opening on mCa 2+ uptake . The further increased ROS production occurring after mPTP opening and observed with either succinate or glutamate + malate may have derived from the Ca 2+ load‐induced inhibition of the respiratory chain activity. This would be consistent with the general actions of Ca 2+ and ROS on CL oxidation, the release of HK bound to the mPTP, the extent of dissociation of SDH/complex II and its associated ROS production combining with cyt c release to ultimately trigger the intrinsic apoptosis pathway.…”

“…131 High Ca 2+ loads (approximately 50 nmol Ca 2+ /mg protein or higher concentrations, depending on the oxidizable substrate) into mitochondria are critically involved along with ROS as triggers for activation of the mPTP and cell death induction (Figure 3). 132 However, it has not been elucidated exactly how the mitochondrial Ca 2+ (mCa 2+ ) overload triggers cell death. Low extramitochondrial free Ca 2+ levels (2 μM), which are still within the physiological range, may promote release of a small cytochrome c (cytc) fraction (approximately 13%) from mitochondria.…”

A Self‐Assembled Fluorescent Nanoprobe for Imaging and Therapy of Cardiac Ischemia/Reperfusion Injury

“…Thus, a ROS-responsive nanoparticle drug-delivery system offers a promising approach for the treatment of various I/R injuries such as stroke and renal infarction. [20] Conjugation of ROS-responsive nanoparticles to an MRI tracer such as gadolinium or to a radiotracer for PET imaging holds great promise for utilizing ROS-responsive nanoparticles for diagnosis of MI and in vivo assessment of ROS levels. [14][15][16][17][18] Therefore, ROS-responsive nanoparticles may also represent a promising strategy for targeted drug delivery in the treatment of a number of diseases in addition to I/R injury.…”