- 作者：Ricardo Quarrie ; Br ; on M. Cramer ; Daniel S. Lee ; Gregory E. Steinbaugh ; Warren Erdahl ; Douglas R. Pfeiffer ; Jay L. Zweier ; Juan A. Crestanello
- 关键词：ischemia ; reperfusion ; reactive oxygen species ; mitochondria ; proton leak ; uncoupling ; heart
- 刊名：Journal of Surgical Research
- 出版年：2011
- 出版时间：January 2011
- 年：2011
- 卷：165
- 期：1
- 页码：5-14
- 全文大小：641 K
Background
Proton leak (H+ leak) dissipates mitochondrial membrane potential (mΔΨ) through the re-entry of protons into the mitochondrial matrix independent of ATP synthase. Changes in H+ leak may affect reactive oxygen species (ROS) production. We measured H+ leak and ROS production during ischemia-reperfusion and ischemic preconditioning (IPC) and examined how changing mitochondrial respiration affected mΔΨ and ROS production.Materials and Methods
Isolated rat hearts (n = 6/group) were subjected to either control-IR or IPC. Rate pressure product (RPP) was measured. Mitochondria were isolated at end reperfusion. Respiration was measured by polarography and titrated with increasing concentrations of malonate (0.5–2 mM). mΔΨ was measured using a tetraphenylphosphonium electrode. H+ leak is the respiratory rate required to maintain membrane potential at –150 mV in the presence of oligomycin-A. Mitochondrial complex III ROS production was measured by fluorometry using Amplex-red.
Results
IPC improved recovery of RPP at end reperfusion (63 % ± 4 % versus 21 % ± 2 % in control-IR, P < 0.05). Ischemia-reperfusion caused increased H+ leak (94 ± 12 versus 31 ± 1 nmol O/mg protein/min in non-ischemic control, P < 0.05). IPC attenuates these increases (55 ± 9 nmol O/mg protein/min, P < 0.05 versus control-IR). IPC reduced mitochondrial ROS production compared with control-IR (31 ± 2 versus 40 ± 3 nmol/mg protein/min, P < 0.05). As mitochondrial respiration decreased, mΔΨ and mitochondrial ROS production also decreased. ROS production remained lower in IPC than in control-IR for all mΔΨ and respiration rates.
Conclusions
Increasing H+ leak is not associated with decreased ROS production. IPC decreases both the magnitude of H+ leak and ROS production after ischemia-reperfusion.