FIG. 13.

Mitochondrial ROS generation during hepatic ischemia/reperfusion (I/R) injury. In vivo two-photon confocal microscopy during partial (70% of the liver) hepatic I/R (90 min of ischemia followed by 60 min of reperfusion) in mice. The generation of ROS, by using 2',7'-dichlorofluorescin (DCF) fluorescence, occurs largely in mitochondria and is accompanied by loss of mitochondrial membrane potential, as detected by using tetramethylrhodamine ethyl ester (TMRE). The imaging of the liver of living mice was performed by the using spectral confocal inverted microscope Leica TCS SP5. DCF (200 μM, incubated for 45 min) and TMRE (5 μM, incubated for 30 min) labelings were imaged by using one-photon excitation at 485 nm and emission at 520 nm for the DCF and excitation at 516 nm and emission at 582–677 nm for the TMRE. The emissions were acquired with internal spectral detector (PMT) at high speed to avoid the movement from animal breathing and heart beating during acquisition. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article at www.liebertonline.com/ars).