Real-time single cell analysis of Smac/DIABLO release during apoptosis

We examined the temporal and causal relationship between Smac/DIABLO release, cytochrome c (cyt-c) release, and caspase activation at the single cell level during apoptosis. Cells treated with the broad-spectrum caspase inhibitor z-VAD-fmk, caspase-3 (Casp-3)–deficient MCF-7 cells, as well as Bax-deficient DU-145 cells released Smac/DIABLO and cyt-c in response to proapoptotic agents. Real-time confocal imaging of MCF-7 cells stably expressing Smac/DIABLO-yellow fluorescent protein (YFP) revealed that the average duration of Smac/DIABLO-YFP release was greater than that of cyt-c-green fluorescent protein (GFP). However, there was no significant difference in the time to the onset of release, and both cyt-c-GFP and Smac/DIABLO-YFP release coincided with mitochondrial membrane potential depolarization. We also observed no significant differences in the Smac/DIABLO-YFP release kinetics when z-VAD-fmk–sensitive caspases were inhibited or Casp-3 was reintroduced. Simultaneous measurement of DEVDase activation and Smac/DIABLO-YFP release demonstrated that DEVDase activation occurred within 10 min of release, even in the absence of Casp-3

Ion transport in C6-glioma-cells during oxidative stress

In this work the dose dependent disorders of ion transport processes and mitochondrial membrane potential caused by hydrogen peroxide were investigated with fluorescencespectroscopic methods in C6-Glioma-Cells. These cells served as a model system for one glial subtype, the astrocytes. The transition pore is known to precede the programmed cell death, apoptosis. The apoptotic pathway is interrupted if all mitochondria of the affected cell undergo depolarisation. The following deficiency of high energy substrates leads to cell swelling and plasmamembrane disrupture, the so called onkosis. The results of this work indicate that the mitochondria are involved in the signal cascade of the oxidative stress response in C6-Glioma-Cells

Ionentransporte in C6-Glioma-Zellen bei oxidativem Streß

In this work the dose dependent disorders of ion transport processes and mitochondrial membrane potential caused by hydrogen peroxide were investigated with fluorescencespectroscopic methods in C6-Glioma-Cells. These cells served as a model system for one glial subtype, the astrocytes. The transition pore is known to precede the programmed cell death, apoptosis. The apoptotic pathway is interrupted if all mitochondria of the affected cell undergo depolarisation. The following deficiency of high energy substrates leads to cell swelling and plasmamembrane disrupture, the so called onkosis. The results of this work indicate that the mitochondria are involved in the signal cascade of the oxidative stress response in C6-Glioma-Cells