Ca2+-induced reactive oxygen species production promotes cytochrome c release from rat liver mitochondria via mitochondrial permeability transition (MPT)-dependent and MPT-independent mechanisms: role of cardiolipin.
Release of cytochrome c from mitochondria is considered a critical, early event in the induction of an apoptosis cascade that ultimately leads to programmed cell death. Mitochondrial Ca(2+) loading is a trigger for the release of cytochrome c, although the molecular mechanism underlying this effect is not fully clarified. This study tested the hypothesis that distinct Ca(2+) thresholds may induce cytochrome c release from rat liver mitochondria by membrane permeability transition (MPT)-dependent and independent mechanisms. The involvement of reactive oxygen species (ROS) and cardiolipin in the Ca(2+)-induced cytochrome c release was also investigated. Cytochrome c was quantitated by a new, very sensitive, and rapid reverse-phase high performance liquid chromatography method with a detection limit of 0.1 pmol/sample. We found that a low extramitochondrial Ca(2+) level (2 microM) promoted the release of approximately 13% of the total alamethicin releasable pool of cytochrome c from mitochondria. This release was not depending of MPT; it was mediated by Ca(2+)-induced ROS production and cardiolipin peroxidation and appears to involve the voltage-dependent anion channel. High extramitochondrial Ca(2+) level (20 microM) promoted approximately 45% of the total releasable pool of cytochrome c. This process was MPT-dependent and was also mediated by ROS and cardiolipin. It is suggested that distinct Ca(2+) levels may determine the mode and the amount of cytochrome c release from rat liver mitochondria. The data may help to clarify the molecular mechanism underlying the Ca(2+)-induced release of cytochrome c from rat liver mitochondria and the role played by ROS and cardiolipin in this process.
Petrosillo
Giuseppe
G
Department of Biochemistry and Molecular Biology and Consiglio Nazionale delle Ricerche Institute of Biomembranes and Bioenergetics, University of Bari, Via E. Orabona, 4, 70126 Bari, Italy.
Ruggiero
Francesca M
FM
Pistolese
Marilva
M
Paradies
Giuseppe
G
D000408:Alamethicin / Q000378:metabolism; D000818:Animals; D017209:Apoptosis; D002118:Calcium / Q000378:metabolism*; D002308:Cardiolipins / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D016572:Cyclosporine / Q000494:pharmacology; D045304:Cytochromes c / Q000378:metabolism*; D007473:Ion Channels / Q000378:metabolism; D008099:Liver / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism; D008930:Mitochondria, Liver / Q000378:metabolism*; D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D051381:Rats; D017382:Reactive Oxygen Species; D012430:Ruthenium Red / Q000494:pharmacology; D013050:Spectrometry, Fluorescence; D013997:Time Factors
D002308:Cardiolipins; D007473:Ion Channels; D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D017382:Reactive Oxygen Species; D012430:Ruthenium Red; D000408:Alamethicin; D016572:Cyclosporine; D045304:Cytochromes c; D002118:Calcium