Cyclosporin A inhibits chromium(VI)-induced apoptosis and mitochondrial cytochrome c release and restores clonogenic survival in CHO cells.
A variety of key events in the molecular apoptotic pathway involve the mitochondria. Cyclosporin A (csA) affects the mitochondria by inhibiting the mitochondrial permeability transition (MPT), thereby preventing disruption of the transmembrane potential. The role of the MPT in apoptosis is not fully understood, but inhibition of the MPT may prevent the release of mitochondrial caspase activators, such as cytochrome c (cyt c), into the cytosol. Certain hexavalent chromium [Cr(VI)] compounds are known occupational respiratory tract toxins and carcinogens. We have previously shown that these compounds induce apoptosis as a predominant mode of cell death and that this effect can be observed in cell culture using soluble Cr(VI). We show here that Cr(VI)-induced apoptosis in Chinese hamster ovary (CHO) cells involves disruption of mitochondrial stability. Using a cyt c-specific monoclonal antibody, we observed a dose-dependent release of mitochondrial cyt c in cytosolic extracts of CHO cells exposed to apoptogenic doses of sodium chromate. Co-treatment of these cells with csA inhibited the release of cyt c and abrogated Cr(VI)-induced apoptosis as determined by a reduction in internucleosomal DNA fragmentation. Co-treatment with csA also markedly increased clonogenic survival of Cr(VI)-treated CHO cells. In contrast, the general caspase inhibitor Z-VAD-FMK markedly inhibited most of the morphological and biochemical parameters of apoptosis but did not prevent cyt c release and did not increase clonogenic survival. These results suggest that the MPT plays an important role in the regulation of mitochondrial cyt c release and that this may be a critical point in the apoptotic pathway in which cells are irreversibly committed to death.
Pritchard
D E
DE
Department of Pharmacology, Program in Genetics and Program in Molecular and Cellular Oncology, The George Washington University Medical Center, 2300 Eye Street NW, Washington, DC 20037, USA.
Singh
J
J
Carlisle
D L
DL
Patierno
S R
SR
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D016466:CHO Cells / Q000187:drug effects; D061945:Caspase Inhibitors; D020169:Caspases / Q000502:physiology; D002470:Cell Survival / Q000187:drug effects; D002840:Chromates / Q000037:antagonists & inhibitors* / Q000633:toxicity*; D006224:Cricetinae; D016572:Cyclosporine / Q000494:pharmacology*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D003574:Cytochrome c Group / Q000378:metabolism*; D003600:Cytosol / Q000187:drug effects / Q000201:enzymology; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D007425:Intracellular Membranes / Q000187:drug effects / Q000378:metabolism / Q000502:physiology; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008928:Mitochondria / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D017670:Sodium Compounds / Q000037:antagonists & inhibitors* / Q000633:toxicity*
D000590:Amino Acid Chloromethyl Ketones; D061945:Caspase Inhibitors; D002840:Chromates; D015853:Cysteine Proteinase Inhibitors; D003574:Cytochrome c Group; D017670:Sodium Compounds; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; C028982:sodium chromate(VI); D016572:Cyclosporine; D020169:Caspases