Abstract |
The mechanism of induction of apoptosis by the novel anti-cancer drug 1-O-octadecyl-2-methyl-rac-glycero-3-phosphocholine (ET-18-OCH3) was investigated in p53-defective SV40 immortalized rat hepatocytes (CWSV1). Exposure to 12 microM ET-18-OCH3 for 36 h induced apoptosis as determined using classical morphological features and agarose gel electrophoresis of genomic DNA. Increased levels of reactive oxygen species (ROS) were detected spectrophotometrically using a nitroblue tetrazolium (NBT) assay in cells treated with ET-18-OCH3. Both the increased generation of ROS and the induction of apoptosis were inhibited when cells were treated concurrently with ET-18-OCH3 in the presence of the antioxidant alpha-tocopherol. Similar results were achieved when cells were switched acutely to choline-deficient (CD) medium in the presence of the antioxidant. The possible role of mitochondria in the generation of ROS was investigated. Both ET-18-OCH3 and CD decreased the phosphatidylcholine (PC) content of mitochondrial and associated membranes, which correlated with depolarization of the mitochondrial membrane as analyzed using 5,5',6,6'-tetramethylbenzimidazolcarbocyanine iodide (JC-1), a sensitive probe of mitochondrial membrane potential. Rotenone, an inhibitor of the mitochondrial electron transport chain, significantly reduced the intracellular level of ROS and prevented mitochondrial membrane depolarization, correlating with a reduction of apoptosis in response to either ET-18-OCH3 or CD. Taken together, these results suggest that the form of p53-independent apoptosis induced by ET-18-OCH3 is mediated by alterations in mitochondrial membrane PC, a loss of mitochondrial membrane potential, and the release of ROS, resulting in completion of apoptosis.
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Authors | A S Vrablic, C D Albright, C N Craciunescu, R I Salganik, S H Zeisel |
Journal | FASEB journal : official publication of the Federation of American Societies for Experimental Biology
(FASEB J)
Vol. 15
Issue 10
Pg. 1739-44
(Aug 2001)
ISSN: 0892-6638 [Print] United States |
PMID | 11481221
(Publication Type: Journal Article, Research Support, Non-U.S. Gov't, Research Support, U.S. Gov't, P.H.S.)
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Chemical References |
- Antigens, Polyomavirus Transforming
- Antioxidants
- Phosphatidylcholines
- Phospholipid Ethers
- Reactive Oxygen Species
- Tumor Suppressor Protein p53
- Vitamin E
- edelfosine
- Nitroblue Tetrazolium
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Topics |
- Animals
- Antigens, Polyomavirus Transforming
(pharmacology)
- Antioxidants
(pharmacology)
- Apoptosis
(drug effects)
- Cell Line, Transformed
- Electrophoresis, Agar Gel
- Hepatocytes
(physiology, ultrastructure)
- Intracellular Membranes
(chemistry, physiology)
- Male
- Membrane Potentials
- Mitochondria, Liver
(chemistry, physiology, ultrastructure)
- Nitroblue Tetrazolium
- Phosphatidylcholines
(analysis)
- Phospholipid Ethers
(pharmacology)
- Rats
- Rats, Inbred F344
- Reactive Oxygen Species
(metabolism)
- Tumor Suppressor Protein p53
(antagonists & inhibitors, physiology)
- Vitamin E
(pharmacology)
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