| ER-targeted Bcl-2 and inhibition of ER-associated caspase-12 rescue cultured immortalized cells from ethanol toxicity. | |
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MedLine Citation:
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PMID: 20727705 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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Alcohol abuse, known for promoting apoptosis in the liver and nervous system, is a major public health concern. Despite significant morbidity and mortality resulting from ethanol consumption, the precise cellular mechanism of its toxicity remains unknown. Previous work has shown that wild-type Bcl-2 is protective against ethanol. The present study investigated whether protection from ethanol toxicity involves mitochondrial Bcl-2 or endoplasmic reticulum (ER) Bcl-2, and whether mitochondria-associated or ER-associated caspases are involved in ethanol toxicity. Chinese hamster ovary (CHO695) cells were transiently transfected with cDNA constructs encoding wild-type Bcl-2, mitochondria-targeted Bcl-2, or ER-targeted Bcl-2. MTT assay was used to measure cell viability in response to ethanol. Ethanol treatments of 1 and 2.5 M reduced cell viability at 5, 10, and 24 h. Wild-type Bcl-2, localized both to mitochondria and ER, provided significant rescue for CHO695 cells treated with 1M ethanol for 24 h, but did not rescue toxicity at 2.5 M. ER-targeted Bcl-2, however, provided significant and robust rescue following 24 h of 1 and 2.5 M ethanol. Mitochondria-targeted Bcl-2 offered no protection at any ethanol concentration and generally reduced cell viability. To follow up these experiments, we used a peptide inhibitor approach to investigate which caspases were responsible for ethanol-induced apoptosis. Caspase-9 and caspase-12 are known to be downstream of mitochondria and the ER, respectively. CHO695 cells were treated with a pan-caspase inhibitor, a caspase-9 or caspase-12 inhibitor along with 1.5 M ethanol, followed by MTT cell viability assay. Treatment with the pan-caspase inhibitor provided significant rescue from ethanol, whereas inhibition of caspase-9 did not. Inhibition of ER-associated caspase-12, however, conferred significant protection from ethanol toxicity, similar to the pan inhibitor. These findings are consistent with our transfection data and, taken together, suggest a significant role for the ER in ethanol toxicity. |
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Authors:
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Andreea G Balan; Barret J Myers; Jansi L Maganti; D Blaine Moore |
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Publication Detail:
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Type: Journal Article; Research Support, Non-U.S. Gov't Date: 2010-08-21 |
Journal Detail:
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Title: Alcohol (Fayetteville, N.Y.) Volume: 44 ISSN: 1873-6823 ISO Abbreviation: Alcohol Publication Date: 2010 Sep |
Date Detail:
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Created Date: 2010-09-17 Completed Date: 2011-01-07 Revised Date: - |
Medline Journal Info:
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Nlm Unique ID: 8502311 Medline TA: Alcohol Country: United States |
Other Details:
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Languages: eng Pagination: 553-63 Citation Subset: IM |
Copyright Information:
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Copyright © 2010 Elsevier Inc. All rights reserved. |
Affiliation:
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Department of Biology, Kalamazoo College, Kalamazoo, MI, USA. |
Export Citation:
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APA/MLA Format Download EndNote Download BibTex |
| MeSH Terms | |
Descriptor/Qualifier:
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Animals CHO Cells Caspase 12 / antagonists & inhibitors*, genetics Cell Line, Transformed Cell Survival Cricetinae Cricetulus Endoplasmic Reticulum / chemistry, enzymology, physiology* Enzyme Inhibitors / pharmacology Ethanol / administration & dosage, toxicity* Genes, bcl-2 / genetics Green Fluorescent Proteins / genetics Microscopy, Fluorescence Mitochondria / chemistry, enzymology, physiology Proto-Oncogene Proteins c-bcl-2 / physiology* Recombinant Fusion Proteins / analysis, genetics Transfection |
| Grant Support | |
ID/Acronym/Agency:
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//Howard Hughes Medical Institute |
| Chemical | |
Reg. No./Substance:
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0/Enzyme Inhibitors; 0/Proto-Oncogene Proteins c-bcl-2; 0/Recombinant Fusion Proteins; 147336-22-9/Green Fluorescent Proteins; 64-17-5/Ethanol; EC 3.4.22.-/Caspase 12 |
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