| Biochemical and morphological events during okadaic acid-induced apoptosis of Tsc2-null ERC-18 cell line. | |
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MedLine Citation:
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PMID: 11950167 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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Several tumor suppressor genes have been shown to regulate cellular susceptibility to proliferation or apoptotic cell death. An essential first step in studies with the long-range goal of determining the effect of a tumor suppressor gene on cellular susceptibility to apoptosis is careful characterization of the cell's response to an apoptotic stimulus. The goals of this study were to characterize the apoptotic response of a tuberous sclerosis complex-2 (Tsc2) tumor suppressor gene-null cell line, to establish valid biochemical events that can be used as apoptosis markers, and to determine how these events correlate with apoptosis-specific morphologic changes. For characterization of apoptosis, we treated Tsc2-null renal epithelial tumor cells (ERC-18) with okadaic acid (OKA, 0.1-0.25 microM), and measured the biochemical and morphologic events during the apoptotic response. Electron microscopic and immunocytochemical evaluation showed an early loss of microvilli and a loss of vinculin and talin staining from focal adhesions within 1 hour. During the first 2 hours of treatment with 0.25 microM OKA, ERC-18 cells rounded and approximately 50% detached from the culture vessel with minimal membrane bleb formation. Phosphatidylserine externalization, chromatin margination and fragmentation, cytochrome C release, and caspase-3 and -7 cleavage were evident at 6 hours. Maximal membrane bleb formation occurred between 6 and 10 hours. Cells progressed to secondary oncotic necrosis between 10 and 24 hours of OKA treatment. Almost all cells had an oncotic phenotype after 24 hours, and 17.5% lost cell membrane integrity. A small subpopulation (< or = 5%) of OKA-treated cells underwent primary oncotic necrosis within 6 hours. Interestingly, the caspase-3 and -7 inhibitor Z-DEVD-FMK did not inhibit or delay OKA-induced apoptosis in these cells. Our results suggest a complex apoptotic model involving 2 or more potentially parallel death pathways. Although caspase-3 and -7 cleavage occurs during apoptosis in this model, this cleavage may not independently regulate cell death in ERC-18 cells. Therefore, measurement of apoptosis in this model requires analysis of both biochemical and morphologic events. |
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Authors:
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Todd M Kolb; Seung H Chang; Myrtle A Davis |
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Publication Detail:
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Type: Journal Article; Research Support, U.S. Gov't, P.H.S. |
Journal Detail:
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Title: Toxicologic pathology Volume: 30 ISSN: 0192-6233 ISO Abbreviation: Toxicol Pathol Publication Date: 2002 Mar-Apr |
Date Detail:
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Created Date: 2002-04-12 Completed Date: 2002-04-25 Revised Date: 2009-07-01 |
Medline Journal Info:
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Nlm Unique ID: 7905907 Medline TA: Toxicol Pathol Country: United States |
Other Details:
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Languages: eng Pagination: 235-46 Citation Subset: IM |
Affiliation:
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Department of Pathology, University of Maryland, School of Medicine, Baltimore 21201, USA. |
Export Citation:
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| MeSH Terms | |
Descriptor/Qualifier:
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Animals Apoptosis / drug effects* Caspases / metabolism Cytochrome c Group / secretion Kidney Neoplasms / pathology* Okadaic Acid / toxicity* Oligopeptides / pharmacology Phosphatidylserines / metabolism Phosphorylation Rats Repressor Proteins / physiology* Talin / analysis Tumor Cells, Cultured Tumor Suppressor Proteins Vinculin / analysis |
| Grant Support | |
ID/Acronym/Agency:
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ESO5925-01/ES/NIEHS NIH HHS; ESO8157/ES/NIEHS NIH HHS; T32 ESO7263/ES/NIEHS NIH HHS |
| Chemical | |
Reg. No./Substance:
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0/Cytochrome c Group; 0/Oligopeptides; 0/Phosphatidylserines; 0/Repressor Proteins; 0/Talin; 0/Tumor Suppressor Proteins; 0/benzoylcarbonyl-aspartyl-glutamyl-valyl-aspartyl-fluoromethyl ketone; 125361-02-6/Vinculin; 169027-60-5/tuberous sclerosis complex 2 protein; 78111-17-8/Okadaic Acid; EC 3.4.22.-/Caspases |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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