Document Detail


Enrichment of non-synchronized cells in the G1, S and G2 phases of the cell cycle for the study of apoptosis.
MedLine Citation:
PMID:  16765323     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
The susceptibility of cells to apoptosis induction is deeply influenced by their position in the cell cycle. Unfortunately, however, current methods for the enrichment of cells in defined phases of the cell cycle are mostly based on the synchronization of cells by agents or conditions that are intrinsically toxic and induce apoptosis on their own. We developed a novel procedure for the purification of cells in distinct phases of the cell cycle. This method is based on the stable transfection of cells with a chimeric protein made up by histone H2B and green fluorescent protein (GFP). Cytofluorometric purification of cells defined by their size and their H2B-GFP-dependent fluorescence (which reflects chromatin and hence DNA content) allowed for the efficient separation of diploid and tetraploid cells in the fluorescence-activated cell sorter (FACS). Moreover, when applied to diploid cells, this method allowed for the enrichment of live, functional cells in the G1, S and G2 phases of the cell cycle. FACS-purified cells were viable and readily resumed the cell cycle upon reculture. While staurosporine was equally toxic for cells in any phase of the cell cycle, camptothecin was particularly toxic for cells in the S phase. Moreover, BAY11-7082, a specific inhibitor of the IKK complex required for NF-kappaB activation, exhibited a particular cell cycle-specific profile of toxicity (G2>S>G1). These results delineate a novel procedure for studying the intersection between cell cycle regulation and cell death mechanisms.
Authors:
Arnaud Coquelle; Shahul Mouhamad; Marie O Pequignot; Thorsten Braun; Gabrielle Carvalho; Sonia Vivet; Didier Métivier; Maria Castedo; Guido Kroemer
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't     Date:  2006-04-29
Journal Detail:
Title:  Biochemical pharmacology     Volume:  72     ISSN:  0006-2952     ISO Abbreviation:  Biochem. Pharmacol.     Publication Date:  2006 Nov 
Date Detail:
Created Date:  2006-11-07     Completed Date:  2007-01-04     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  0101032     Medline TA:  Biochem Pharmacol     Country:  England    
Other Details:
Languages:  eng     Pagination:  1396-404     Citation Subset:  IM    
Affiliation:
Centre National de la Recherche Scientifique, UMR8125, Institut Gustave Roussy, 39 rue Camille-Desmoulins, F-94805 Villejuif, France.
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MeSH Terms
Descriptor/Qualifier:
Apoptosis* / drug effects
Bromodeoxyuridine / metabolism
Camptothecin / pharmacology
Cell Line, Tumor
Cell Survival / drug effects
Colonic Neoplasms / genetics,  metabolism,  pathology*
Dose-Response Relationship, Drug
Enzyme Inhibitors / pharmacology
Flow Cytometry / methods*
Fluorescent Antibody Technique
Humans
Interphase* / drug effects
Nitriles / pharmacology
Ploidies
Staurosporine / pharmacology
Sulfones / pharmacology
Chemical
Reg. No./Substance:
0/3-(4-methylphenylsulfonyl)-2-propenenitrile; 0/Enzyme Inhibitors; 0/Nitriles; 0/Sulfones; 59-14-3/Bromodeoxyuridine; 62996-74-1/Staurosporine; 7689-03-4/Camptothecin

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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