title: Detection of environmental clastogens and aneugens in human fibroblasts by cytokinesis-blocked micronucleus assay associated with immunofluorescent staining of CENP-A in micronuclei.
abstract: The cytokinesis-blocked micronucleus (CBMN) assay, in combination with fluorescent in situ hybridization (FISH) of human pan-centromeric DNA probes, or with CREST antibodies that specifically stain kinetochore proteins, is widely used on several cell types. It distinguishes micronuclei containing one or several whole chromosomes, which are positively labeled (centromere positive micronucleus, C+MN, due to aneugenic effect), or acentric chromosome fragments, which are unlabeled due to the absence of centromere (centromere negative micronucleus, C-MN, due to clastogenic effect). However, the very slight level of the centromeric signals obtained with the FISH technique on primary human fibroblasts, a cell type commonly used in environmental genetic toxicology, leads to great difficulties in distinguishing C+MN and C-MN. Furthermore, the CREST technique may lead to inappropriate results particularly with regards to variations in antibody composition between patient sera. Our results show that the in vitro CBMN, in combination with immunofluorescence staining of CENP-A (centromere protein A), efficiently screens genotoxicants for their ability to induce clastogenic and/or aneugenic effects. We propose the in vitro CBMN assay in combination with immunofluorescence staining of CENP-A as a suitable tool in environmental genotoxicity testing of primary human fibroblasts.
authors.affiliation: Laboratoire de Biogénotoxicologie et Mutagenèse Environnementale (EA 1784/FR CNRS 3098 ECCOREV), Aix-Marseille Université, Faculté de Médecine, Marseille, France. laila.benameur@univmed.fr
authors.forename: Laila
authors.initials: L
authors.lastname: Benameur
authors.forename: Thierry
authors.initials: T
authors.lastname: Orsière
authors.forename: Jérôme
authors.initials: J
authors.lastname: Rose
authors.forename: Alain
authors.initials: A
authors.lastname: Botta
mesh_terms: D024422:Aneugens / Q000378:metabolism / Q000633:toxicity; D001324:Autoantigens / Q000378:metabolism*; D017675:CREST Syndrome / Q000097:blood; D000076247:Centromere Protein A; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism*; D048749:Cytokinesis; D004784:Environmental Monitoring; D004785:Environmental Pollutants / Q000378:metabolism*; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D015162:Micronucleus Tests / Q000379:methods*; D009153:Mutagens / Q000378:metabolism* / Q000633:toxicity; D015534:Trans-Activators / Q000378:metabolism
chemical_list: D024422:Aneugens; D001324:Autoantigens; C000619700:CENPA protein, human; D000076247:Centromere Protein A; D002868:Chromosomal Proteins, Non-Histone; D004785:Environmental Pollutants; D009153:Mutagens; C502810:SS18L1 protein, human; D015534:Trans-Activators