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West African Journal of Pharmacology and Drug Research

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Simultaneous Determination of Mutagenicity and Toxicity of Chemicals to Chinese Hamster Cells Using Delayed Toxicity Method

AO Asita, MG Clare, SK Obuya, MR Budda, CK Atterwill

Abstract


Induction of chromosome damage in mammalian cells by chemicals generally requires DNA replication and this leads to delayed toxic effect, i.e. cell death. A demonstration of cytotoxicity is required (measurement of cell number, culture confluency and inhibition of mitotic index) for in vitro cytogenetic assays. The study therefore investigated whether delayed cytotoxicity can be used to simultaneously predict mutagenicity and cytotoxicty. Chinese hamster lung cells were cultured in 96-well microtitre plates, incubated until they were actively dividing, then exposed for 3 hours to Methylmethane sulfonate (MMS), Cyclophosphamide (CPH), Dimethyl sulfoxide (DMSO) or 3-Methylcholanthrene (3-MC). Cytotoxicity was assessed indirectly at intervals from 0 to 72 hours after exposure, using MTT uptake and reduction, with optical density values from the purple formazan product being expressed relative to the concurrent solvent control. The results showed experimental variability from one time point to the next, which may mask subtle changes such as cell cycle delay. Toxicity profile for tests with MMS or CPA changed as time passed. Toxicity profile for DMSO tests remained similar throughout, while that for the relatively non-toxic 3-MC treated cells was not definitive because any effects were too subtle for the test system to detect. This approach appears to be suitable for detection of substances that damage chromosomes and are evidently toxic.

Key Words: Chromosome damage, delayed toxicity, screen for genotoxins

Résumé


L'induction des lesons chromosomale chez les mamiferes par les substance chimique generalement necesite la replication de l'AND et ceci genere un effet toxique retarde i.e la mort de la cellule .la mesure de la cytotoxicite est requise.( la mesure du nombre de cellule, la confluence de la culture et l'inhibation de l'index mitotique)de l'assy cytogenique invitro. L'etude a ainsi invitigue si la cytotoxicite retarde peu etre simultanement utilize pour predire la mutagenicite et la cytotoxicite. Les cellule pulmonaire du Chinese hamster onete cultive dans 96 plaquettes bien microtitre. incube jusqu'au point ou elle se divise activement,ensuite expose pendant 3 heures au sulfonate de methylmethane (SMM), cyclophosphamide (CPH), dimethyl sulfoxide(DMSO)ou3-methylcholanthre(3-MC). La cytotoxicite a ete mesure indirectement a des intervals de 0 a 72heurs après exposition utilisant l'absorption et la reduction du MTT,avec la densite optique des valeurs du produitfromazan marron etant relativement exprime au olvent de controlles resutats ont demontre experimentalement la variabilite d'un temps de point a l'autre, qui peut masque des changement subtile telque le retardement du cycle cellulaire. Le profile de test toxicite avec SMM ou CPA au fur et a mesure que le temps passé.le profile de test de toxicite pour le DMSO restait similair tout au long alors celui des cellule traite au 3-MC relativement non toxiquen'etait pas definitive parceque les effet etaient trop subtile pour etre detecte par ce systeme de test. Cette approchhe aparait tres approprie pour detecte les substances qui endomage les chromosome et ils sont evidement tocique.

Mot cles : Lesion chromosomique, toxicite retarde, depistage de genotoxine

West Afr. J. Pharmacol. Drug Res. Vol.19 (1&2) 2003: 5-8



http://dx.doi.org/10.4314/wajpdr.v19i1.14725
AJOL African Journals Online