The human lymphocyte micronucleus assay: a comparison between whole-blood and separated-lymphocyte cultures

Migliore, Lucia; Nieri, M.; Amodio, S.; Loprieno, N.

doi:10.1016/0165-7992(89)90041-9

Cited by 45 publications

(5 citation statements)

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“…For each donor, one series of cultures was prepared with two parallel cultures (`duplicates') for every concentration of mutagen tested. Whole blood cultures were done as recommended by Migliore et al (1989). For each culture, 0.8 mL of heparinized blood samples was added to 8 mL of RPMI-1640 medium (Sigma Chemical Co., St. Louis, MO, USA) containing 10% fetal calf serum, 1% penicillin/streptomycin and 80 μL of phytohemaglutinin (10 μL/mL) (PHA, Gibco, New Zealand).…”

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confidence: 99%

“…This suggestion was based on a previous study that mincronucleus induction occurred exclusively in binucleated cells and was related to DNA breaks. Aneugenic drugs such as diethylstilbestrol, griseofulvin and vincristine sulphate increase frequencies of micronuclei in mononucleated and binucleated cells, whereas clastogens such as mitomycin C, bleomycin and doxorubicin increase the frequency of micronuclei only in binucleated cells (Migliore et al , 1989; Rosefort et al , 2004). …”

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Micronuclei induced by reverse transcriptase inhibitors in mononucleated and binucleated cells as assessed by the cytokinesis-block micronucleus assay

et al. 2010

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This study evaluated the clastogenic and/or aneugenic potential of three nucleoside reverse transcriptase inhibitors (zidovudine - AZT, lamivudine - 3TC and stavudine - d4T) using the cytokinesis-block micronucleus (CBMN) assay in human lymphocyte cultures. All three inhibitors produced a positive response when tested in binucleated cells. The genotoxicity of AZT and 3TC was restricted to binucleated cells since there was no significant increase in the frequency of micronuclei in mononucleated cells. This finding indicated that AZT and 3TC caused chromosomal breakage and that their genotoxicity was related to a clastogenic action. In addition to the positive response observed with d4T in binucleated cells, this drug also increased the frequency of micronuclei in mononucleated cells, indicating clastogenic and aneugenic actions. Since the structural differences between AZT and 3TC and AZT and d4T involve the 3' position in the 2'-deoxyribonucleoside and in an unsaturated 2',3',dideoxyribose, respectively, we suggest that an unsaturated 2', 3', dideoxyribose is responsible for the clastogenic and aneugenic actions of d4T.

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Micronuclei induced by reverse transcriptase inhibitors in mononucleated and binucleated cells as assessed by the cytokinesis-block micronucleus assay

et al. 2010

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“…Our results suggest that cyt B treatment interferes with chromosome missegregation in human lymphocytes treated in vitro with spindle poison (as first suggested by Eastmond and Tucker, 1989;Migliore et al, 1989). The observed 'cyt B effects' are likely to be explained if it is assumed that in cytokinesis-blocked cells the absence of the actin ring interferes with anaphase-B, leading to a shorter distance between the poles, as suggested by Norppa et al (1993), Surrallés et al (1996) and Falck et al (1997) and measured by Cimini et al (1997) in human fibroblasts.…”

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confidence: 66%

Effect of cytochalasin B on the induction of chromosome missegregation by colchicine at low concentrations in human lymphocytes

Minissi

Gustavino²,

Degrassi³

et al. 1999

Mutagenesis

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The aim of the present work was to investigate the possible interference of cytochalasin B (cyt B) with low concentration treatment with colchicine in the induction of chromosome/chromatid loss and micronuclei in human lymphocytes mitotically activated in vitro. Thus, cells from a single female donor were treated with colchicine (10 or 25 nM, from 24 h after PHA addition to fixation at 66 h) either in the presence or absence of cyt B. Single lagging chromosomes/chromatids were scored in bipolar ana-telophases and greater damage (disrupted and c-anaphases) was scored in cells at anaphase. Micronuclei were scored in the first 4000 nuclei observed in both cyt B-treated (in mononucleate and binucleate cells) and untreated cultures. With the same criterion, FISH analysis was performed on 2000 nuclei where chromosome 7 and 11 centromeric DNA probes were used in pairs. Our results showed that: (i) the frequency of laggards and of micronuclei increased with colchicine concentration but in the presence of cyt B there was a lower frequency of both (with a mean reduction of 49%); (ii) FISH analysis showed a colchicine concentration-dependent increase in nuclei with three spots for chromosome 7; (iii) a colchicine concentration-dependent increase in tetraploid cells was observed. This increase was particularly remarkable (5-fold) in cells grown in the presence of cyt B compared with cyt B-untreated cells. The observed 'cyt B effects' can be explained if it is assumed that in cytokinesis-blocked cells there is a shorter distance between the poles. As a consequence: (i) laggards would be engulfed in the nearest daughter nucleus with a consequent lower induction of micronuclei; (ii) segregating sister chromatids in heavily impaired anaphases would not travel a sufficient distance to give rise to two daughter nuclei, leading to an increased frequency of polyploid nuclei.

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“…contribution of dividing binucleate cells, especially when tested for non-disjunction. However, in human lymphocyte cultures References treated with 3 µg/ml cytochalasin B it was demonstrated (Falck et al, 1997) (Migliore et al, 1989), the evaluation of MN formation in of metabolic activation on the genotoxic and cytotoxic activity Eur. J. Med.…”

Section: Discussionmentioning

confidence: 99%

Evaluation and characterization of micronuclei induced by the antitumour agent ASE [3beta-hydroxy-13alpha-amino-13,17-seco-5alpha-androstan-17-oic-13,17-lactam-p-bis(2-chloroethyl)amino phenylacetate] in human lymphocyte cultures

et al. 2000

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increased lifespan over controls in the treatment of L1210 N.A.Demopoulos leukemia in mice and a 150% increased lifespan in the treatment of P388 leukemia (Wampler and Catsoulacos, 1977).Division of Genetics, Cell and Developmental Biology, Department of Moreover, ASE has given good activity against B 16 melanoma Biology, University of Patras, 26500 Patras, Greece in C 57 b 1 mice and T 8 Guerrin tumors in rats (Catsoulacos and 3β-Hydroxy-13α-amino-13,17-seco-5α-androstan-17- Wampler, 1982). In addition, ASE alone or in combination oic-13,17-lactam-p-bis(2-chloroethyl)amino phenylacetate with a different modified steroid, the homo-aza-steroidal ester (ASE) is a homo-aza-steroidal ester of p-bis(2-chloroethyl)of o-bis(2-chloroethyl)aminobenzoic acid, proved to exert amino phenyl acetic acid and has been shown to display enhanced antitumor activity in a synergistic manner antineoplastic, mutagenic and genotoxic activity. In the (Papageorgiou et al., 1999). It is noteworthy that although present study an effort has been made to evaluate the ASE exerts about the same antineoplastic activity as ability of ASE to induce micronuclei (MN) in human chlorambucil, it is less toxic with a more prolonged action lymphocytes treated in vitro using the cytokinesis-block (Stevenson and Patel, 1973; Catsoulacos et al., 1978). assay. Lympocytes were treated with different concentraAdditionally, ASE has shown mutagenic properties. It was tions of ASE (0.1, 0.25, 0.5, 1, 2.5, 5, 10 and 20 µg/ml) at found to be positive in strains TA1535 and TA100 as well as two different cell culture times, 21 and 41 h after culture in strain TA102 with and without metabolic activation in the initiation. ASE treatment lasted until cell harvest, for Salmonella/microsome system (Athanasiou and Arzimanoglou, 51 and 31 h, respectively. Two types of cultures were used, 1986). The compound has also been shown to induce sister whole blood and isolated lymphocyte cultures. The content chromatid exchange in human lymphocytes (Mourelatos et al., of induced MN was identified by FISH analysis, using an 1987) as well as in CHO cells (Athanasiou et al., 1983). ASE α-satellite DNA probe, in binucleate cells. Our results was also determined to decrease protein synthesis rate in suggest that ASE is capable of increasing MN frequencies ovaries of Drosophila melanogaster (Stephanou et al., 1991). in human lymphocytes under both culture conditions. ThisFinally, ASE was identified as displaying genotoxic, cytostatic increase is related to the concentration in a linear doseand antineoplastic properties (Nikolaropoulos et al., 1997). dependent manner and is also dependent on the duration Human lymphocytes cultured in vitro have been widely of treatment. FISH analysis has shown that the induced used as an appropriate system to identify the genetic activity MN resulted mainly from breakage events. Additionally, of various chemical compounds. They are a more or less a weak aneugenic effect was found at the higher conhomogeneous cell population, being all in the G 0 phase, an...

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The human lymphocyte micronucleus assay: a comparison between whole-blood and separated-lymphocyte cultures

Cited by 45 publications

References 5 publications

Micronuclei induced by reverse transcriptase inhibitors in mononucleated and binucleated cells as assessed by the cytokinesis-block micronucleus assay

Micronuclei induced by reverse transcriptase inhibitors in mononucleated and binucleated cells as assessed by the cytokinesis-block micronucleus assay

Effect of cytochalasin B on the induction of chromosome missegregation by colchicine at low concentrations in human lymphocytes

Evaluation and characterization of micronuclei induced by the antitumour agent ASE [3beta-hydroxy-13alpha-amino-13,17-seco-5alpha-androstan-17-oic-13,17-lactam-p-bis(2-chloroethyl)amino phenylacetate] in human lymphocyte cultures

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