CYTOKINESIS-BLOCK MICRONUCLEUS (CBMN) ASSAY/CBMN CYTOME ASSAY IN HUMAN LYMPHOCYTES AFTER IN VITRO IRRADIATION AND ITS USE IN BIODOSIMETRY

Micromeria pulegium (Rochel) Benth. is an endemic species of Lamiacea family that includes frequently used plants in culinary and folk medicine. As cytotoxic potential of some species of Micromeria genus has been confirmed, this study aimed to test unknown antiproliferative and genotoxic potential of M. pulegium, endemic bh species, aqueous leaf extract in normal (human lymphocytes) and cancer (human melanoma GR-M) cells in order to protect small populations of native M. pulegium populations or promote its controlled micropropagation or cultivation. Cytokinesis-block micronucleus cytome assay was applied for human lymphocyte cultures, while trypan blue exclusion assay was used for evaluation of cytotoxicity in human GR-M melanoma cells. Results demonstrate no genotoxic effects up to concentration of 0.2 mg/ml in human lymphocyte in vitro but significant reduction of cell viability in human GR-M melanoma cell line cultures treated with 0.3 mg/ml of Micromeria extract.

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Cytokinesis-Block Micronucleus Cytome Assay Evolution into a More Comprehensive Method to Measure Chromosomal Instability

Genes ◽

10.3390/genes11101203 ◽

2020 ◽

Vol 11 (10) ◽

pp. 1203

Author(s):

Michael Fenech

Keyword(s):

Chromosomal Instability ◽

Ex Vivo ◽

Chromosome Breakage ◽

Comprehensive Method ◽

Cbmn Assay ◽

Dividing Cells ◽

Chromosome Fragments ◽

Painting Probes ◽

Cytome Assay

This review describes the cytokinesis-block micronucleus (CBMN) cytome assay and its evolution into a molecular cytogenetic method of chromosomal instability (CIN). Micronuclei (MNi) originate from whole chromosomes or chromosome fragments that fail to segregate to the poles of the cell during mitosis. These lagging chromosomes are excluded from the daughter nuclei and are enveloped in their own membrane to form MNi. The CBMN assay was developed to allow MNi to be scored exclusively in once-divided binucleated cells, which enables accurate measurement of chromosome breakage or loss without confounding by non-dividing cells that cannot express MNi. The CBMN assay can be applied to cell lines in vitro and cells such as lymphocytes that can be stimulated to divide ex vivo. In the CBMN assay, other CIN biomarkers such as nucleoplasmic bridges (NPBs) and nuclear buds (NBUDs) are also measured. Use of centromere, telomere, and chromosome painting probes provides further insights into the mechanisms through which MNi, NPBs and NBUDs originate. Measurement of MNi is also important because entrapment within a micronucleus may cause chromosomes to shatter and, after nuclear reintegration, become rearranged. Additionally, leakage of DNA from MNi can stimulate inflammation via the cyclic GMP-AMP Synthase—Stimulator of Interferon Genes (cGAS-STING) DNA sensing mechanism of the innate immune system.

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Estimating genotoxic effects of anticancer drugs using cytokinesis-block micronucleus assay on human lymphocytes

Fundamental and Clinical Medicine ◽

10.23946/2500-0764-2019-4-3-95-101 ◽

2019 ◽

Vol 4 (3) ◽

pp. 95-101

Author(s):

V. I. Minina ◽

V. Yu. Buslaev

Keyword(s):

Anticancer Drugs ◽

Human Lymphocytes ◽

In Situ Hybridisation ◽

Micronucleus Assay ◽

Cochrane Library ◽

Dependent Manner ◽

Fluorescence In Situ Hybridisation ◽

Genotoxic Effects ◽

Cbmn Assay

Here we review the current experience of using cytokinesis-block micronucleus (CBMN) assay on cultures of human lymphocytes to evaluate genotoxic effects of anticancer drugs. Having performed search in PubMed, Scopus, Web of Science, TOXLINE, and the Cochrane Library, we identified a total of 172 relevant studies. Out of them, 89 were conducted in vitro, and 41 were published within the last decade. The mentioned studies concordantly demonstrated a significant increase in micronuclei, protrusions, nucleoplasmic bridges, and a decrease in proliferation in cells treated with anticancer drugs in a time- and dose-dependent manner. Notably, the results of CBMN assay are consistent with the data obtained from other cytogenetic techniques (comet assay, chromosomal aberration analysis, analysis of mutations in housekeeping genes, and fluorescence in situ hybridisation). Conclusion. CBMN assay permits a reliable evaluation of the mutagenic effects related to anticancer drugs.

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Jatrophane Diterpenoids With Protective Effect on Human Lymphocytes DNA

Natural Product Communications ◽

10.1177/1934578x19848168 ◽

2019 ◽

Vol 14 (5) ◽

pp. 1934578X1984816

Author(s):

Gordana Krstić ◽

Milka Jadranin ◽

Miroslava Stanković ◽

Ivana Aljančić ◽

Ljubodrag Vujisić ◽

...

Keyword(s):

Protective Effect ◽

Human Lymphocytes ◽

Structural Features ◽

Positive Control ◽

Sar Studies ◽

Structure Activity ◽

Minimal Dose ◽

Cbmn Assay ◽

Sar Study

Two sets of structurally different jatrophanes (1-11 and 13-16), jatrophane 12, and latex extract of 2 Euphorbia species (17 and 18) were tested for in vitro protective effect against chromosome aberrations in peripheral human lymphocytes using the cytokinesis-block micronucleus (CBMN) assay. Jatrophanes 1-6 in minimal doses of 1 µg/mL prominently decreased micronuclei (MN) frequency in the range 44.86% to 34.29% and manifested considerable protective effect. From the other set of jatrophanes, 13 in the same minimal dose notably decreased MN frequency by 31.05%, while extracts 17 and 18 at a concentration of 4 µg/mL remarkably decreased the frequency of MN by 37.94% and 36.12%, respectively. Jatrophanes 12, 14, and 16 showed moderate protection, while 7-11 and 15 were less active than positive control. The structure-activity relationship (SAR) studies of the tested jatrophanes (1-16) indicated the favorable position of benzoate at C-8 or C-9 (3, 4, and 13) and a preference of isobutanoyloxy group at C-3 (1-3) rather than propanoyloxy at the same position (4-6) for pronounced protective effect on human lymphocytes DNA. In a previous SAR study on 11 jatrophanes (1, 3-8, and 13-16), the same structural features in 3, 4, and 13 influenced powerful inhibition of P-gp, while growth inhibition of cancer cells was more than doubled in 1 (isobutanoyloxy group at C-3) compared to 6 (propanoyloxy at C-3).

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