Cytokinesis-block micronucleus assay in human glioma cells exposed to radiation

english

2004

volume 23, issue 3

Radiotherapy    Pathology    Tumor Cells, Cultured   Micronucleus Tests 

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Biological tests are efticient in reflecting the biological influences of several types of generally harmful exposures. The micronucleus assay is widelyused in genotoxicity studies or studies on genomic damage in general. Wepresent methodological aspects of cytokinesis-block micronucleus assay performed in human gliomas irradiated in vitro. Eight human glioblastoma cell lines obtained from DSMZ (Deutsche Sammlung von Mikroorganismen and Zellkulturen GmbH, Germany) were gamma-irradiated (60Co) over a dose range of 0-10 Gy. Cytokinesis-block micronucleus assay was performed to quantitate cytogenetic damage. The cells were fixed directly on dishes, stained with fluorochrome DAPI and evaluated under fluorescent and phase contrast microscope. The micronucleus frequency was expressed as a micronuclei (MN) perbinucleated cell (BNC) ratio, calculated after scoring at least 100 BNC perdish. The frequency of spontaneous MN ranged from 0.17 to 0.613 (mean: 0.29 0.14). After irradiation increase of MN frequency in the range of 0.312 - 2.241 (mean: 0.98 +- 0.68) was found at 10 Gy. Gliomas are extremely heterogenous in regard to cytogenetic effects of irradiation, as shown in thisstudy by cytokinesis-block micronucleus assay. This test is easily performed on irradiated glioma cell lines and can assist in determining their radiosensitivity. However, in order to obtain reliable and reproducible results, precise criteria for MN scoring must be strictly followed. Simultaneous use of fluorescent and phase contrast equipment improves imaging of morphological details and can further optimize MN scoring.

16.09.2009