Analysis of Radiation Dose-Response Curve Obtained with Cytokinesis Block Micronucleus Assay (original) (raw)
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Mutation Research-genetic Toxicology and Environmental Mutagenesis, 2010
There are conflicting data regarding the effect of culturing time of human peripheral blood lymphocytes on the yield of chromosomal aberrations induced by sparsely ionising radiation in the G 0 phase of the cell cycle. While some authors find that the yield of aberrations does not change with time, others find increased frequencies of aberrations with harvesting time. The reasons for the conflicting results are not known, but the majority of studies were performed with lymphocytes of a single donor collected at one time point. We performed a study to verify if individual variability could be a confounding factor. As a positive control, lymphocytes were also exposed to high LET radiation (neutrons and alpha-rays), where an effect of harvesting time on the level of damage is expected to be seen.
2011
Aim: Cytokinesis-block micronucleus (CBMN) assay and its comprehensive variant CBMN cytome assay are cytogenetic methods. CBMN is based on assessment of micronuclei in nucleated cells that have completed only one nuclear division. Besides micronuclei, CBMN cytome assay analyzes additional genotoxic (nucleoplasmic bridges and nuclear buds), cytostatic (nuclear division index), and cytotoxic (amount of necrotic and apoptotic cells) parameters. The aim of this study is to evaluate these parameters in human blood lymphocytes after in vitro irradiation and to assess its contribution to biodosimetry. Material and methods: Human blood from 6 donors was in vitro irradiated by 0, 1, 2, 3, 4, or 5 Gy and cultivated for 72 hours. Blood lymphocytes were stimulated with phytohemagglutinin and their cytokinesis was blocked by cytochalasin B. After cultivation, cultures were hypotonically treated, dropped onto glass slides and stained with Giemsa. Slides were evaluated by microscope. Results: We observed significantly increased amount of micronuclei, nucleoplasmic bridges, and nuclear buds measured in binucleated cells, significantly increased amount of micronuclei measured in mononucleated cells and significantly decreased nuclear division index after irradiation by 1, 2, 3, 4, and 5 Gy. Amount of death cells (apoptotic and necrotic) significantly increased after irradiation by 4 and 5 Gy. Conclusion: Although all parameters assessed by CBMN cytome assay have biodosimetric potential, practically feasible is only evaluation of micronuclei in binucleated cells. This parameter was used to construct in vitro linear-quadratic dose-response calibration curve which could be used as a biodosimetric tool for triage of radiation casualties.
Teratogenesis, Carcinogenesis, and Mutagenesis, 1994
Usingthecytokinesis-blocktechnique, lymphocytesfrom healthy volunteers (n = 9) were evaluated for 1) the radiation dose-response curve for micronuclei (MN) expression; 2) technique variables on the yield of MN; and 3) the shortest lymphocyte incubation time required for the MN assay. We found that the best fitting of relationships between increasing MN production and increasing irradiation dose (U. 0 GJ) was the linear-quadratic model as expressed by the yield equation Y = C + aD + PD (P = 0.0003). When lymphocytes were irradiated in vitro with 2.0 Gy and harvested at various time intervals, MN increased during the entire 84 hr culture time. The radiation caused a division delay in lymphocyte as indicated by an increased frequency of mononucleated cells and a decreased number of mitotic indices. The data showed that a shortened culture time (60 hr) for the MN assay is possible and that binucleated cells with 2 3 MN were found only in cells irradiated at 2 2. 0 Gy. These findings suggest that scoring of MN in lymphocytes may be a practical biological dosimeter for the rapid screening of accidental radiation exposure victims, especially when their clinical manifestations are not obvious.
Mutation Research/Fundamental and …, 1986
Study of the different aspects of protection against the exposure of ionizing radiation has always been an active area of research. High cost and toxicity of radioprotective drugs have limited their use. So, search for new drugs with a high degree of protection and lower cost and side effects seem a necessity. In this study radioprotective effect of aqueous as well as alcoholic extracts of the Mann of Cotoneaster nummularia( Shirkhesht), regarding their high accessibility and possibly low side effects, against 2 Gy Gamma irradiation, was analyzed using micronucleus assay on bone marrow cells of male mice (Balb/c). Different doses of 250, 500, 1000 mg/kg/BW for aqueous and 3750, 7500, 15000 mg/kg/BW for alcoholic extract of Shirkhesht were administered IP for five constitutive days prior to 2 Gy gamma irradiation. The result compared with the known radioprotective effect of vitamin E after the same treatment schedule. High frequency of micronucleus was observed in non treated gamma-exposed mice, which represented the clastogenic effect of irradiation. Vitamin E, aqueous and alcoholic extracts of Shirkhesht treated mice represented a 5.56, 3.32 and 2.1 times decrease in the gamma-induced micronucleus frequency respectively. The data suggest a radioprotective effect of shirhkesht compared to vitamin E.
The quantification of micronuclei in lymphocytes is a method for estimating of individual exposure to ionizing radiation, which can be used complementary to physical dosimetry or when this later cannot be performed. In this work, the quantification of micronuclei was carried out using cytogenetic analyzes of peripheral blood samples from 5 patients with cervical uterine cancer following radiotherapy, in order to evaluate the absorbed dose as a result of exposure to 60 Co source. From each patient, blood samples were collected in three phases of the treatment: before irradiation, 24 h after receiving 0.08 Gy and 1.8 Gy, respectively. A good agreement was obtained between doses estimated by calibration curve of dose versus the frequencies of micronuclei and the values previously planned to the radiotherapy. The results presented in this report emphasizes biological dosimetry, based on the quantification of micronuclei in lymphocytes from peripheral blood, as an important methodology f...
Radiation Oncology Investigations, 1993
We obtained peripheral lymphocytes from both healthy donors (n = 7) and cancer patients (n = 14) for the cytokinesis-blocked micronucleus (MN) assay. Lymphocytes were irradiated with 13'Cs to 2 Gy. Cytochalasin B (6 &ml) was added to the incubation at 44 hr and the cells were harvested a t 72 hr. We found that compared to healthy donors, lymphocytes of cancer patients revealed no significant difference in the MN baseline level (P = 0.86). Apparently DNA damage manifested as MN in the lymphocytes of cancer patients is not more severe than that of healthy donors. In addition, 2 Gy in vitro irradiation of lymphocytes caused no difference in the MN formation between cancer patients and healthy donors (P = 0.9). Radiation induced cell cycle delay as evaluated by the number of mononucleate, binucleate, and cells with >2 nuclei and by the mitotic indices showed similar results in both populations. However, the difference for MN frequency in lymphocytes before and after 2 Gy in vitro irradiation was highly significant for both cancer patients and healthy donors (P = 0.0001). These studies in general indicate that the MN frequency in cytochalasin B-blocked human lymphocytes has the potential of serving as a quantitative biological dosimeter for the screening of radiation exposure to either healthy individuals or cancer patients.
REVIEW The micronucleus assay as a biological dosimeter of in vivo ionising radiation exposure
2010
Biological dosimetry, based on the analysis of micronuclei (MN) in the cytokinesis-block micronucleus (CBMN) assay can be used as an alternative method for scoring dicentric chromosomes in the field of radiation protection. Biological dosimetry or Biodosimetry, is mainly performed, in addition to physical dosimetry, with the aim of individual dose assessment. Many studies have shown that the number of radiation-induced MN is strongly correlated with dose and quality of radiation. The CBMN assay has become, in the last years, a thoroughly validated and standardised technique to evaluate in vivo radiation exposure of occupational, medical and accidentally ex-posed individuals. Compared to the gold standard, the dicentric assay, the CBMN assay has the important advantage of allowing economical, easy and quick analysis.
The present state and perspectives of micronucleus assay in radiation protection. A review
International Journal of Radiation Applications and Instrumentation. Part A. Applied Radiation and Isotopes, 1987
The occurrence of micronuclei proved to be a sensitive biological indicator of clastogenic effects of many chemical and physical agents. The possibility of using the micronucleus technique in radiation protection as an alternative to the traditional chromosomal analysis has recently been followed with increasing interest. This review outlines the main biological and practical aspects of the micronucleus assay and discusses its potential to serve as a rapid and reliable measure of radiation overexposures and
Mutation research, 1991
To evaluate the application of the cytokinesis-block (CB) micronucleus (MN) assay as a biological dosimeter following in vivo exposure to ionising radiation we determined the micronucleus frequency in spleen and peripheral blood lymphocytes of the mouse, serially, for 14 days following acute whole-body irradiation. The baseline MN frequency of spleen lymphocytes (7.86 +/- 0.68, mean +/- 1 SD) was significantly (p less than 0.001) elevated when compared to that for peripheral blood lymphocytes (4.10 +/- 0.53). Immediately after irradiation there was a substantial dose-related increase in MN, but the MN frequencies in spleen lymphocytes (120.2 +/- 9.4 for 1 Gy; 409.5 +/- 38.4 for 2 Gy) were significantly (p less than 0.009) elevated compared to those in peripheral blood lymphocytes (78.0 +/- 7.0 for 1 Gy; 200.2 +/- 10.9 for 2 Gy). During the 14 days after irradiation, the MN frequency in spleen lymphocytes declined gradually to approximately half of the value observed immediately afte...
Establishment of a dose-response curve for X-ray-induced micronuclei in human lymphocytes
Genome Integrity, 2016
the recent past, additional assays have been worked out and validated including the now well-established cytokinesis-block micronucleus (CBMN) assay. Even though micronuclei (MN) can arise from exposure to a variety of clastogenic agents and are not necessarily radiation specific, ionizing radiation is considered as a clastogen and it is an efficient inducer of MN. The CBMN assay is established as a very reliable, thoroughly validated and is a standardized technique in the field of radiation biology to assess in vivo radiation exposure of occupational, medical, and accidentally exposed individuals and to determine individual in vitro radiosensitivity or cancer susceptibility. [2-4] To improve our cytogenetic laboratory capability as the National Cytogenetic Biodosimetry Laboratory in Indonesia, we have established our own dose-response standard curves of unstable chromosome aberrations for gamma radiation [5] and X-rays. [6] In vitro MN test with CBMN of human peripheral blood lymphocytes has been used extensively to study chromosomal damage induced by ionizing radiation or chemicals. [7] Because radiation-induced MN showed a radiation dose and quality dependence, MN can be used as a biological dosimeter for radiation protection purposes [8] and MN assay has been recommended by