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1

Application of ionizing radiation in studies of biomarkers of individual susceptibility

Cebulska-Wasilewska A.

Nukleonika

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2005

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Vol. 50,suppl.2

3-8

EN

Human biomonitoring, as a tool to identify health risk from environmental exposures, has gained increasing interest especially in the areas of cancer risk assessment and diseases treatment. Chromosome aberrations resulting from direct DNA breakage or from inhibition of DNA repair or synthesis, measured in peripheral blood lymphocytes, have been used successfully in the assessment of health risk associated to environmental genotoxic exposures. A faster but sensitive and reliable method for detection of DNA damage, or DNA repair capacity, might be crucial to many fields from molecular epidemiology and toxicology to preventive and clinical medicine. There are reports that results of DNA measures with the use of single cell gel electrophoresis (SCGE) correlate, on the one hand, with physical measures of genotoxins, and on the other hand, with cytogenetic damage that is a biomarker associated to the alteration of the health risk. This review is based on studies in which exposure to radiation was applied as a challenging treatment and DNA damage induced and repaired was analyzed with the use of the alkaline version of SCGE assay. Results from studies on susceptibilities and repair competence carried out in various groups of exposed workers, controls, and cancer patients (more than 700 donors) show variability between donors both in a response to challenging treatment and in the efficiency of repair process. Influences of the occupational exposures and factors depending on genotypes or life style on cellular capacities are observed. Discussed results suggest that study in vitro with the challenging cells by radiation exposure and measuring, with the SCGE assay, the DNA damage before and after repair, may develop a good biomarker of the individual susceptibility to various genotoxins and exposures (environmental, occupational, therapeutic). Such a biomarker may have a potential use in a molecular epidemiology and preclinical identification.

2

Efficiency of a 252Cf source in normal or in B-10 enriched lymphocytes evaluated by SCGE assay, classical cytogenetics and FISH technique

Cebulska-Wasilewska A., Niedźwiedź W., Florjan D., Schneider K., Kopec M., Kreft A.

Nukleonika

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2001

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Vol. 46, nr 2

41-49

EN

Biological effectiveness of a californium-252 source was evaluated after irradiations in vitro of normal or pretreated cells with compound enriched in the B-10 ion (Na210B12H11SH also known as BSH) in order to check the possibility of any enhancement effect due to the process of boron neutron capture. Peripheral blood lymphocytes were used as a model for human cells. Human blood samples or isolated lymphocytes were irradiated with the isotopic source of 252Cf, at the Faculty of Physics and Nuclear Techniques at the University of Mining and Metallurgy, Kraków, (both the neutron source and the samples were placed in an "infinite" polyethylene block). DNA and chromosomal damage were studied to compare the biological effectiveness of irradiation. Single cell gel electrophoresis also known as the Comet assay was done to investigate the DNA damage. Classical cytogenetic analysis was applied to assess the frequencies of unstable aberrations (dicentrics, rings and acentric fragments). To evaluate the frequencies of stable aberrations the fluorescence in situ hybridisation (FISH) with probes for chromosomes 1, 4 (14,3% of the whole genome) was performed. Linear (or close to linear) increases with radiation doses were observed for the DNA damage and aberration frequencies in lymphocytes both untreated or pretreated with BSH. Levels of translocations evaluated for the whole genome were comparable with the frequencies of dicentrics and rings. No significant differences were detected due to radiation dose in the frequencies of sister chromatid exchanges (SCE) detected in the second mitosis. Statistically no significant differences were observed in various biological end-points between normal or boron pre-treated cells.

3

Czy dokonana metodą kometową ocena podatności osobniczej na promieniowanie i efektywności procesów naprawy indukowanych uszkodzeń DNA będzie pomocna w diagnostyce przedklinicznej

Cebulska-Wasilewska A.

Postępy Techniki Jądrowej

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2000

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vol. 43, z. 1

5-14

PL

Artykuł przedstawia rezultaty badań prowadzonych metodami cytogenetycznymi oraz testem kometowym, które pozwalają ocenić uszkodzenia DNA powstające w dowolnych typach komórek.

4

Biological efficiency of the Brookhaven Medical Research Reactor mixed neutron beam estimated from gene mutations in Tradescantia stamen hair cells assay

Cebulska-Wasilewska A., Schneider K., Capała J., Coderre J.

Nukleonika

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2000

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Vol. 45, nr 2

115-119

EN

The relative biological effectiveness of low energy neutrons for the induction of various abnormalities in Tradescantia stamen hair mutation (Trad-SH) assay was studied using two clones (T-4430 and T-02), heterozygous for flower color. Dose response relationships for gene mutations induced in somatic cells of Trad-SH were investigated after irradiation with a mixed neutron beam of the Brookhaven Medical Research Reactor (BMRR), currently used in a clinical trial of boron neutron capture therapy (BNCT) for glioblastoma. To establish the RBE (relative biological effectiveness) of the BMRR beam in the induction of various biological end-points in Tradescantia, irradiation with various doses of g-rays was also performed. After irradiation all plants were cultivated several days at Brookhaven National Laboratory (BNL), the transported to Poland for screening the biological end-points. Due to the post-exposure treatment, all plants showed high levels of lethal events and alteration of the cell cycle. Plants of clone 4430 were more reactive to post-treatment conditions, resulting in decreased blooming efficiency that affected the statistics. Slope coefficients estimated from the dose response curves for gene mutation frequencies allowed the evaluation of ranges for the maximal RBE values of the applied beam vs. g-rays as 6.0 and 5.4 for the cells of T-02 and T-4430, respectively. Estimated fraction of doses from neutrons and corresponding biological effects for the clones T-02 and T-4430 allowed to evaluate the RBE values for neutrons part in the beam as 32.3 and 45.4, respectively.

5

Application of tsh bioindicator for studying the biological efficiency of radiation

Cebulska-Wasilewska A., Rękas K., Kim J.K.

Nukleonika

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1999

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Vol. 44, nr 1

15-30

EN

The effectiveness of neutrons from a californium-252 source in the induction of various abnormalities in the Tradescantia clone 4430 stamen hair cells (TSH-assay) was studied. Special attention was paid to check whether any enhancement in effects caused by the process of boron neutron capture is visible in the cells enriched with boron ions. Two chemicals (borax and BSH) were applied to introduce boron-10 ions into cells. Inflorescences, normal or pretreated with chemicals containing boron, were irradiated in the air with neutrons from the 252 Cf source at KAERI, Taejon, Korea. To estimate the relative biological effectiveness (RBE) of the beam under the study, Tradescantia inflorescences without chemical pretreatment were irradiated with various doses of X-rays. The ranges of radiation doses used for neutrons were 0-0.1 Gy and for X-rays 0-0.5 Gy. After time needed to complete the postirradiation repair Tradescantia cuttings were transferred to Krakow, where screening of gene and lethal mutations in somatic cells of stamen hairs have been done, and dose response relationships were plotted. In two independent experimental studies an alteration of dose-response curves was observed, probably due to slight changes in the postexposure plant treatment. However, it has not resulted in the change of the maximal RBE values, which for the induction of gene mutations were estimated as 5.6 in the pilot studies, and 5.8 one year later. Inflorescences pretreated with borax and BSH responded to neutrons differently. The values of RBE have changed from 5.6 to 7.9 in the case of plants pretreated with 240 ppm of B-10 from borax, and 5.8 to 7.2 in the case of 400 ppm of B-10 from BSH. The results showed an increase, although statistically insignificant, in biological efficiency of radiation from the 252 Cf source in the samples pretreated with boron containing chemicals.

PL

Oszacowano skuteczność jonów ze źródła 252 Cf w indukowaniu mutacji genowych w komórkach pręcików Tradescantia (klonu 4430). Sprawdzono także czy proces wychwytu neutronów powoduje widoczny wzrost efektu w komórkach kwiatostanów traktowanych związkami boru przed napromieniowaniem neutronami. W doświadczeniu użyto dwa związki wprowadzające jony 10 B do komórek: boraks i BSH. Kwiatostany normalne oraz traktowane związkami boru były napromieniowane neutronami ze źródła 252 Cf w KAERI, Taejon, Korea. W celu oszacowania względnej skuteczności biologicznej (RBE) kwiatostany Tradescantia napromeniowano różnymi dawkami promieniowania rentgenowskiego. Zakresy dawek wynosiły: 0-0.1 Gy w przypadku neutronów i 0-0.5 Gy w przypadku promieniowania rentgenowskiego. Po czasie potrzebnym na naprawę popromienną rośliny przetransportowano do Krakowa, gdzie badano mutacje genowe i letalne w komórkach somatycznych, oraz wyznaczono zależności dawka-efekt. Maksymalne wartości RBE (5.6-5.8) szacowane w oparciu o współczynniki nachylenia krzywych dawka-efekt zależne były od różnych warunków po napromienieniu. W pracy przedstawiono analizę zmiany parametrów opisujących procesy komórkowe. Uzyskane rezultaty wykazują niewielki, statystycznie nieznaczący wzrost skuteczności biologicznej źródła 252 Cf w komórkach roślin traktowanych związkami zawierającymi bor. Wartości RBE zmieniały sie od 5.6 do 7.9 w przypadku roślin traktowanych boraksem (240 ppm 10B) i od 5.8 do 7.2 w przypadku BSH (400 ppm 10B).

6

DNA and chromosomal damage estimate in blood of people suspected of exposure to radiation

Cebulska-Wasilewska A., Niedźwiedź W., Nowak D., Kasper E., Wierzewska A., Wójcik A., Boużyk E.

Nukleonika

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1998

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Vol. 43, nr 1

65-72

EN

In 1996 four persons were suspected of accidental exposure to ionizing radiation. In order to estimate the absorbed doses, peripheral lymphocytes were analyzed for the presence of unstable chromosomal aberrations. Additionally, the comet assay was applied for the analysis of DNA damage. Chromosomal aberrations were scored in the first postirradiation metaphase and the absorbed doses reconstructed on the basis of a calibration curve established for lymphocytes irradiated in vitro. All absorbed doses were found to be below 1 Gy. In the comet assay, the absorbed doses were assessed on the basis of dose response relationship and calibration coefficient obtained from the comparison of two DNA damage detection systems. In the case of two persons who received the highest doses, a good agreement was found between chromosomal aberrations and the comet assay. In the case of the other persons, a high level of DNA damage observed in the comet assay did not correlate with a low level of chromosomal aberrations. In the case of one person additional analysis of the sister chromatid exchange (SCE) suggested exposure to chemicals (e.g. long medical treatment or smoking). On the basis of the obtained results we assume that, although the comet assay is sensitive to DNA damage, it is not appropriate for a low radiation dose estimate because, unlike in cytogenetic analysis, one cannot distinguish between DNA damage caused by radiation and other agents.

PL

W 1996 r. cztery osoby były podejrzane o przypadkową ekspozycję na promieniowanie jonizujące. Stosując elektroforezę pojedynczych komórek w żelu agarozowym oraz cytogenetykę klasyczną oceniono potencjalną dawkę pochłoniętego promieniowania na podstawie anlizy uszkodzeń DNA oraz aberracji chromosomowych. Dawki pochłonięte przez poszczególne osoby zostały oszacowane na podstawie krzywych kalibracyjnych wykreślonych dla limfocytów napromieniowanych in vitro. Wszystkie pochłonięte dawki nie przekraczały 1 Gy. W przypadku metody kometowej dawki oszacowano na podstawie współczynnika kalibracji uzyskanego z porównania dwu metod oceny uszkodzeń DNA. Dla dwu osób, które otrzymały najwyższe dawki, zaobserwowano bardzo dużą zgodność w ocenie pochłoniętej dawki na podstawie aberracji chromosomowych i uszkodzeń DNA. W przypadku pozostałych osób poziom uszkodzeń DNA nie był skorelowany z niską frekwencją aberracji chromosomowych. Dla jednej z tych osób dodatkowa analiza wymian chromatyd siostrzanych wskazywała na ekspozycję na czynniki chemiczne (leki, papierosy). Słaba korelacja pomiędzy niskimi dawkami oszacowanymi z wykorzystaniem tych dwu metod sugeruje ograniczenia w zastosowaniu metody kometowej w środowiskowej dozymetrii biologicznej.