A decrease in the level of unscheduled DNA synthesis in preparations of irradiated rat hepatocyte nuclei under the effect of polyamines

The results obtained show the possibility of polyamine (spermine and spermidine) utilization to stabilize the nuclear chromatin and to protect it against nuclease degradation in the course of isolation of liver cell nuclei. Besides, the inhibition of the unscheduled DNA synthesis induced by nuclease incision and by gamma- or UV-radiation was demonstrated in the presence of polyamines.     

Study of a mtDNA deletion in BALB/c mice exposed to ionizing radiation

A novel large mtDNA deletion of 5914 bp was detected in mice exposed to X-radiation. The regions flanking the deleted fragment were characterized by the method of sequencing. The possibility of using a minimum sample of the mouse auricle tissue for detecting mtDNA deletions in the same animals at different postradiation times is demonstrated.     

Genome instability in the F1-progeny of mice irradiated by ionizing radiation as determined by micronucleus assay

The F1-progeny of BALB/c male mice chronically exposed to low-dose gamma-radiation (0.1; 0.25 and 0.5 Gy; dose rate 0.01 Gy/day) as well as the F1-progeny of females exposed to acute X-radiation (0.5; 1.0 and 2.0 Gy; dose rate 0.1 Gy/min) shown the significant elevated micronuclei frequencies in bone marrow erythrocytes, as compared to the F1-progeny of unirradiated males and females. The increase in the micronuclei frequency in the F1-progeny was determined by the dose of irradiation of parents. The values of elevated micronuclei frequency in the F1-progeny of chronically irradiated males and acutely irradiated females for a dose of 0.5 Gy were comparable. The micronuclei frequencies in the F1-progeny of irradiated females and males for this dose were in 1.5 and in 1.6 times higher than ones in the F1-progeny of unirradiated mice correspondingly. The results suggest the possibility of transfer of genome instability from irradiated parents to the somatic cells of the F1-progeny via non-lethally damaged germ cells of parents.     

Computer analysis of band variability in DNA fingerprints

Through the example of the distribution of PCR products DNA matrices of mouse tail tissue, a method of comparative analysis of DNA fingerprints is described. The PCR products were obtained using a 20-mer random primer flanking the Atp1b2 locus on mouse chromosome 11. A software program was designed that permits the simplification of comparison of DNA fragments variability or polymorphism detected on electrophoregrams from different individuals.     

Delayed and transgenerational molecular and genetic effects of prolonged influence of ionizing radiation in nuclear plant workers

Genome variability and changes in immune homeostasis, induced in man in the course of long-term industrial contact with ionizing radiation (IR) sources were studied by using unique biomaterials stored in the Radiobiological Repository for Human Tissues at the Southern Urals Biophysics Institute, FMBA. The biomaterials, peripheral blood samples and blood DNA were obtained from the "Mayak" PA employers occupationally exposed to prolonged external gamma-radiation and/or internal alpha-radiation from incorporated 239Pu in a wide range of accumulated doses. A significant increase in the polymorphism of microsatellite-associated peripheral blood DNA repeats was revealed in a group of persons with accumulated doses of external gamma-radiation above 2.0 Gy, as well as in the descendants of parents with preconceptive doses of higher than 2.0 Gy. In persons whose parents had a preconceptive dose above 2.0 Gy, an increase in the gene p53 mutation rate was observed, and descendants of persons with dose of 3.0 Gy and higher showed mtDNA heteroplasmy, regardless of the sex of an exposed parent. Changes in the expression of membrane markers for the effector and regulatory T-lymphocytes depending on radiation type and dose load were determined. The growth factor level variations (TGF-beta1, EGF, HGF, FGF) in peripheral blood serum in persons exposed to radiation from gamma- or alpha-sources, allow us to consider them as biomarkers of radiation-induced disturbances in immune homeostasis. The concentration changes of TGF-beta1, apoptosis proteins (p53, TPA-cyk, sAPO-1/Fas), and the adhesion molecule sCD27 in the case of cardiovascular diseases in the serum of both irradiated and non-irradiated "Mayak" PA employers point to the information value of these immune response characteristics as specific biomarkers of cardiac disorders. It is proposed that the revealed changes in immune homeostasis and in the variability of somatic cell genome may provoke development of tumors and cardiovascular diseases in man in delayed periods after prolonged exposure to IR.     

Mitochondrial DNA Deletion in Offspring of Female Mice Exposed to X-Rays

Biophysics - Abstract—The effects of X-ray exposure on the mitochondrial genome were studied in the offspring of female mice exposed in the preconception period at doses of 0.5 and 2 Gy....     

AP-PCR assay of DNA alterations in the progeny of male mice exposed to low-level gamma-radiation

By comparative analysis of fingerprints of arbitrarily primed polymerase chain reaction (AP-PCR) products, DNA alterations in somatic cells of the progeny (F1 generation) of male mice chronically exposed to low-doses of gamma-radiation was investigated. Male BALB/c mice exposed to 10-50 cGy were mated with unirradiated females 15 days after irradiation. DNA was isolated from biopsies taken from tail tips of 2-month-old progeny. Preliminary AP-PCRs were carried out with 17 primers representing core sequences of micro- and/or minisatellites or their flanking oligonucleotides. Best quantitatively reproduced AP-PCR fingerprints of genomic DNA were obtained with one of these primers, a 20-mer oligonucleotide flanking the micro-satellite locus Atplb2 on mouse chromosome 11. Comparative analysis of individual fingerprints of AP-PCR products obtained on DNA templates from the progeny of irradiated and intact males revealed an increased variability of micro-satellite-associated sequences and an increased frequency of "non-parental bands" in DNA-fingerprints from the progeny of males chronically exposed to gamma-radiation 15 days before mating (at the postmeiotic stage of spermatogenesis). The results show that increased micro-satellite instability can be initiated by irradiation of the male parent to subsequently arise or be transmitted to the soma of the F1 generations.     

Increased genomic instability in somatic cells of the progeny of female mice exposed to acute X-radiation in the preconceptional period

The level of genome instability (GI) was studied in the progeny of female mice exposed in the preconceptional period to radiation doses of 0.5, 1, and 2 Gy in comparison to that in the progeny of the same parent pairs born before irradiation of the females. To assess the level of genome instability, we analyzed polymorphism of DNA fragments from postmitotic (blood and brain) and proliferating (spleen and tail tip) tissues amplified by AP-PCR (PCR amplification with an arbitrary primer). It was found that polymorphism of the spectrum of AP-PCR products, which is a multilocus genetic marker (MGM), in the genome of somatic cells in the progeny of female mice exposed to 2 Gy was higher than in the progeny of male mice exposed to the same doses. In the progenies of female mice born before and after irradiation, tissue-specific variations in the level of DNA polymorphism were detected. The maximum value of this polymorphism (with respect to the frequency of “nonparental bands”) was determined for peripheral blood DNA in comparison with the other tissues. Estimations of the MGM polymorphism with the AP-PCR method demonstrate an increased level of genome instability in somatic cells of offsprings from female mice exposed to a single acute dose of X-rays (0.5, 1, and 2 Gy) in the preconceptional period. Radiation-induced transgenerational genome instability with an increase in the dose of preconceptional irradiation of female mice was more pronounced in DNA of the postmitotic tissues (blood and brain DNA) than in DNA of the proliferating tissues (spleen and tail tip epithelium).     

Study of genome instability using DNA fingerprinting of the offspring of male mice subjected to chronic low dose gamma irradiation

By a polymerase chain reaction with an arbitrary primer (AP-PCR), the possibility of transmission of genome instability to somatic cells of the offspring (F1 generation) from male parents of mice exposed to chronic low-level gamma-radiation was studied. Male BALB/c mice 15 days after exposure to 10-50 cGy were mated with unirradiated females. Biopsies were taken from tale tips of two month-old offspring mice and DNA was isolated. The primer in the AP-PCR was a 20-mer oligonucleotide flanking the microsatellite locus Atp1b2 on chromosome 11 of the mouse. A comparative analysis of individual fingerprints of AP-PCR products on DNA-templates from the offspring of irradiated and unirradiated male mice revealed an increased variability of microsatellite-associated sequences in the genome of the offspring of the males exposed to 25 and 50 cGy. The DNA-fingerprints of the offspring of male mice exposed to chronic irradiation with the doses 10 and 25 cGy 15 days before fertilization (at the post-meiotic stage of spermatogenesis) showed an increased frequency of "non-parent bands". The results of the study point to the possibility of transmission to the offspring somatic cells of changes increasing genome instability from male parents exposed to chronic low-level radiation prior to fertilization.     

Specialized software product for comparative analysis of multicomponent DNA fingerprints

GelAnalyzer software, which is used to identify and correctly compare DNA fingerprints consisting of a large number of discrete bands, has been developed by the project to study the quantitative changes in DNA polymorphism patterns in animals and humans exposed to gamma radiation. The actual capabilities of this program are much broader and include the possibility to analyze the images of any multicomponent gels containing fragments of DNA, RNA, and proteins. This software product runs on Windows. GelAnalyzer allows one to analyze gel images obtained by a scanner, camera, or digital camera and ensures the visual control of the identification and comparative analysis of bands; it also makes it possible to take into account the bands that are poorly identified automatically and exclude the artifacts (incidental marks) on images. The operation of GelAnalyzer software is based on the determination of the values of normalized coordinates of bands with allowance for the relative electrophoretic mobility (Rf) of PCR products and comparison of their spectra (set of bands in gel lanes) to reveal the similarities or differences in their components with subsequent statistical data processing and display the results of the analysis.