[The study of aberrant methylation in blood leukocytes of liquidators of the Chernobyl accident].

The study of aberrant methylation of CpG islands in the promoter regions of genes (P16/CDKN2A, P14/ARF, RASSF1A, GSTP1) in blood leukocytes of liquidators of the Chernobyl accident (n = 83, 38-76 years of age) and control subjects of two groups (n = 48, age ≤ 35 and n = 65, age > 35) was carried out using methylation-sensitive restriction endonuclease analysis followed by PCR. The total number of AciI sites in the analyzed fragments ranged from 2 to 7 for different genes. Only 1 subject (2.1%) from the control group (healthy young individuals, age ≤ 35) has methylation of the studied CpG--dinucleotides of RASSF1A gene. Promoter methylation of at least one of the genes analyzed was observed in 28.92% liquidators and significantly exceeded (p = 0.016) such rate in a one-age (> 35 years of age) control group (12.31%). A significantly elevated frequency (p = 0.023) of individuals with abnormal methylation of GSTP1 gene in the group of liquidators as compared to the control group was revealed. The occurrence of promoter methylation of RASSF1A gene significantly correlated with aging both in the control group (r = 0.214; p = 0.023) and in the liquidators of the Chernobyl accident (r = 0.230; p = 0.036). No similar trend was found for other genes. Multiple regression analysis showed that the growth in the number of methylated loci of a set of genes p16, p14 and GSTP1 is exclusively due to the fact of exposure (OR = 7.32, 95% CI = 2.49-25.83, p-value = 2.7 x 10(-5)). The results obtained demonstrate for the first time the reality of the radiation-induced aberrant methylation of CpG islands in promoters of genes involved in the basic protective, functions of cells in the human body in remote periods after radiation exposure.

[The alterations in profiles of RAPD- and of ISSR-markers in progeny of the single gamma-irradiated mice of BALB/c strain].

Molecular-genetic effects in the offspring of BALB/c male mice exposed to single radiation doses of 1, 2 and 3 Gy were studied. Induced genetic variability was studied using such methods as assessment of variation RAPD- and ISSR-profiles. Comparative analysis of genetic radiosensitivity of stem spermatogonia and of spermatids is presented in the work. The frequency of changes in the patterns of the offsprings of irradiated mice was significantly different from the analogous parameters in the offsprings of the control group already at a dose of 1 Gy. Comparative analysis of genetic radiosensitivity at different stages of spermatogenesis revealed the similar sensitivity of spermatogonia and of spermatids at 1 and 3 Gy and a higer sensitivity of spematogonia at 2 Gy.

[The analysis of mutations at mini- and microsatellite DNA loci in the family members of a group of occupationally exposed to tritium and tritium oxide plant workers].

Mini/microsatellite (MNS/MCS) loci are efficient tools in solving basic and applied problems in different spheres of biology and medicine due to their unique characteristics - a high frequency of tandem repeats in combination with their wide variability. Specifically, they have been found use as potential markers of genetic effects of ionizing radiation on animals and on human. However there is no general agreement as to the influence of irradiation on the frequency of mutations in hypervariable repetitive DNA sequences up to now. The present work is the study of the mutation frequency at MCS/MNS loci in 19 families of workers occupationally exposed to chronic beta-radiation from tritium and tritium oxide (examined group), and in the control group included 23 families. The results have indicated that the average frequency of microsatellite mutations in the examined group made up 4.7% and exceeds about 7-fold the same parameter of the control group (0.7%). This differences is statistically significant (p = 0.004). The average frequency of minisatellite mutation in the examined group made up 3% while in the control group it was 2 time lower (1.5%), but this difference is not statistically significant. Mutations for 4 MCS and 2 MNS loci were revealed in two children from one family (the total reconstructed dose in their father was about 1000 mSv). If we exclude this family from statistical analysis the frequencies of MCS and MNS mutations in the children of nuclear workers do not statistically differ from the control values.

[Development of amplification technology-based methods for tick-borne encephalitis virus detection and genotyping].

The development of methods for tick-borne encephalitis (TBE) virus genotyping, such as determination of nucleotide sequences, analysis of restriction fragment length polymorphism, polymerase chain reaction using genotype-specific probes with hybridization-fluorescence detection, is under consideration. The capacities and application area of varions typing techniques, such as TBE detection and genotyping in the field, autoptic, and cultural materials; study of circulating genotypes in different natural foci and in various viral vectors and carriers, are described. Each technique was tested using a large number of strains isolated from ill humans and ticks in different regions of the country and employed to investigate abundant materials.

[The influence of chronic and of acute irradiation on the polymorphism of RAPD-markers in offspring for irradiated mice].

On mice lines BALB/c and CBA/lac was performed the study of molecular-genetics effects in mice progeny after the chronic (dose rate -0.0017 Gy/day, total dose -0.36 Gy) and acute (dose range 1-3 Gy) exposure of y-radiation on the parents. For variability analysis was used technique of amplification DNA with series of random primers (RAPD-assay). Random primers were used as single primer and in mixture of ones. In this work were held the comparative analysis of the genetic radiosensitivity for stem spermatogonia and spermatides. After the acute exposure the dose dependence for levels of polymorphism of RAPD-markers were obtained. After the chronic irradiation, significant differences from control group were obtained only by use primers mixture M1. Comparative analysis of the genetic radiosensitivity of different stages of mice spermatogenesis are display is similar sensitivity of stem spermatogonia and spermatides after doses of irradiation 1 Gy and 3 Gy. Indicated that after irradiation by dose 2 Gy, spermatogonia are more sensitivity than spermatides.

[Estimation of genetic effects of chronic exposure to low-dose rate gamma-radiation by cytogenetic methods and DNA-comet assay]. / Otsenka geneticheskikh éffektov khronicheskogo vozdeistviia nizkointensivnogo gamma-izlucheniia tsitogeneticheskimi metodami i metodom DNK-komet.

The study of genetic effects in CBA/lac mice exposed for 1 year to constant low dose-rate gamma-radiation at a dose-rate 63 cGy/year has been carried out. We have shown the significant increase in the DNA breaks' level in spleen lymphocytes by comet-assay beginning from the total absorbed dose of 20 cGy. It is possible that the DNA breaks' level increase resulted from the structural rearrangement of chromatin or induction of lymphocyte proliferation. The results obtained by micronucleus test have proved that the mutagenic effect of chronic low dose-rate gamma-radiation depends on cell type and respectively on cell proliferation rate, cell differentiation, etc. So, by the end of experiment the significant increase in the frequency of PCE with micronuclei (MN) was observed. However, in contrast, the frequency of NCE with MN was not increased. No significant increase in the percent of lung cells with MN was registered.

[Assessment of single-stranded DNA breaks in spleen lymphocytes using the DNA comet assay in the long term after acute irradiation of mice at sublethal and moderately-lethal doses]. / Otsenka metodom DNK-komet odnonitevykh razryvov DNK limfotsitov selezenki v otdalennye sroki posle ostrogo oblucheniia myshei v subletal'nykh i sredneletal'nykh dozakh.

The single-strand DNA level in spleen lymphocytes of BALB/c male-mice after 11 month acute exposure to gamma-radiation at doses 1, 3 and 6 Gy has been investigated by comet assay. The results of our study showed that at 11 month after irradiation at different doses a significant increase in the level of DNA breaks in spleen lymphocytes and decrease in the total number of these cells in mice was registered. It is possible that the increase in the DNA breaks is due to the effort of the compensatory proliferation process in lymphoid system that can give the increase in the number of different genetic disturbances in lymphocytes.