[Molecular-biological properties of blood lymphocytes of Hodgkin's lymphoma patients. Plausible possibility of treatment effect prognosis].

Hodgkin's lymphoma (HL) patients were investigated before and during chemical and radiation therapy. The properties of peripheral blood lymphocytes of the HL patients before treatment have been compared with healthy donors and the patients during the treatment. The genetic damage--frequency of cells with micronuclei (MN), the level of DNA single- and double-strand breaks (SSB and DSB), DNA-protein cross-links (DPC) have been studied. Biochemical and physiological parameters have been compared as well: the concentration of reactive oxygen species (ROS), the ability to the adaptive response induction. The radiosensitivity of lymphocytes in vitro exposed to the 1 Gy irradiation has also been determined (by MN test). It was shown that in Hodgkin's lymphoma patients' lymphocytes (in comparison with healthy donors) the frequency of cells with MN does not change, the level of SSBs and DSBs increases, the amount of DPC does not change, and ROS concentration (on average) significantly increases because of the part of the population that have high ROS content. The ROS concentration decreases to control level, the frequency of cells with MN increases, the level of DSBs does not change but the level of DPCs (which prevents the determination of DSB) increases in the patients during treatment. It was also discovered that lymphocyte radiosensitivity correlates with the MN cells frequency before treatment and the ROS concentration. These results make it possible to suppose that the high MN frequency and high ROS concentration in Hodgkin's lymphoma patient lymphocytes (before treatment) can serve as prognostic factors for the effectiveness of radio and chemical therapy.

[Genome damage and reactive oxygen species production in the progenies of irradiated CHO-K1 cells].

A series of experiments to study the delayed effects of gamma-radiation exposure in different generations of the progenies of the Chinese hamster ovary CHO-K1 irradiated cells has been conducted. It has been shown that in the progenies of the cells irradiated with a dose of 1 Gy, the following effects are observed: in the 9-27 cell generations - increase in the genome damage, intracellular production of reactive oxygen species, apoptotic cells percentage and cell sensitivity to additional exposure (irradiation at a dose of 10 Gy); in the 30-42 cell generations - decrease of the studied parameters up to control values and increased resistance to additional exposure. It is assumed that the decrease of the studied parameters up to the control values in the 30-42 postirradiation generations of cells is caused by elimination of damaged cells or transition of genomic instability into a hidden (latent) condition.

[Molecular and cellular consequences of the Chernobyl accident].

In this paper the results of the Chernobyl accident investigation 5-10 and 24 years after are summarized. The genomic instability, adaptive response formation, genome damage and oxidative status have been investigated. The studies were performed on cells in culture, mice, children and adults living in contaminated areas and liquidators. On cells in culture after exposition in the accident zone and culturing thereafter in laboratory conditions the cell proliferative activity decrease; the late cell death, the frequency of cells with micronuclei and giant cells increasing have been observed. In the progeny of exposed cells the enhancement of radiosensitivity has been noticed. So we can suppose that in cultured cells exposition in the zone of the accident the genomic instability is induced which results in many disturbances. At the organism level in mice exposed in the Chernobyl zone the radiosensitivity increase and the decrease of endotheliocytes density in brain tissue has been observed. On the stimulated by PHA blood lymphocytes of children the increase of the frequency of cells with micronuclei more than 2 time have been noticed. In all groups investigated, the decrease of individuals with significant adaptive response was observed. In children and adults inhabitants the increase of radiosensitivity after low dose of irradiation has been noticed. 24-year after the accident it was discovered that in liquidators lymphocytes the frequency of cells with micronuclei, with chromosome type aberrations, with DNA double strand breaks have been increased; the reactive oxygen species (ROS) were decreased in comparison with the control population. We can suppose that genomic instability induced in residents of contaminated regions and liquidators long after the accident results in the genetic apparatus damage, radiosensitivity enhancement, hypoxia that represent risk factors and increase the probability of tumour and non-tumour diseases. The development of these pathological processes may happen in much more remote periods.

[The connection between molecular-cellular parameters and immune status of liquidators after Chernobyl accident].

The genome damage (the frequencies of cells with micronuclei (MN), chromosome aberrations, the level of DNA double-strand breaks (DSB DNA), the concentration of reactive oxygen species (ROS) and 28 immunological parameters have been studied on the blood lymphocytes of Chernobyl accident liquidators. The purpose of this article was the investigation of cytogenetic, molecular changes of blood lymphocytes of irradiated individuals 24 years after accident, examination it there are correlation between genome damage and immunological parameters. It was shown that in lymphocytes of liquidators the frequencies of cells with MN and with all type of chromosome aberrations didn't differ from the lymphocytes of nonirradiated individuals, but the frequency of chromosome aberration type was increased, the level of DSB DNA was increased too. The concentration of ROS is decreased. The percent of cytotoxic CD8(+)-T-lymphocytes, natural killer cells (CD16(+)-lymphocytes), CD3+ CD16+ CD56+ (NK-T-cells), that posses antivirus and antitumor activity--HLA-DR+, regulatory T-lymphocytes (CD4+ CD25+high) in liquidators significantly increases. The level of serum immunoglobulin (Ig A) significantly increases too. The index of immune regulation, meaning of phagocyte neutrophil (FAN) and macrophage activity decreases. In liquidators there are significant correlation between the frequencies of cells with MN and the content of regulatory T-lymphocytes (p < 0.05), between the concentrations of ROS and activated T-lymphocytes. More connection is on the tendency level (p < 0.10): the frequency of chromosome aberrations, the DSB DNA level with natural killer cells and regulatory T-lymphocytes; the frequency of cells with MN and DSB DNA and FAM. We can suppose that genomic instability induced by the liquidators of Chernobyl accident consequences 24 years ago manifests now as increased genome damage and oxidative status decrease that can result in imbalance of cells and humoral immune status, disturbancies of health.

[Molecular-cellular characteristic of blood lymphocytes in Hodgkin lymphoma].

The molecular-cellular parameters complex has been studied on the blood lymphocytes of malignant Hodgkin's lymphoma (HL) patients: the frequency of cells with micronuclei (MN) and chromosome aberrations; the level of DNA single and double strand breaks - OR and DR DNA (DNA comet assay), oxidative status--the content of reactive oxygen species (ROS) by using nonfluorescent dye that is oxygenated in the cells to fluorescent reagent and detection of fluorescence intensity after there. It was shown that the patients with LH had the increased level of DR and OR DNA, the increased frequency of cells with chromosome aberrations and the number of aberrations per cell was increased too. The concentration of ROS is increased too for the most individuals with intoxication. In the process of the chemical and radiation therapy the increase of OR DNA level, the frequency of the cell with MN has been registered. The ROS concentration correlates with the level of DNA-strand breaks. So the blood lymphocytes of HL patients before treatment differ from the lymphocytes of healthy donors. The damage of genome and the change of oxidative status have been observed that can be additive markers for the HL diagnosis, their sensitivity to the treatment and the characteristic of lymphocytes changes by this disease.

[Genomic damage, adaptive response induction in blood lymphocytes at the prostate cancer patients. Connection with the tumour radiotherapy efficiency].

On the blood lymphocytes of prostate cancer (PCa) patients before and during radiotherapy: DNA damage by DNA commet assay (DNA double strand breaks - DSB); the frequency of cells with micronuclei (MN) with cytokinetic cytochalasin block; the adaptive response induction by the additional irradiation of PHA stimulated lymphocytes in the doses of 0.05 and 1.0 Gy 24 h and 48 h after stimulation were studied. Changes of these parameters with the decreasing of prostate specific antigen (PSA) have been compared. PSA decreasing is an adequate of the radiotherapy efficiency. It was shown that in oncological patients the DSB level and the frequency of cells with MN have been increased. During radiotherapy (in 3 months) the DNA DSB level and the frequency of cells with MN is enhancing. The degree and direction change of these parameters coincide. It was discovered the significant correlations between the enhancing of DNA DSB level and the cell frequency with MN during therapy and degree of the PSA level decreasing. Then it was shown that when the cell frequency with MN before treatment is higher the radiotherapy efficiency is worse. These results can have great significance for the evaluation of the prognosis of the treatment efficiency. The investigation of lymphocytes for the adaptive response ability has shown that in the patients with the pronounced adaptive response before radiotherapy the decrease of PSA level during treatment was not significant (in mean 3.5-3.6 ng/ml); when the adaptive response is absent or the phenomenon of enhanced radiosensitivity was observed the PSA level (in the most cases) was decreased very essential (in mean 0.07 ng/ml). We can suppose that prognosis of the treatment efficiency of the prostate cancer patients with the pronounced adaptive response in blood lymphocytes will be worse.

[Double-strand DNA breaks induction and repair in human blood lymphocytes irradiated with adapting dose].

Using a DNA-comet assay was shown that irradiation of human blood lymphocytes at G1 cell cycle with a low conditioning dose (5 cGy) induces an adaptive response (AR) manifested in reduction of the double-strand DNA (DSB) amount induced by challenging dose at 10 Gy. 24 h after conditioning irradiation (48 h after PHA addition) in cells irradiated at both conditioning and challenging doses a relative DBS amount was approximately 24% less in comparison to versus a control irradiated at challenging dose only. 48 h after adapting irradiation this index increased to approximately 35%, while 72 h after was decreased to approximately 29%. AR observed by us during 72 h after its induction did not accompanied by statistically significant changes in DBS repair enhancing. It is possible to assume that basic role in AR forming in lymphocytes under experimental conditions used by us playing the processes preventing radiation-induced DBS formation (antioxidant defense system activation, chromatin conformation changes ets).

[Adaptive response in the different mitotic cycles after irradiation].

The frequency of cells with chromosome aberrations and the number of aberrations per cell by metaphase analysis have been studied in the nonirradiated progeny of irradiated human blood lymphocytes. The DNA fragmentation (DNA double strand breaks) have simultaneously been investigated by the DNA comet assay. PHA stimulated lymphocytes have been irradiated in the adaptive dose 0.05 Gy 24 h and in the challenge dose 1 Gy 48 h after stimulation to study the adaptive response (AR). 5-bromodeoxyuridine have been added for the identification the first--the fourth mitoses. It has been discovered that the frequency of chromosome aberrations is increased is all mitotic cycles after challenge irradiation, the level of double strand breaks is increased too. The adaptive response in induced by the adaptive and challenge irradiation in the first and the second mitotic cycles (fixation 48 and 72 h after stimulation) for the most parts of individuals, but it is absent in the third and the fourth mitosis. Only chromatid aberrations are observed in the first mitosis, but chromosome aberrations--in the following mitosis. Investigation by the DNA comet assay have showed the adaptive response is noticed 48-72 h after stimulation but it is insignificant 96 h. The conclusion is that the genomic instability is observed in nonirradiated progeny irradiated lymphocytes; the adaptive response is manifested up to third mitosis and is explained by the decreasing of the number of the chromatid and chromosome aberrations and DNA fragmentation. We can suppose that double strand DNA breaks can be signaling damage for the adaptive response induction.

[Influence of chronic exposure to low doses of gamma-radiation and 90Sr on the level of DNA breaks and cell sensitivity to hydrogen peroxide in the mouse spleen].

Using comet assay, a statistically significant increase (p < 0.05) in the level of DNA breaks in spleen cells was revealed in male CBA/lac mice exposed to gamma-radiation (1.7 cGy/day) or 90Sr (150-250 Bq/day) for 210 days. The level of DNA breaks also increased under combined exposure to both gamma-radiation and 90Sr (p < 0.05), but to a lesser degree than under exposure to each of these factors alone. Upon additional in vitro treatment of spleen cells with hydrogen peroxide, the relative increase in the level of DNA breaks was smaller in cells of irradiated mice than in the control. The ratio of the level of DNA breaks after hydrogen peroxide treatment to that before this treatment in control mice was 4.2 +/- 0.9, compared to 1.4 +/- 0.6 in gamma-irradiated mice, 1.9 +/- 0.8 in 90Sr-irradiated mice, and 2.3 +/- 0.8 in mice exposed to both gamma- and 90Sr-irradiation.