[Distribution of individuals by spontaneous frequencies of lymphocytes with micronuclei. Particularity and consequences].

By micronucleus (MN) assay with cytokinetic cytochalasin B block, the mean frequency of blood lymphocytes with MN has been determined in 76 Moscow inhabitants, 35 people from Obninsk and 122 from Chelyabinsk region. In contrast to the distribution of individuals on spontaneous frequency of cells with aberrations, which was shown to be binomial (Kusnetzov et al., 1980), the distribution of individuals on the spontaneous frequency of cells with MN in all three massif can be acknowledged as log-normal (chi2 test). Distribution of individuals in the joined massifs (Moscow and Obninsk inhabitants) and in the unique massif of all inspected with great reliability must be acknowledged as log-normal (0.70 and 0.86 correspondingly), but it cannot be regarded as Poisson, binomial or normal. Taking into account that log-normal distribution of children by spontaneous frequency of lymphocytes with MN has been observed by the inspection of 473 children from different kindergartens in Moscow we can make the conclusion that log-normal is regularity inherent in this type of damage of lymphocytes genome. On the contrary the distribution of individuals on induced by irradiation in vitro lymphocytes with MN frequency in most cases must be acknowledged as normal. This distribution character points out that damage appearance in the individual (genomic instability) in a single lymphocytes increases the probability of the damage appearance in another lymphocytes. We can propose that damaged stem cells lymphocyte progenitor's exchange by information with undamaged cells--the type of the bystander effect process. It can also be supposed that transmission of damage to daughter cells occurs in the time of stem cells division.

[Adaptive response of blood lymphocytes as a marker of hemopoiesis status in exposed persons].

An obvious correlation between the type of reaction manifested by peripheral blood lymphocytes to low dose irradiation in vitro (adaptive potential), the RBM cell composition (during the period of the major exposure), and the peripheral blood cell composition (at a late time period coincident with the studies of induced radioresistance) has been found in the Techa riverside residents in the later periods after the onset of a long-term low-dose rate radiation exposure (55-60 years later) within a range of individual red bone marrow doses from 0.01 to 1.79 Gy. The nature of these dependences observed in chronically exposed individuals differs from that revealed in the controls. It can be suggested based on the results of the study that the capacity for the adaptive response shown by peripheral blood lymphocytes donated by exposed persons in the remote period after exposure can be regarded as a biological marker of the functional state of the hemopoietic stem cell pool.

[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.

[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.

[The radiosensitivity change of human blood lymphocytes in different mitotic cycles after a low dose irradiation].

The adaptive response (AR) in stimulated blood lymphocytes (8 donors) have been studied by two methods: analyses of unstable chromosome aberrations in metaphases and micronuclei assay (MN) with cytochalasine cytokinetic block. The adaptive irradiation in the dose of 5 cGy have been conducted 24 h and challenge irradiation after 48 h after stimulation. For the metaphase analysis of the first and subsequent mitosis cells were incubated with BrdU and were fixed 50, 72 and 96 h after the stimulation. In the MN test cells were fixed 72 or 96 h after the stimulation and cytochalasin B was added in the cell culture 24 h before fixation. Was shown that in the cells of first mitoses fixed at 50 h after stimulation only chromatid aberrations are presented in the lymphocytes of all donors and AR in all donors was noticed; but when fixation was conducted 72 h after stimulation the chromosome type aberrations are prevailing and AR is absent, 96 h after--one donor has AR. Was discovered that in the cells of the second mitosis fixed at 72 h after stimulation the only chromosome type aberrations observed, their frequency is higher that in the lymphocytes of 1 and 3 mitosis in the same fixation time, in 7 of 8 donors there is AR registered. In the cells of 3th mitosis 72 h fixation time only 1 donor has AR, in cells of the 4th mitosis nobody have AR. By the MN assay AR in two donors is observed, in the first--the increase in radiosensitivity after adapting irradiation is noticed in the rest of the radiosensitivity is not changed. 78 percent of results coincidence by MN assay with time fixation 72 and 96 h with the results of metaphase analysis in the cells of first mitosis in the same time fixation was observed. The proposition of these data explanation is that the decrease of chromatid and chromosome aberration frequency is the one is result of different enzyme systems function and the ability to the these system induction in different cells is different. If the one is decreasing the chromatid aberrations frequency is induced in lymphocytes of most donors, the second is induced rarely only in the part of cells.

[The adaptive response of first generation offsprings irradiated parents].

In offspring's of first generation irradiated inhabitants of Techa river (fathers, mothers and both parents) the spontaneous level of damaged blood lymphocytes, sensitivity of lymphocytes to the additional acute irradiation in dose 1.0 Gy and radioinduced adaptive response after adaptive (5 cGy) and challenge (1.0 Gy) irradiation 5 h after was studied. The micronuclei test with cytochalasin B as a criteria of the effect have been used. It was shown, that descendents of irradiated parents differ from the control group. The main difference is the significant decrease of the adaptive response frequency in the progeny. In the offspring's of the irradiated fathers and mothers there is no one individuals with the adaptive response; in the offspring's of both irradiated parents the frequency of individuals with adaptive response decreases in control from 19.5% to 6.8%. The distribution of descendents according to response on adaptive irradiation differ significantly from the control distribution and from the each other. And the tendency to the radiosensitivity increase after adaptive irradiation was observed. In the whole joint group of progeny the mean spontaneous cell frequency with micronuclei decreased, but the sensitivity of lymphocytes to the additional acute irradiation doesn't differ from the control. The results of the paper permit to suppose that transgenerational genome instability in human can be determined. Earlier discovered decrease of the adaptive response frequency in the Techa river livings is observed in the offspring's of irradiated fathers, mothers and both parents.

[Determination of catechelamines, serotonin, and 5-hydroxyindoleacetic acid in a blood].

The specific features of chromatographic electrochemical detector determination of plasma catecholamines, serotonin, and 5-hydroxyindoleacetic acid (5-HIAA) are considered. Platelet-poor plasma catecholamines were concentrated on alumina. The concentrate was injected into a chromatograph. Serotonin and 5-HIAA were measured in platelet-poor plasma and thrombocytic clot. The data available in the literature and their authors' own findings of the sample preparation parameters that could minimize the artificially increased platelet-poor plasma concentrations of serotonin via thrombolysis, as well its loss on protein sedimentation.

[On the response of a cell population to low-dose irradiation].

The cell composition of a population of human blood lymphocytes was studied after irradiation at doses of 5 cGy, 1.0 Gy and 5 cGy + 1.0 Gy and the use of a cytokinesis block. The frequencies of uni-, bi- and multinucleate lymphocytes with and without micronuclei (MN) were taken into account. By the standard criterion the frequency of binucleate lymphocytes with MN among binucleate lymphocytes--the donors were characterized as follows: in with reduction of radiosensitivity after irradiation with 5 cGy + 1.0 Gy as compared to the values of radiosensitivity after irradiation with 1.0 Gy only (an adaptive response, AR); in with no change of radiosensitivity after exposure to these doses (no AR); and with an increased ofradiosensitivity after exposure to these doses (syndrome of increased radiosensitivity, IRS). It was found that upon exposure to 1.0 Gy and 5 cGy + 1.0 Gy in some donors with AR, without AR and with IRS the total numbers of damaged cells in the population and the number of binucleate cells with MN were equal. This result calls in question the involvement of the repair mechanism in the alteration of radiosensitivity of lymphocytes in these donors. It was also observed that in the same donors a simultaneous increase (or a decrease in the case of IRS) of the portion of undamaged binucleate cells in the population took place. Our results demonstrate the existence of a new, populational, mechanism involved in the alteration of radiosensitivity after exposure to the adaptive and challenge doses.

[An individual variability of the adaptive response to irradiation in human cells. Approach to its determination].

On human blood lymphoxytes with micronuclei (MN) assay and cytokinetic cytochalasin block and analysis of chromosome aberrations the change of cell population composition, adaptive response (AR) and phenomenon of enhanced radiosensitivity after low dose (5 cGy) and challenge doses (1.0 Gy) have been studied. Irradiation have been carried out in G1 and G2 phases of cell cycle (24 h and 48 h after PHA stimulation). Fixation of cells have been conducted after 50 h (2 h after demecolcin adding) and 72 h (24 h after cytochalasin adding) chromosome and MN assay. Evaluation criteria were the frequency of binucleated cells with MN on 1000 binucleated cells and the frequency of cells with chromatid aberration on 100 metaphases. It was shown that cell population constitution change, AR occurring depended on the individual peculiarity. The evaluation of AR presence by the indexes of bimucleated cells with MN frequency and cells with chromatid aberrations don't coincide (coincidence is observed in 3 cases from 15). It is supposed that in G2 phase after irradiation in challenge dose the MN assay and metaphase analysis can register different cells (24 h and 2 h after mitotic block). The cell population constitution change can probably influence on the AR evaluation but in isn't the AR mechanism. The main mechanism of AR forming * the protection from the damages by different ways. AR depends on many factors, individual peculiarities observes by the use of definite evaluation criteria, in individuals with definite genetic constitution. Perhaps these considerations permit to discuss the problem of AR universality.

[Effect of low dose irradiation on the reaction of blood lymphocytes of individuals with the somatic diseases].

The reaction of blood lymphocytes on adaptive irradiation in vitro (0.05 Gy) and challenge irradiation (1.0 Gy) 5 h after has been studied among 320 children 3-16 years old by micronuclei test with cytochalasin B cytokinetic block. The adaptive response or the hypersensitivity phenomenon (HS) has been determined by the calculation of the number of binucleated cells with micronuclei (MN) in 1000-2000 binucleated cells. For each individual by the chi2 criteria significant differences between the frequencies of damaged cells by irradiation in challenge dose and by combined action of adaptive and challenge doses have been detected. On the base of the results obtained the connection between AR or HS and the somatic diseases of different type and the disturbancies in systems of humoral and cell immunity has been studied. It was shown that in the row of allergic diseases and tuberculous infection among the significant part of the children (approximately 50%) the HS phenomenon was registered. The connection between the HS phenomenon and inflammatory processes (without allergic components), the logopedic disturbancies wasn't observed. It was shown in the group of HS children the deviation in the cell immunity system is registered. Significant deviations in HS group in the system of the humoral immunity is observed. So by the HS phenomenon significant number of the children have the somatic diseases and the deviation in the systems of cell and humoral immunity.

[Adaptive response of blood lymphocytes of the inhabitants of the South Ural chronically exposed to radiation]. / Adaptivnye sposobnosti limfotsitov krovi u zhitelei iuzhnogo urala, podvergshikhsia khronicheskomu oblucheniiu.

The inhabitants of Techa river villages exposed to injured radiation action and inhabitants of uncontaminated regions were examined by micronuclei (MN) assay. The initial damage of blood lymphocytes, the role of radiation in the induction of sensitivity to the acute irradiation and the ability to form the adaptive response were evaluated. It was shown that the initial level of damaged lymphocytes in the inhabitants of the contaminated area did not differ significantly from the spontaneous level. But in these people the sensitivity to acute irradiation was decreased. The ability to develop the adaptive response was decreased too. It was suggested that the radiation plays the main role in the development of radiosensitivity and adaptive response in chronically irradiated people.

[The cytogenetic damage, radiosensitivity and adaptive response in children living in the different districts of Moscow]. / Tsitogeneticheskie povrezhdeniia, radiochuvstvitel'nost' i adaptivnyi otvet u detei, prozhivaiushchikh v raznykh raionakh Moskvy.

The spontaneous level of blood lymphocytes with micronuclei (MN), the sensitivity to 1.0 Gy irradiation and adaptive response (AR) after adaptive irradiation with a dose of 0.05 Gy 5 hr later have been studied in children population living in different districts of Moscow. It was shown that spontaneous frequency of cells with MN, the sensitivity to 1.0 Gy acute irradiation and the AR manifestation have significant differences in samples taken from children living in different districts. The individual variability is significant also. In each group of children the individuals with the enhanced radiosensitivity after adaptive irradiation have been observed. In conformance with the data of radioecological inspection the radiation situation in different Moscow districts is quite safe on overage but in some districts the spontaneous level of lymphocytes with MN, and radiosensitivity after 0.05 Gy irradiation were enhanced, the AR was not found.

[On the new mechanism of adaptive response]. / O novom mekhanizme formirovaniia adaptivnogo otveta.

The role of changes in cell composition of population of human blood lymphocytes in the forming of an adaptive response (AR) has been studied. By micronuclei assay and cytokinetic block with cytohalasin B the frequency of mono-, bi- and multinuclear cells with micronuclei (MN) and without MN were determined in the initial population. The same parameters have been studied after exposure of the population to the adaptive (0.05 Gy), challenge (1.0 Gy) doses and to doses 0.05 + 1.0 Gy 5 hours after. 13 from 23 investigated individuals manifested the AR: the decreasing of the ratio of damaged binuclear cells to the all binuclear cells after the adaptive and challenge exposure. It was shown that the ways of an AR forming are different: in 7 of 13 individuals with AR the number of binuclear cells with MN did not decrease but the amount of binuclear undamaged cells increased. The ratio of these parameters enhances but not for the account of cells with MN decreasing. There is the linear correlation between the frequency of cells with MN and the frequency of binuclear cells in population (spontaneous, after irradiation with doses of 0.05, 1.0 and 0.05 + 1.0 Gy) with the coefficient of correlation about -1. These results show the presence of new mechanism of AR forming, which is not connect with the induction of damage repair and rather with the stimulation of cell division. In the another group of individuals the decrease in damaged cells number after irradiation with doses of 0.05 + 1.0 Gy have been observed. Probably the stimulation of repair system occurred to the moment of 1.0 Gy irradiation. Thus, the mechanism of an AR forming depends on the individual properties of organism. The work was suppoted by RFBR grant 03-04-48325a.

[The reaction of cell population to low level of irradiation]. / Reaktsiia populiatsii kletok na obluchenie v malykh dozakh.

The results of long-term investigations of the effectivity of low level irradiation (below 0.5 Gy) carried out on the cells in culture and blood human lymphocytes (adults and children) have been brought. In the experiments conducted in the laboratory conditions and in the contaminated with radionuclides regions (after Chernobyl accident) the genomic instability have been discovered. The cell manifestations of the genomic instability have been registered in the progeny of irradiated cells as the decreasing of proliferative activity, the increasing of the frequency of cells with micronuclei, the increasing of cells with sister chromatid exchanges, the late cell death, the absence of the adaptive response ability, the enhancement of the radiosensitivity. The results of the investigations of the adaptive response of blood lymphocytes have been presented. It was shown that in all populations investigated there are individuals without the adaptive response and the individuals with the enhancement of radiosensitivity after adaptive irradiation (0.05 Gy). On the basis of own results and the data of literature the possible mechanisms of low level irradiation effects are discussed. The conclusion is that: a. The population with new properties can be formed after low level irradiation; b. The effects and mechanisms of this effect realization can be different from that after irradiation with high doses.

[The cell composition in the lymphocyte population and the radiation adaptive response]. / Kletochnyi sostav populiatsii limfotsitov i radiatsionnyi adaptivnyi otvet.

Using lymphocytes of 5 healthy individuals the ability to adaptive response (AR), cell composition of population after PHA stimulation, changes in cell composition population after irradiation in the dose of 1.0 Gy and after irradiation in adaptive (0.05 Gy) and challenge (1.0 Gy) doses have been studied. AR observed in 2 of the 5 individuals only. After PHA stimulation the persons with AR have the total amount of cells after mitosis or during mitosis (the number of binucleated cells + the number of multinucleated cells + the whole cells with micronuclei + the number of mitotic cells) on average is higher than in persons without AR. In individuals with AR the linear correlation between the number of binucleated cells with micronuclei (on the 1000 scored binucleated cells) and the part of binucleated cells in the population is observed with coefficients of correlation -0.89 and -0.91. In the humans without AR this correlation is absent. The correlation observed permits to suppose that AR may occur at the expense of not only the decrease in number of damaged lymphocytes, but also the increase in the share of not damaged binucleated cell with the stable number of damaged cells.

[Cytogenetic results of irradiation of Southern Ural residents]. / Tsitogeneticheskie posledstviia oblucheniia zhitelei iuzhnogo Urala.

The paper summarizes the results of studies of 85 individuals exposed in the Southern Ural region. The spontaneous frequency of the cells with micronuclei (MN) in a population of human blood lymphocytes after PHA stimulation and cytokinetic block with cytochalasin B has been determined. The sensitivity of lymphocytes to the irradiation at the dose of 1.0 Gy and the adaptive response (AR) after the irradiation at the low adaptive dose of 0.05 Gy, and the challenge dose of 1.0 Gy 5 h later have been studied too. It was shown that the peculiarity of the Urals population consists in a higher individual variability of the frequency of cells with MN in all groups have been investigated (spontaneous, after acute irradiation in the dose 1.0 Gy) in comparison with Moscow people. The proportion of persons with a significant AR in the Urals groups was considerably lower than that identified among Moscow residents, and the number of persons with enhanced radiosensitivity increased following low-dose irradiation. We can suppose that prolonged action of low level radiation with another ecological factors, living in the contaminated regions result in the enhancement of the sensitivity to the genotoxic agents in the separate individuals.

[Level of spontaneous and radiation-induced cytogenetic damage in blood lymphocytes of children depending on age and life style]. / Uroven' spontannykh i indutsirovannykh oblucheniem tsitogeneticheskikh povrezhdenii v limfotsitakh krovi detei v zavisimosti ot vozrasta i uslovii zhizni.

On blood lymphocytes the frequency of cell with micronucleus was examined in different population of children. The groups with different lifestyle living in different ecological situation, and their reaction to the radiation exposure was investigated. It was shown that when the increased level of spontaneous cells with micronucleus was observed the increased radiosensitivity of the cells was found. The connection between the increased spontaneous level of the damaged cells with the enhanced sensitivity to irradiation in adaptive doses are discussed.

[Increased radiosensitivity after irradiation of lymphocytes low adaptive doses]. / Fenomen povysheniia radiochuvstvutel'nosti posle oblucheniia limfotsitov v malykh adaptiruiushchikh dozakh.

The variability of blood lymphocyte reaction on the adaptive irradiation (0.05 Gy at first, then 1.0 Gy 5 h later) was investigated by micronuclei assay. Blood samples were obtained from 700 children. It was shown that in all groups studied there were children with enhanced radiosensitivity ("radiosensitivity syndrome"-RS) after exposure to adaptive low dose of radiation. The radiosensitivity syndrome occurred more often in groups of ill children; part of them was characterized by the enhanced blood content of immunoglobulin E, enhanced level of T helpers and T suppressors. A high spontaneous level of lymphocytes with micronucleus is a factor of radiosensitivity formation. The possible factors resulted in radiosensitivity syndrome are discussed.

[Radiation-induced adaptive response in children and effect of external and internal factors]. / Radioindutsirovannyi adaptivnyi otvet u detei i vliianie na nego vneshnikh i vnutrennikh faktorov.

434 children 3-13 old age and 4 adult people 20-40 old age have been investigated by the use of micronucleus (MN) assay and cytokinetical block by cytochalasin B on the ability to the adaptive response (AR). The frequency of lymphocytes with MN was scored in 1000 binucleated cells after irradiation in challenge dose of 1 Gy and together in adaptive (0.05 Gy) and challenge doses. The significance of differences have been determined by chi 2 criteria. It was shown that definite AR was determined in approximately 20 percent of "normal" children, when in adult population--in approximately 50 percent cases. The part of children with AR decreases in the groups of children with defects of development, with mentally deficiencies, with hard heredity (the parents are the chronic alcoholics), with the high pressure of genotoxic agents. It was discovered that in about the half of the children there is opposite effect observed-irradiation in adaptive dose induce the increasing of radiosensitivity; this effect is insignificantly expressed in adult people. The problems of genetic determination of AR, influence of different factors, the significance of AR as the phenomen of protection from extremal action have been discussed.

[Inability of large granular lymphocytes to recirculate through blood-lymph barriers]. / Nesposobnost' bol'shykh granuliarnykh limfotsitov retsirkulirovat' cherez gematolimfaticheskie bar'ery.

The level of large granular lymphocytes (LGL) has been determined in thoracic lymph duct and peripheral blood of cattle. It has been shown that in contrast to the blood, these cells are present in the lymph in minor quantities. Unlike blood LGL, lymph LGL have smaller granules in cytoplasm. It is concluded that LGL do not recirculate from blood to central lymph.