[The reentrant binomial model of nuclear anomalies growth in rhabdomyosarcoma RA-23 cell populations under increasing doze of rare ionizing radiation].

Distributions of nuclear morphology anomalies in transplantable rabdomiosarcoma RA-23 cell populations were investigated under effect of ionizing radiation from 0 to 45 Gy. Internuclear bridges, nuclear protrusions and dumbbell-shaped nuclei were accepted for morphological anomalies. Empirical distributions of the number of anomalies per 100 nuclei were used. The adequate model of reentrant binomial distribution has been found. The sum of binomial random variables with binomial number of summands has such distribution. Averages of these random variables were named, accordingly, internal and external average reentrant components. Their maximum likelihood estimations were received. Statistical properties of these estimations were investigated by means of statistical modeling. It has been received that at equally significant correlation between the radiation dose and the average of nuclear anomalies in cell populations after two-three cellular cycles from the moment of irradiation in vivo the irradiation doze significantly correlates with internal average reentrant component, and in remote descendants of cell transplants irradiated in vitro - with external one.

The karyotypic structure of cell populations in vitro as an integral system.

This review describes regularities of karyotypic variability maintaining karyotypic stabilization of continuous cell lines. Statistical analysis of individual karyotypes of "marker" and "markerless" cell lines show that survival of cell population in vitro is maintained by a certain ratio of cells with different structural variants of karyotype (SVK). Characteristic feature of karyotypic variability in the "markerless" cell lines during long-term cultivation under various conditions is dicentric formation due to telomeric associations. These dicentrics seem to form genetical structures providing adaptation to conditions in vitro of the cell population as an autonomous system. Correlations between the numerical variability reflecting in SVK, and structural variability (dicentric formation) are manifestations of an integral cell-populational function. Experimental data allow to suggest that integrity of the karyotypic structure of cell populations is maintained not only by selection of random variations, but also by programmed (adaptive) changes of karyotype. As a whole, in the cell population the state is realized that can be called karyotypic homeostasis; the observed phenomena characterize processes maintaining such homeostasis.

[Structural changes of the nucleolar organizing regions in rhabdomyosarcoma RA-23 tumor cells induced by okadaic acid]. / Strukturnye preobrazovaniia interfaznykh IaOR v opukholevykh kletkakh rabdomiosarkomy PA-23, indutsirovannye okalaikovoi kislotoi.

It has been recently shown that okadaic acid (OA), an inhibitor of dephosphorilation, is capable of inducing changes in the nucleolar organizer regions (AgNORs) of some mammalian cells. Our work was focused on studying the structural changes in AgNORs of tumour cells of rat rhabdomyosarcoma RA-23 by their exposure to 100 nM OA. A standard silver staining procedure of interphase AgNORs in tumour cells was used. We measured no less than 100 tumour cells in each clone. In the examined tumour cell populations, the index of interphase AgNORs varied from 1.54 to 4.35. A clear structure and form of AgNORs was not observed in 30% okadaic acid-treated tumour cells, as opposed to 10% of the control ones. AgNORs in these cells looked like a mixture of thin threads encompassing some dark dots lying, mostly, separately. Additional procedures of tumour cell staining with Giemsa and fluorescent dye Hoechst 33,258, respectively, revealed that such structures were not chromosomes. Meanwhile, the frequency of cells at the stage of prophase exceeded 3%, as opposed to the control, where the frequency of cells at this stage was less than 0.5%. Thus, we can conclude that we have detected specific changes in AgNORs and chromatin structure of okadaic acid-treated tumour cells.

[Internuclear chromosomal bridges and nuclei with protrusions in cellular populations of rat PA-23 rhabdomyosarcoma]. / Mezh''iadernye khromosomnye mosty i iadra s protruziiami v kletochnykh populiatsiiakh rabdomiosarkomy PA-23 krys.

Internuclear chromosome bridges (CB) and nuclear protrusions (NP) were investigated in cell populations of RA-23 rat rhabdomyosarcoma. A morphological classification of different types of CB and NP has been offered. The obtained data suggest a morphological similarity between CB and NP. It is likely that NP could arise from CB after its break. So, NPs may be regarded as remains of broken CB.

[Chromosome bridges and tailed nuclei in malignant cell populations]. / Khromosomnye mosty i "kvostatye" iadra v populiatsiikh zlokachestvennykh kletok.

Cell clones of the finite organospecific rat rhabdomyosarcoma RA-23 were selected in vivo for high and low frequency of interphase cells with chromosome bridges. After selection for high frequency of cells with bridges, the frequency of cells with anomalies of nuclear form sharply increased in the cell populations. These anomalies were manifested by long nuclear protrusions into the cytoplasm. This type of anomaly was termed tailed nuclei. In the studies populations of RA-23, the frequency of cells with "tailed" nuclei positively correlates with the frequency of interphase cells with chromosome bridges and the frequency of ana- and telophases with chromosome bridges. These parameters might be genetically associated: dicentric chromosomes form chromosome bridges in ana- and telophases of mitosis. Then, in some cases, the bridges are maintained, which result in the appearance of interphase cells with bridges, and, in other cases, the bridges are ruptured, which result in the appearance of cells with tailed nuclei.

[The possibility of performing artificial selection in vivo for a change in the mutability level in populations of mouse bone marrow stem cells]. / O vozmozhnosti provedeniia iskusstvennogo otbora in vivo na izmenenie urovnia mutabil'nosti v populiatsiiakh stvolovykh kostnomozgovykh kletok myshei.

Bone marrow cells of CBA males were transplanted to irradiated syngeneic females and after 13 days the mean frequency of blood erythrocytes with micronuclei, cloning efficiency of haemopoietic stem cells (HSC), and variability range of frequency of cells with micronuclei in spleen colonies were studied in 44 primary recipients. A wide range of variability has been shown for the indicators of mutability and cloning efficiency (0.1-1.8% for frequency of erythrocytes with micronuclei, 200-300-1500 for cloning efficiency, 0.1-0.5% for frequency of cells with micronuclei in spleen colonies of individual transplants). A weak negative correlation between the frequency of erythrocytes with micronuclei and the cloning efficiency was observed. We conclude that the difference in frequency of erythrocytes with micronuclei between HSC subpopulations is sufficiently high to carry out an artificial selection for high and low mutability in the course of transplantations. The cloning efficiency can be increased when spleen colonies are studied by micronuclear analysis. Data on the importance of such a selection in studying the behavior of "mutability" and "cloning efficiency" characters, immortalization, "aging", and death of HSC populations of CBA mice have been reported.

[Selection in vivo in cell populations of rat RA-23 rhabdomyosarcoma at the trait "frequency of cells with bridges"]. / Selektsiia in vivo v kletochnykh populiatsiiakh rabdomiosarkomy RA-23 krys po priznaku "chastota kletok s mostami".

Selection in vivo of cell clones of rat transplantable organospecific rhabdomyosarcoma RA-23 for increased and decreased frequencies of cells with chromosomal bridges (FCB) was performed. The initial average FCB in clones was 0.8% at a range of variation of from 0 to 3.0%. Selection for an increase, as well as for a decrease, in the FCB in RA-23 clones was effective. After one step of selection for an increase in the FCB, the average FCB increased to 3.0%. Thereafter, selection for an increase in the FCB was impracticable due to loss of transplantability in cell populations in which the FCB exceeded 5.0%. Over five steps of selection for a decrease in the FCB, the average FCB significantly decreased to 0.3% (P < 0.001). The heritability coefficient h2 of the trait FCB upon selection ranged from 0.25 to 0.30. The high and low FCBs attained by selection correlated with the index "frequency of cells with micronuclei." The population of RA-23 cells after selection for a decrease in the FCB differed from the population of cells of the original RA-23 strain by significantly lower karyotypic heterogeneity. The values of h2 obtained upon selection for the FCB and the effectiveness of selection for an increase, as well as for a decrease, in the FCB show that, with selection at the trait "frequency of cells with bridges," which characterizes the stability of the karyotype, existing cell strains can be subjected to karyotypic stabilization or destabilization.

[The heterogeneity of cell populations by the cytochemical characteristics of the cellular chromatin]. / Geterogennost' kletochnykh populiatsii po tsitokhimicheskim kharakteristikam khromatina kletok.

Cell heterogeneity of cytochemical characteristics of chromatin in the norm and after provocative factors in vitro (a heating at 45 degrees C, X-irradiation and joint actions of heating and X-ray irradiation) was investigated in the peripheral blood mononuclear cells of sheep and mouse, the lymphoid cell populations of the murine bone marrow, spleen and thymus. As a criterion of cytochemical heterogeneity were used the distinctions in the rate of staining of cell nuclei with DNA-specific dyes, that was registered by means of measurement of coefficient variation of the DNA-histogram basis peak, with a flow cytometry. Heterogeneity of cytochemical characteristics are dealt with showed a variability of spatial organization of interphase nuclei of single cells. The provocative factors bring about a decrease in the level of the cytochemical characteristics heterogeneity in the sheep mononuclear cells. In the murine cell populations studied no such equalizing of the marker of variability was revealed.

[Pathological mitoses in clones of the RA-2 subline of rat transplantable rhabdomyosarcoma selected for increased and decreased frequencies of the formation of spontaneous micronuclei]. / Patologicheskie mitozy v klonakh sublinii perevivnoi rabdomiosarkomy krys RA-2, otselektirovannykh na povyshennuiu i ponizhennuiu chastotu formirovaniia spontannykh mikroiader.

Cell substrains with increased and decreased spontaneous levels of micronuclei have been obtained by artificial selection. Clones of the substrains were investigated for mitotic division fidelity. About 200 mitoses were investigated in each clone. Frequencies of mitotic abnormalities such as ana- and telophases with bridge, chromosome and fragment delays at ana- and telophases, scattered chromosomes, chromosomes dislocated from spindle at metaphases and tripolar ana- and telophases were significantly higher in substrains with increased level of micronuclei. This finding indicates that reasons for arising of spontaneous micronuclei and alterations in mitotic division fidelity are the same or closely related. These reasons can be amplified as a result of the artificial selection.

[Interclonal and interpopulational differences in the frequency of spontaneous karyotypic changes in cell populations of transplantable rat rhabdomyosarcoma RA-2]. / Mezhklonal'nye i mezhpopuliatsionnye razlichiia po chastote spontannykh kariotipicheskikh izmenenii v kletochnykh populiatsiiakh perevivnoi rabdomiosarkomy RA-2 krys.

Rat organospecific transplantable RA-2 rhabdomyosarcoma substrains RA-2H (high metastatic potential), RA-2L (low metastatic potential) and RA-2T (thermal resistance) were investigated using single cell cloning technique for frequency of micronucleated cells (FMC). Significant interclonal and interpopulational differences were established. Average FMC for RA-2H was 2.96 +/- 0.13%; 5.94 +/- 0.24% for RA-2L and 1.89 +/- 0.12% for RA-2T. Clones showed 5-10 times differences in FMC in each substrain. FMC versus growth time and clone size was studied in RA-2H: average FMC dropped from 2.7% on day 9 to 0.5% on day 20 after administration and the lowest values were recorded in larger clones at each stage. Perspectives of FMC studies in tumor cell populations are discussed.

[The dependence of the incidence of encountering cells with micronuclei in the experimental metastases of rat rhabdomyosarcoma on the duration of the growth and on the size of these metastases]. / Zavisimost' chastoty vstrechaemosti kletok s mikroiadrami v éksperimental'nykh metastazakh rabdomiosarkomy krys ot dlitel'nosti rosta i razmerov étikh metastazov.

Cell suspensions of rat transplantable rhabdomyosarcoma RA-2 were injected to white rats. The experimentally induced lung metastases (ELM) were estimated according to the frequency of micronucleated cells (MNC) that appeared 9-20 days after injection (a. i.). The frequency of MNC in ELM decreased from 2.7% (9 days a. i.) to 0.7% (20 days a. i.). The ELM which were of the same size but were examined at different time after injection had equal frequencies of MNC. At each investigated time after injection the least frequencies of MNC were observed in the ELV of the biggest size. The data provided allow to infer that the growth time and site of ELM may influence the spontaneous karyotype instability rates.

[Changes in the frequency of cells with micronuclei in cell populations of rat transplantable rhabdomyosarcoma RA-2 as a result of selection for an increased and a decreased frequency of such cells]. / Izmenenie chastoty kletok s mikroiadrami v kletochnykh populiatsiiakh perevivnoi rabdomiosarkomy RA-2 krys v resul'tate otbora na povyshennuiu i ponizhennuiu chastotu takikh kletok.

Artificial selection for increasing and decreasing frequencies of cells with micronuclei (FCM) was conducted in populations of cell clones of the rat transplantable RA-2 rabdomyosarcoma. The selection was effective in both the opposite directions. The average FCM increased from 3.3 +/- 0.2 to 6.9 +/- 0.6% and decreased from 3.3 +/- 0.2 to 1.6 +/- 0.2%. Heritability coefficient, h2, varied from 0.2 to 0.4. In the cell population with increased FCM, 37% of anaphases and early telophases had lagged chromosomes and fragments. In contrast, in the cell population with decreased FCM, lagging were observed in 11% of cases. The DNA-content in G1 cell varied greatly in cell population with increased FCM (P < 0.01). The selection for increased FCM has led to a significant decrease in metastatic capacities of cells.

[Clonal analysis of karyotypic instability and metastatic potential in tumor cell populations]. / Klonal'nyi analiz kaiotipicheskoi nestabil'nosti i metastaticheskogo potentiala v populiatsiiakh opukholevykh kletok,

Rat organospecific transplantable RA-2 rhabdomyosarcoma strains RA-2H and RA-2L were investigated using single cell cloning technique for "Micronuclear containing cell frequency" (MCF) and "Metastatic potential" (MP). The RA-2H was obtained from RA-2 during selection for increasing metastatic potential, whereas the RA-2L was the result of selection in opposite direction. MP was assessed by lung colony formation technique and in the case of RA-2H, it was one hundred times greater than in the case of RA-2L (2490 +/- 280 and 22 +/- 11 lung colonies i.v. after injection of 10 cells, respectively). Average MCE for RA-2H was 3 and for RA-2L 5.3%. Also, significant differences were observed between these substrains in percentage of anaphase and telophase cells with bridges and fragments (24 +/- 4 for RA-2H and 56 +/- 5% for RA-2L, P less than 0.05). The selection of RA-2H clones with high level of MCF was successful. In the population obtained MCF was increased up to 7.8% and MP decreased (79 +/- 44). So, the selection for decreasing of MP led to significant increase in genome instability characteristics and the selection for increasing of MCF--to the abrupt decline of MP, which proved the existence of negative correlation between these two characters in populations studied. It is concluded that correlation between malignancy and genome instability in tumor populations with different levels of malignancy is not, as a rule, positive.

[The radiosensitivity and thermoresistance of the cells of substrains of rhabdomyosarcoma RA-2 in rats]. / Radiochuvstvitel'nost' i termoustoichivost' kletok subshtammov rabdomiosarkomy RA-2 krys.

The data obtained show that clonogenic RA-2T cells are 2-3 times more thermoresistant than clonogenic cells of the original thermosensitive RA-2T strain as estimated by D0 value upon heating up to 43-45 degrees C. After X-irradiation of rat rhabdomyosarcoma, a decrease in the capacity of forming pulmonary colonies is more pronounced in cells of the thermosensitive RA-2 strain cells than in those of the thermoresistant strain RA-2T (D0 = 1.6 Gy and D0 = 2.4 Gy, respectively). In all appearance, within one and the same tumor cell population, the hereditarily thermoresistant cells are more radioresistant than the thermosensitive ones.