[Dynamics of elements distribution in blood, depending on age, by example of Moscow Region residents].

Elemental status of a person determines the qualitative and quantitative content of chemical elements in the human body. This marker allows us to estimate the level of imbalance of chemical elements and therefore health risks. The method for simultaneous quantitative and qualitative analysis of 67 elements in biomaterials has been proposed. The detailed elemental analysis of whole blood samples of 1711 healthy people (age range 0-100 years) of Moscow Region has been performed. A number of patterns of age-related changes of the element status conditionally healthy people has been estimated. Na content in the samples increased with the age of the person. Presumably, this result reflects the studied populations nutrition disorders associated with immoderate consumption of table salt. The maximum content of Ca was observed in blood samples of people age range 0-20 years (66-69 mg/kg), the Ca content in the blood samples of people age range 26-85 years was significantly lower (59-62 mg/kg). The maximum decrease of Ca was detected in blood samples of people age range of 85-100 years (57-59 mg/kg). Thisreductionin the concentration of Ca, apparently due to age-related changes of Ca balance, correlates with decrease of bone mineral density and bone mass. Iron content decreased in the blood samples of people age range 10-100 years from 480 to 390 mg/kg. Selenium content in blood of people age range 0-25 years linearly increased, remained stable high in the blood of people age range 25-55 years (0,13-0,136 mg/kg) and then gradually decreased. A graph of As content dependence from a person's age is a mirror image of the graph of Se content dependence from a person's age, which is evidence of the antagonistic effects of these elements. Graphic changes in the content of rare earth elements Eu and Ho reflect the unidirectional trend of these elements accumulation. The maximum content of these elements was observed in blood samples of people age range of 25-65 years. Perhaps a reduction of Eu and Ho in the age range 65-100 years age reflects a downward trend in bone mineral density and decrease in bone mass, which correlates with the Ca content in the blood depending on the age of people. The data obtained showed a significant increase of U and V in the blood of people age range of 85-100 years. The compounds of vanadium and uranium normally relatively easily filtered by the kidneys and excreted in the urine. This result seems to demonstrate age-related deterioration in the functioning of the excretory system. A list of recommendations for nutrition correction of elemental imbalance of the observed population has been proposed.

[Selective effect of inductors of apoptosis on the endoribonuclease activity of 26S proteasomes and alpha-RNP particles in K562 cells: possible involvement of 26S proteasomes and alpha-RNP in the regulation of RNA stability]. / Differentsial'naia reguliatsiia éndoribonukleaznoi aktivnosti 26S-proteasom i alpha-RNP-chastits pri deistvii induktorov apoptoza na kletki linii K562: vozmozhnoe uchastie alpha-RNP-chastits i proteasom v kontrole nad stabil'nost'iu RNK pri programmirovannoi kletochnoi gibeli.

It has been shown that endoribonuclease activity of alpha-RNP particles and 26S proteasomes are changed under the action of inductors of programmed cell death. Treatment of K562 cells with inductors of apoptosis--doxorubicin (adriamycin) and diethylmaleate--lead to a significant stimulation of RNAse activity of alpha-RNP and to reduction of proteasome RNase activity. The enzymatic activity under study has been shown to be specifically and selectively dependent on phosphorylation of subunits of alpha-RNP particles and 26S proteasomes. The characteristics of RNAse activity of different subpopulations of proteasomes differ. The specificity of a subpopulation of proteasomes exported from the cell has been demonstrated. Proteasome and alpha-RNP involvement in the coordinated control of stability of various specific messenger RNA molecules is suggested, and one of the mechanisms of this control might be the export of specific subpopulation of proteasomes from the cell.

[Immortalized cell lines from murine embryos are characterized by progressive destabilization of their karyotype structure]. / Immortalizovannye linii kletok, poluchennye iz transgennykh émbrionov myshei, kharakterizuiutsia progressiruiushzchei destabilizatsiei struktury kariotipa.

The karyotype structure was studied for three cell lines obtained from cells of transgenic murine embryos at early stages of their establishment. The first line was obtained from a transgenic embryonic explantant containing oncogen v-sis under promotor MMTV, two other lines originated from cells of transgenic embryos containing oncogen k51. The karyotypic analysis of G-banded metaphase chromosomes revealed deviations from the normal mouse karyotype as early as by the third passage of cultivation of independent embryonic cell lines that contained a foreign oncogene in their genome. The repeated analysis that involved 15-22 passages revealed similar abnormalities: variability and progression in chromosome number with the appearance of hyperpolyploid combinations, and a large number of rearranged chromosomes, both marker and unique ones. It is concluded that introduction of a foreign oncogene into murine cell genome leads to its enhanced and progressive non-specific destabilization. Oncogen v-sis produces a more valuable karyotype destabilization than oncogen k51.

Overexpression of P-glycoprotein is associated with a decreased mitochondrial transmembrane potential in doxorubicin-selected K562 human leukemia cells.

We tested the hypothesis of whether overexpression of P-glycoprotein (Pgp) could be coupled with changes in specific mechanisms of antioxidant defense (in particular, transition of mitochondrial transmembrane potential, MTP) in tumor cells chronically exposed to anticancer drugs known to exert their cytotoxicity via oxidative stress. We show elevation of Pgp associated with decreased MTP in doxorubicin-selected K562Dox subline as compared with parental K562 cells. The low MTP was not due to a fewer number of mitochondria in K562Dox cells, nor was it associated with altered content of Bcl-XL protein. We discuss a model for coordinated up-regulation of Pgp and MTP transition in cells that survived chemotherapy-induced oxidative stress.

Resistance to adriamycin in human chronic promyeloleukemia line K562 correlates with directed genome destabilization--amplification of MDR1 gene and nonrandom changes in karyotype structure. / Ustoichivost' kletok khronicheskogo promieloleikoza cheloveka linii K562 k adriamitsinu korreliruet s napravlennoi destabilizatsiei genoma--amplifikatsiei gena MDR1 i ne sluchainymi izmeneniiami v strukture genotipa.

Three independent subclones (B2, B3 and C9) of human myelogenous leukemia cell line K562 selected with adriamycin (ADM) were analysed. Cross-resistance of these ADM-resistant cells was examined for a resistance to the number of drugs including colchicine, actinomycin D and ethidium bromide. MDR 1 gene amplification in B3 and C9 subclones was detected using Southern-hybridization with specific probe. Additional genetical material was found in genomes of resistant cells by analysis of G-banded metaphase chromosomes. An extraordinarily long marker chromosome was observed in every C9 metaphase plate. The character of this chromosome G-banding suggests that it may be a derivative of chromosome 5 containing a large homogeneously staining region (HSR) in locus 5q15. Both B2 and B3 subclones expressed double minute chromosomes (DMs) in 5% of cells. In the course of a prolonged cultivation (about 2 years) of C9 cells in the presence of ADM a progressive karyotype destabilization was observed: the frequency of new markers formation in C9 cells increased, cells having additional copies of marker chromosome with HSR appeared, the length of HSR varied, coexistence of HSR and DMs being found in several C9 cells. These karyotypical changes may be regarded as patterns of genome destabilization due to the multidrug resistance of K562/ADM cells.

Alterations in immunoglobulin synthesis during adriamycin-induced apoptosis in mouse hybridoma cells.

Changes in the immunoglobulin synthesis in mouse hybridoma 1F7 cell line following exposure to the anticancer drug adriamycin were studied. It was shown that adriamycin in concentrations up to 0.1 microgram/ml causes dose-dependent reversible enhancement of immunoglobulin heavy gamma 2b chain synthesis, and to a lesser extent, kappa light chain synthesis. Enhancement of gamma 2b heavy chain synthesis accompanies an increase in Ig gamma 2b mRNA levels. Enhancement of DNA-binding activity of another B-cell specific protein, octamer transcription factor Oct-2 was also observed. Cell cycle analysis revealed accumulation of the ADR-treated cells in G2/M phase of cell cycle. These changes in hybridoma cells preceded the onset of internucleosomal DNA fragmentation and appearance of morphological characteristics of apoptosis. These findings suggest, that differentiation-related processes occur during the latent phase of apoptosis induced by adriamycin in mouse hybridoma cells.

[A new class of small RNP (alpha-RNP) containing antisense RNA in K-562 cells. I. Their characteristics and changes during erythroid differentiation]. / Novyi klass malykh RNP (al'fa-RNO), soderzhashchokh antismyslovuiu RNK, v kletkakh linii K-562. I. Kharakteristika i izmeneniia pri differentsirovke po éritroidnomu puti.

A new class of small RNP (alpha-RNP) has been detected and identified in nuclei and cytoplasm of A-562 erythroid leukemia cell line; these RNPs have a characteristic spectrum of proteins containing conservative and specific components and a special RNA component, which contains a small antisense component (alpha-RNA), a homolog of short dispersed Alu repeats. alpha-RNP is highly stable, tightly associated with chromatin in the nucleus, and is found in the free state in cytoplasm. The composition of nuclear and cytoplasmic alpha-RNP differ and have a specific pattern of changes in response to dimethylsulfoxide, an agent causing differentiation.

[New immortalized cell lines from transgenic rat embryos. I. The isolation and characteristics of cell lines from rat embryos with an injected v-mos oncogene coupled with or without the gene for genecitin resistance]. / Novye linii immortalizovannykh kletok iz transgennykh émbrionov krysy. I. Poluchenie i kharakteristika linii kletok iz émbrionov krysy s in''etsirovannym onkogenom v-mos odnovremenno s genom ustoichivosti k genetitsinu ili bez nego.

A collection of established cell lines was made by means of their explanation into 15 day old transgenic rat embryos. Some of these cell lines were characterized by measuring the cultivated population redoubling time, the saturation density and oncogenicity. A cytogenetic analysis was also carried out. The phenotypical analysis and studies of reproduction permit to define these lines as transformed immortalized non-oncogenic lines capable of contact inhibition. Cytogenetic studies were performed only on Mos N3, N6 and Mos+Neo N1, N6 lines. The karyotypes of cells in Mos lines were normal, and the karyotypes of cells in Mos+Neo lines had chromosomal markers (2 and 3, resp.). These markers result from arrangements of chromosomes 6, 9, 14, 15 and 17. the "thru deletions" of region q1 2qter (line N1) and region q22qter (line N6) of chromosome 15 were revealed in Mos+Neo lines by the summarized reconstruction karyotype method. We propose that these deletions of chromosome 15 and other chromosomes rearrangements may play an important role in transformation of cells from transgenic embryos in vitro, because RB1 antioncogene was mapped on rat chromosome 15.

[The effect of adriamycin and ethidium bromide on the Ca2(+)-dependent K channel of human erythrocytes]. / Vliianie adriamitsina i bromistogo étidiia na Ca2(+)-zavisimyi K-kanal éritrotsitov cheloveka.

The influence of cytotoxic agents adriamycine (AD) and ethydium bromide (EB) on Ca(2+)-dependent K+ channels of erythrocytes was investigated. The incubation of erythrocytes with agliconic part of AD (without aminosugar residue) increased Ca(2+)-dependent K+ efflux induced by low concentration of propranolol, while EB suppressed the activating effect of propranolol. EB, verapamil and triphluoroperazine inhibited Ca(2+)-dependent K+ efflux induced by high doses of propranolol. The incubation of erythrocytes with AD took off the inhibitory action of EB and verapamil but did not influence the blocking effect of triphluoroperazine. Both AD and EB did not influence Ca(2+)-dependent K+ channels induced by Ca(2+)-ionophore A23187, and Pb(2+)-dependent K+ efflux from erythrocytes. It is suggested that opposite effects of AD and EB on Ca(2+)-dependent K+ channels are due to activation by AD and inhibition by EB of system of Ca2+ transport into cells, but not on their action directly on K+ channel.

[The karyotypic variability of Chinese hamster CHLV-79 RJK cells characterized by multiple drug resistance resulting from the amplification of the mdr gene family]. / Izmenchivost' kariotipa kletok kitaiskogo khomiachka CHLV-79 RJK, kharakterizuiushchikhsia mnozhestvennoi lekarstvennoi ustoichivost'iu, obuslovlennaia amplifikatsiei genov semeistva mdr.

Variability in karyotype structure of Chinese hamster lung V-79 RJK cells and of their six cell sublines, selected for increasing concentrations of ethidium bromide (EB), was investigated in addition to the number of mdr gene copies in cells, both EB sensitive and resistant. It is shown that EB resistant cells exhibit cross-resistance to different drugs resulting from mdr genes amplification. Southern DNA blot hybridization has shown that in Vebr-2 cells (the 1st step of selection) the number of mdr gene copies increased by 10 times, whereas in Vebr-30 cells (the 6th step of selection) the number of mdr gene copies remained the same as in Vebr-2 cells. The level of mdr genes expression in Vebr-30 cells being higher than in Vebr-2 cells. In Vebr-2 cells, homogeneously and differentially stained regions (HSRs) were detected in loci 1p31 and 1q26 of chromosome 1 material (markers Z1 and Z6, respectively). On the following selection steps (prolonged cultivation or increased drug concentration) additional HSRs appeared in chromosome 2 (locus 2qter), in derivatives of chromosome 5 (marker Z7, locus Z7pter) and chromosome X (marker Z2, locus Z2qter). In the course of prolonged cultivation, chromosome 2 and derivatives of chromosomes 1, 2, 5 and X, in which HSRs were found, participated in the formation of new markers resulting from deletions, inversions, insertions and translocations of the chromosomal material. It is supposed that mdr genes amplification in V-79 RJK cells resistant to EB may be regarded as a factor inducing subsequent genome destabilization and eventual progressive changes in the karyotype structure.

[The kinetics of rhodamine 123 efflux from cells with multiple drug resistance under the action of energy metabolism inhibitors]. / Kinetika vykhoda rodamina 123 iz kletok s mnozhestvennoi lekarstvennoi ustoichivost'iu pri deistvii ingibitorov énergeticheskogo metabolizma.

A study was made of the effects of inhibitors of ATP synthesis on the process of rhodamine 123 (R-123) release from sensitive sp2/0-Ag14 cells, multidrug-resistant mouse myeloma spEBR-5 cells and hybridoma IF7, derived from spEBR-5 cells. It has been shown that IF7 cells are cross-resistant to ethidium bromide, colchicine, actinomycin D and adriamycin. However, hybridoma IF7 cells, compared to parental spEBR-5 cells, show a lower resistance index. When studying the dependence of the R-123 efflux rate on glycolysis intensity (effect of 2 mM 2-deoxyglucose) and on the level of oxidative phosphorylation activity (effect of 2 mM KCN and 30 microM dinitrophenol), the following distinctive properties of the R-123 transport system of IF7 cells (compared to spEBR-5 cells) were detected: 1) uptake of R-123 into IF7 cells is similar to that observed for the sensitive sp2/0-Ag14 cells; 2) efflux of R-123 from IF7 cells takes place more intensely; 3) R-123 transport is dependent on the rate of glycolysis and may be inhibited by KCN. It is found that 2,4-dinitrophenol inhibits the R-123 efflux from all the cells. Verapamil reverses the multidrug resistance both in spEBR-5 and IF7 cells. The mechanisms of multidrug resistance of cells are discussed.

[The isolation and phenotypic characteristics of heat-resistant cells from the Chinese hamster CHO-K1 line]. / Poluchenie i fenotipicheskie osobennosti termorezistentnykh kletok kitaiskogo khomiachka linii CHO-K1.

By sequential selection for a resistance to a short-term heating at 45 degrees C, two independent clones, CHSR5 and CHSR6, have been isolated from ovarian cells of the Chinese hamster (CHO-K1). A subline CHO-A40, capable of proliferation at 40 degrees C, has been obtained as mass culture by culturing CHO-K1 cells for 2 months at 40 degrees C. The lines obtained have no cross-resistance to the temperature used in their selection: the CHSR5 and CHSR6 cells are not capable of reproduction at 40 degrees C, while the CHO cells do not survive over a 60 min heating at 45 degrees C. This bears evidence that mechanisms, underlying the cell resistance under different thermal regimens used, may be different. The heat resistant sublines maintain their heat resistance for 6 months of cultivation at 37 degrees C. The acquisition of heat resistance by cells correlates with their high primary resistance, and with their resistance to colchicine and actinomycin D.

[The amplification and overexpression of mdr-family genes in ethidium bromide-resistant Chinese hamster CHO-K1 cells and in the hybrids of sensitive and resistant cells]. / Amplifikatsiia i sverkhékspressiia genov semeistva mdr v ustoichivykh k bromistomu étidiiu kletkakh kitaiskogo khomiachka CHO-K1 i v gibridakh chuvstvitel'nykh i ustoichivykh kletok.

Stable mutant cells Cebr-1 and Cebr-2, resistant to ethidium bromide (EB) in concentration of 2 micrograms/ml, have been isolated by a multistep selection in Chinese hamster ovary cells. It was shown that Cebr-1 and Cebr-2 cells acquired a cross-resistance to unrelated drugs. Stable changes in the structure of chromosomes 1, 2, 5 and 8 were revealed by karyological analysis. Overexpression and amplification of mdr genes were detected in Cebr-2 cells using Northern RNA and Southern DNA blot hybridization. Two independent hybrids Hybr-1 and Hybr-2 were obtained by fusion of Cebr-2 cells with Chinese hamster lung V-79 RJK cells, sensitive to EB. Hybr-2 cells were characterized by the same level of EB-resistance as Cebr-2 cells. Hybr-1 cells have a lower level of EB-resistance than Cebr-2 cells. Hybr-2 cells have demonstrated amplification and overexpression of mdr gene, the same as in Cebr-2 cells, whereas in Hybr-1 cells no mdr gene amplification was observed, but the level of mdr gene expression was higher than in sensitive cells. The data suggest that resistance of Chinese hamster cells to EB is mediated by amplification and overexpression of mdr genes.

[A cytogenetic analysis of the cells of spontaneously transformed LREC rat cell lines. I. Chromosomal rearrangements detected in the early stages of rat embryonic cell transformation in vitro]. / Tsitogeneticheskii analiz kletok spontanno transformirovannykh kletochnykh linii krys LREC. I. Perestroiki khromosom, vyiavliaemye na rannikh stadiiakh transformatsii émbrional'nykh kletok krys in vitro.

Seven spontaneously transformed cell lines LREC of rat embryo fibroblasts were obtained by an originally elaborated cloning technique (method 2T7). Six lines (1-6) were obtained from Rattus norvegicus cells, and one line (LREC-7) from Wistar rats. Method 2T7 was based on a serial propagation of rat embryo cells, brought to a "crisis" stage under conditions of a higher cell density and followed by the appearance of actively proliferating cell clones. These lines were analysed cytogenetically at the earliest stages using the Giemsa G-banding technique. In the karyotypes of six LREC (1-6) lines two abnormal chromosomes 7 were revealed: one marker chromosome M1-t(7; 19) results from translocation between chromosomes 7 and 19, the other marker chromosome M2--del(7) is a result of deletion of the second homolog of chromosome 7 in the q11.2 q22.1 loci; besides an extra normal homolog of chromosome 7 was revealed. There are only two marker chromosomes M2 in the LREC-7 line karyotype. Cells of LREC (1-3) lines could be transformed from the immortalized stage to the malignant one by the 30-45th passages. The cells of LREC (4, 7) lines became malignant at the 10-8th passages, resp. The rearrangements of chromosome 7 are supposed to be specific for LREC lines obtained by our method. A hypothesis is put forward that the translocation of chromosome 7 may play an important role for the immortalization of the rat embryo cells. The deletion of chromosome 7 may be associated with a malignant transformation of cells, as it is possible that the deleted loci have a recessive oncogene. Method 2T7 allows to obtain constantly spontaneously transformed cell lines of rat embryo cells with the least abnormal karyotype.

[The isolation and genetic characteristics of the new cell line of mouse myeloma spEBR-5 selected for ethidium bromide resistance and characterized by multiple drug resistance]. / Poluchenie i geneticheskaia kharakteristika novoi linii kletok mielomy myshi spEBR-5, otselektirovannykh na ustoichivosti k bromistomu étidiiu i kharakterizuiushchikhsia mnozhestvennoi lekarstvennoi ustoichivost'iu.

Stable mutant cells spEBR-5, resistant to ethidium bromide in concentration of 5 micrograms/ml, have been isolated by multistep selection in mouse myeloma cells sp2/0-Ag14. The spEBR-5 cells, selected with ethidium bromide, appeared to be cross resistant to unrelated drugs in connection with mdr/lb gene amplification and overexpression. Five specific chromosomal markers were detected in the karyotype of spEBR-5 cells as a result of chromosome structural rearrangement. No cytological manifestation of gene amplification such as HCR of chromosomes or DMs was found. A diffuse location of amplified sequences in chromosome(s) is suggested. The new mutant cell lines spEBR-5 can be used as a model for investigation of multidrug resistance mechanisms.

[The interaction of human natural killers with target cells of the K562 line and its sublines characterized by multiple drug resistance and thermoresistance]. / Vzaimodeistvie estestvennykh killerov cheloveka s kletkami-misheniami linii K562 i ee subliniiami, kharakterizuiushchimisia mnozhestvennoi lekarstvennoi ustoichivost'iu i teploustoichivost'iu.

Target cells, K562 strain and its sublines characterized by multiple drug resistance (MDR) do not differ in their susceptibility to human natural killer cells (NK) but MDR cells are more susceptible to cytotoxic action of lymphokine-activated cells (LAC) and to NK cells in the presence of a selective agent adriamycin. Target cells death is characterized by fragmentation of nuclear DNA. It has been established that K562 thermotolerant subclone is more resistant to NK and LAC than other clones. Heat shock protein synthesis may have a protective impact in target cells death during interaction with NK and LAC cells.

[The adaptive response of B cells in culture to the action of cytotoxic agents. I. The stimulation of immunoglobulin synthesis and secretion by myeloma cells, hybridomas and human lymphocytes under the influence of adriamycin and ethidium bromide]. / Adaptivnyi otvet B-kletok v kul'ture na deistvie tsitotoksicheskikh agentov. I. Stimuliatsiia sinteza i sekretsii immunoglobulinov kletkami mielom, gibridomy i limfotsitami cheloveka pod vliianiem adriamitsina i bromistogo étidiia.

Effects of cytotoxic drugs, doxorubicin (adriamycin) and ethidium bromide, on immunoglobulin production have been studied in B-cell hybridomas, myelomas and human lymphocytes. Doxorubicin, and in a lesser extent ethidium bromide, are shown to enhance immunoglobulin production both in drug-resistant and drug-sensitive cell lines. Enhancement of immunoglobulin production is accompanied with that of the number of antibody forming cells and with immunoglobulin synthesis in these cells.

[The adaptive response of B cells to the action of cytotoxic preparations in culture. II. Immunoglobulin interaction with adriamycin and ethidium bromide]. / Adaptivnyi otvet B-kletok na deistvie tsitotoksicheskikh preparatov v kul'ture. II. Vzaimodeistvie immunoglobulinov s adriamitsinom i bromistym étidiem.

The binding of adriamycin and ethidium bromide coupled to sepharose beads with intracellular and serum immunoglobulins is detected. These drugs do not bind with Fab and Fc fragments of immunoglobulin. The binding of ethidium bromide to immunoglobulins is competitive. Possible adaptive features of this phenomenon are discussed.