[Division of regulatory cellular systems (Lvov)]. / Viddilennia rehuliatornykh sistem klityny (L'viv).

Two departments of the A. V. Palladin Institute of Biochemistry of the National Academy of Sciences of Ukraine were founded in 1969 in Lviv. These were: the Department of Biochemistry of Cell Differentiation headed by Professor S. I. Kusen and Department of Regulation of Cellular Synthesis of Low Molecular Weight Compounds headed by Professor G. M. Shavlovsky. The Lviv Division of the A. V. Palladin Institute of Biochemistry of the National Academy of Sciences of Ukraine with Professor S. I. Kusen as its chief, was founded in 1974 on the basis of these departments and the Laboratory of Modelling of Regulatory Cellular Systems headed by Professor M. P. Derkach. The above mentioned laboratory which was not the structural unit obtained the status of Structural Laboratory of Cellular Biophysics in 1982 and was headed by O. A. Goida, Candidate of biological sciences. From 1983 the Laboratory of Correcting Therapy of Malignant Tumors and Hemoblastoses at the Institute of Molecular Biology and Genetics, Academy of Sciences of Ukraine (Chief--S. V. Ivasivka, Candidate of medical sciences) was included in the structure of the Division. That Laboratory was soon transformed into the Department of Carbohydrate Metabolism Regulation headed by Professor I. D. Holovatsky. In 1988 this Department was renamed into the Department of Glycoprotein Biochemistry and headed by M. D. Lutsik, Doctor of biological sciences. In 1982 one more Laboratory of Biochemical Genetics was founded at the Department of Regulation of Cellular Synthesis of Low Molecular Weight Compounds, in 1988 it was transformed into the Department of Biochemical Genetics (Chief--Professor A. A. Sibirny). In 1989 the Laboratory of Anion Transport was taken from A. V. Palladin Institute of Biochemistry, Academy of Sciences of Ukraine to Lviv Division of this Institute. This laboratory was headed by Professor M. M. Veliky. One more reorganization in the Division structure took place in 1994. The Department of Glycoprotein Biochemistry (Chief--M. D. Lutsik, Doctor of biological sciences) was renamed the laboratory and a new Department of Regulation of Cellular Proliferation headed by R. S. Stoika, Doctor of biological sciences, was founded. Since 1992 the Division has its present name and exists as an independent research institution with the rights of the Institute of the National Academy of Sciences of Ukraine. At present about 100 researchers work at the Division, among them there are 8 Doctors and 27 Candidates of biological sciences. For the period of the Division existence 6 doctor's theses and 30 candidate theses were defended.(ABSTRACT TRUNCATED AT 400 WORDS)

[Synergism in the stimulating effect of transforming growth factor beta and insulin on the substrate-independent proliferation of the CHO-719 cell line]. / Sinergizm stimuliruiushchego vliianiia transformiruiushchego faktora rosta beta i insulina na substratnezavisimuiu proliferatsiiu kletok linii CHO-719.

Cells of line CHO-719 from the Chinese hamster ovary have no epidermal growth factor receptors. It was detected that these cells were significantly stimulated to proliferate in a semisolid culture medium containing 0.33% agar by transforming growth factor beta (TGF-beta) in combination with insulin. This effect was even more pronounced when the cells of line CHO-719, adapted to the growth in the presence of low concentration (3%) of fetal bovine serum, were used. TGF-beta and insulin utilized separately exerted no influence on the cellular growth.

[Heat shock modulates the ability of A-549 cell line from human lung adenocarcinoma to regulate the intensity of DNA and protein biosynthesis by an autocrine mechanism]. / Teplovoi shok moduliruet sposobnost' kletok linii A-549 iz adenokartsinomy legkikh cheloveka autokrinno regulirovat' intensivnost' biosinteza DNK i belkov.

A-549 cells of human lung adenocarcinoma were subjected to heat shock (30 min, 44 degrees C) which caused substantial decreases in the rates of biosynthesis of the great bulk of cellular proteins with simultaneous increases in the synthesis rates of the 70 kDa protein predominantly localized in cell cytosol. By the 6th hour after the heat shock cessation this protein synthesis reached its maximum; by the 18th hour it was no longer detectable, while the protein itself was not denatured. During the recovery after the heat shock the ability of the serum-free culture medium conditioned by A-549 cells in autocrine regulation of [3H]thymidine incorporation into DNA and [3H]leucine incorporation into proteins changed also. The conditioned medium obtained within 1-3 hours after the heat shock did not influence the intensity of DNA synthesis, while the medium obtained 4-48 hours after the heat shock stimulated this process, the maximal effect (3.3-fold stimulation) being observed in the case of the 48-hour conditioned medium. Temporary (1 hour) acidification of the conditioned media down to pH 2.0 resulted in complete inhibition of the stimulating activity. Besides, these media acquired an ability to inhibit [3H]thymidine incorporation into the DNA of tracer cells. Study of effects of conditioned media on the rate of [3H]leucine incorporation into A-549 cell proteins revealed that the media obtained 1-4 hours after the heat shock inhibited this process, while the media obtained 6-18 hours thereafter stimulated it 1.2-2.1-fold. In the test systems under study temporary acidification of the media increased their stimulating influence on [3H]leucine incorporation into cellular proteins.

[The detection and characteristics of substances with fibroblast growth factor properties in the cells of loach embryos]. / Vyiavlenie i kharakteristika veshchestv so svoistvami faktora rosta fibroblastov v kletkakh zarodyshei v'iuna.

The substances extracted and partially purified from embryos of the loach (Misgurnus fossilis L.) at the late blastula stage are shown to possess properties characteristic of the fibroblast growth factor (FGF): 1) they bind with Heparin-Sepharose; 2) induce DNA synthesis in mouse NIH-3T3 and Swiss-3T3 fibroblast cell lines and in the primary culture of human embryonic fibroblasts; 3) have an apparent molecular weight of 15-16 kDa (according to SDS-electrophoresis); and 4) show positive reaction with antibodies against bovine basic FGF.

[The autocrine regulation of the proliferation of cell line A-549 under conditions facilitating the acidification of the culture medium]. / Autokrinnaia reguliatsiia proliferatsii kletok linii A-549 v usloviiakh, sposobstvuiushchikh zakisleniiu kul'tural'noi sredy.

The influence of serum-free media, previously conditioned by A-549 line cells of the human lung adenocarcinoma (c-medium), on the intensity of 3H-thymidine incorporation into DNA of the same cells was studied. It was found that, depending on the duration of conditioning (2, 4 and 6 days), the c-media were obtained with corresponding values of pH--7.2, 6.9 and 6.3, in the latter case the contact inhibition of cell growth being seen. On culturing the A-549 line cells in the c-medium at pH 7.2 and 6.9, the intensity of DNA biosynthesis was shown to be 2.4 and 1.8 times higher, respectively, compared to that under condition of the fresh serum-free medium. The cell culturing in the c-medium at pH 6.3 (here and in the case of pH 6.9 c-medium the media pH were made up to 7.2 before utilization) leads to the inhibition of DNA biosynthesis intensity in the cells. It was also detected that a temporary acidification of the pH 7.2 c-medium to pH 4.0 or 2.0, using, respectively CO2 bubbling or HCl titration, caused the growth inhibiting manifestation in this medium. The results obtained testify that the carcinoma cells of A-549 line are able to secrete into the cultured medium both stimulators and inhibitors of DNA biosynthesis, with a transforming growth factor beta being of primary importance among the latter.

[The effect of epidermal growth factor and insulin on the specific binding of transforming growth factor beta by NRK-49F and A-459 cell lines]. / Vliianie épidermal'nogo faktora rosta i insulina na spetsificheskoe sviazyvanie transformiruiushchego faktora rosta beta kletkami linii NRK-49F i A-459.

The influence of the epidermal growth factor (EGF, 5 ng/ml) and(or) insulin (1 microgram/ml) on the number of transforming growth factor beta (TGF-beta) specific binding sites in cells with different proliferative response on the TGF-beta action and on the affinity (Kd) of these sites to the ligand was studied. The total amount of TGF-beta binding sites (49,300 +/- 3000) in the cells of NRK-49F line, which are stimulated to proliferation by TGF-beta in the presence of EGF and insulin, decreased under the influence of the mixture of the latter factors (31,300 +/- 4000). In the presence of EGF, this amount did not change (40,900 +/- 6000), while in the presence of insulin it increased (81,200 +/- 9000). Kd of TGF-beta receptors (27.3 +/- 3.0 pM) in the above mentioned variants of experiments did not change. When the cells of A-549 line from human lung adenocarcinoma were investigated (the proliferation of these cells is inhibited by TGF-beta), it was shown that EGF induced the elevation of the amount of TGF-beta binding sites from 9700 +/- 700 to 21,600 +/- 900. Insulin or its mixture with EGF did not change that amount. EGF and(or) insulin did not influence the Kd (27.0 +/- 6.0 pM) of TGF-receptors in these cells. The obtained data are compared with the results of the influence of TGF-beta and of its combinations with EGF, and (or) insulin on the intensity of DNA synthesis and on proliferation in both the investigated cell lines.(ABSTRACT TRUNCATED AT 250 WORDS)

[Transforming growth factor beta--a new type of inhibitor of proliferation of normal and tumor cells]. / Transformiruiushchii faktor rosta beta--novyi tip ingibitora proliferatsii normal'nykh i opukholevykh kletok.

Based on literature data and on the results of the authors' investigation, the analysis of the transforming growth factor beta action as an inhibitor of proliferation of normal and tumor cells was carried out. The possible mechanisms of modulation of this action by other growth factors and by the cultivation conditions were considered. An attempt was made to determine the role of the transforming growth factor beta in the system of regulators of cell proliferation on the early stages of ontogenesis and during carcinogenesis.

[Detection of protease-sensitive factors with properties of beta-type transforming growth factor in early loach embryos]. / Obnaruzhenie proteazochuvstvitel'nykh faktorov so svoistvami transformiruiushchego faktora rosta beta-tipa v rannikh zarodyshakh v'iuna.

The blastoderm and yolk of Misgurnus fossilis L. late blastulae were subjected to acid-ethanol extraction and gel filtration on biogel P10 in 1 M CH3COOH. This resulted in isolation of polypeptides that induce substrate-independent proliferation of NIH-3T3 mouse fibroblasts in presence of epidermal growth factor. They did not compete for epidermal growth factor receptors of A-431 cells and inhibited substrate-independent proliferation of human lung carcinoma A-549 cells. These properties of the growth factor-like polypeptides from loach embryos are similar to those of transforming growth factor beta.

[The effect of transforming growth factor beta on the intensity of cellular DNA synthesis in relation to the type and conditions of cultivation]. / Vliianie transformiruiushchego faktora rosta beta na intensivnost' sinteza DNK kletok v zavisimosti ot ikh tipa i uslovii kul'tivirovaniia.

A study was made of the influence of transforming growth factor beta (0.05-50.00 ng/ml) on the intensity of 3H-thymidine incorporation in the DNAs of pseudonormal mouse fibroblasts of NIH 3T3 line, of cells of NRK-49F line from normal rat kidney, and of cells of A-549 line from human lung adenocarcinoma. The experiments were carried out in the absence and in the presence of epidermal growth factor (5 ng/ml), and(or) insulin (1 micrograms/ml), as well as in the presence of different concentrations of fetal calf serum, and while using different time of incubation of cells with the above mentioned growth factors. It was shown that depending on the culture conditions the transforming growth factor beta exerted stimulatory, inhibitory or no action on the intensity of DNA synthesis in the cells of the same type. An attempt was made to analyse the reasons, which may significantly determine the direction of regulatory influence of the transforming growth factor beta on DNA replication in the cells.

[The effect of the beta-type transforming growth factor and its combination with epidermal growth factor and insulin on substrate-dependent proliferation of normal and tumor cells]. / Vliianie transformiruiushchego faktora rosta beta-tipa i ego kombinatsii s épidermal'nym faktorom rosta i insulinom na substratnezavisimuiu proliferatsiiu normal'nykh i opukholevykh kletok.

The influence of different concentrations of a transforming growth factor of type beta (TGF-beta) and of its combinations with the epidermal growth factor (EGF) and insulin exerted on proliferation of different types of cells in the culture medium with semisolid agar was determined. The following cell lines were tested: mouse fibroblasts of NIH-3T3 and Swiss-3T3 lines, fibroblastic line NRK-49F from rat kidney, cells of A-549 line from human lung carcinoma, HT-1080 line from human fibrosarcoma, and PS-103 line (clone 384/5) from sarcoma stimulated by polychlorvinyl plate implantation to mouse. It is detected that TGF-beta alone does not affect the substrate-independent proliferation of pseudonormal lines of fibroblastic cells, but stimulates it significantly in sarcoma and inhibits in carcinoma cells. If EGF and/or insulin are added to the culture medium besides TGF-beta, certain proliferative effect specific of either type of pseudonormal and malignant cells is detected. The results of the action of TGF-beta, and of its combinations with the most important polypeptide growth factors on several types of cells of different origin may be useful for determination of regulatory functions of TGF-beta in cell proliferation in vivo to promote better grounding of its utilization in the practice of medicine.

[Epidermal growth factor stimulates protein kinase activity, specific for ribosomal protein S6 in loach embryo blastoderm]. / Epidermalnyi faktor rosta stimuliruet proteinkinaznuiu aktivnost', spetsifichnuiu k ribosomnomu belku S6, v blastoderme zarodyshei v'iuna.

The activity of a protein kinase specific to ribosomal protein S6 was determined in early loach embryos in basal conditions and after their treatment with epidermal growth factor (EGF). The cytosol of loach blastoderms isolated at the early gastrula stage possessed a high level of protein kinase activity catalysing incorporation of 0. 33 pmol.min-1.mg-1 of Pi into exogenous S6 protein of rat liver ribosomal 40S subunit. The treatment of embryos for 30 min with EGF (10 ng/ml) added to the incubation medium caused an 25% increase of total S6-kinase activity in cytosol compared with its counterpart in non-stimulated embryos. After chromatography of loach embryos cytosol on DE-52 three fractions possessing S6-kinase activity were revealed, which were eluted with 10 microM cAMP (I), 150 mM NaCl (II) and 300 mM NaCl (III), respectively. After treatment of blastoderms with EGF in the described conditions the enzymatic activity 2-fold decreased in fraction I, increased in fraction II 4-fold and remained practically unchanged in fraction III. The mitogen-stimulated kinase, apart from S6 protein, phosphorylated also casein and but not histone H1.

[Insulin-degrading neutral protease from plasma membranes of rat liver cells and erythrocytes]. / Insulinrasshchepliaiushchaia neitral'naia proteinaza plazmaticheskikh membran kletok pecheni i éritrotsitov krys.

Insulin-degrading neutral proteinase with molecular weight of 70 kDa was partly purified from the rat liver and erythrocyte plasma membranes. Incubation of membranes with [gamma-32P]ATP resulted in the enzyme phosphorylation. Intensity of this process greatly increased in the presence of insulin (100 microU/ml), and correlated with the elevation of the insulin-degrading activity in proteinase. Ca2+, Mn2+, dithiothreitol, cysteine were shown to have a stimulatory effect on insulin degradation; p-chloromercuribenzoate significantly repressed this process. Phenylmethylsulphonyl fluoride and soybean trypsin inhibitor did not affect the activity of the proteinase. It was concluded that the investigated enzyme was a calpain and may participate in the mechanism of insulin action.

[Insulin-degrading neutral proteinases of the plasma membrane of loach liver and embryo cells]. / Neitral'nye proteinazy plazmaticheskikh membran kletok pecheni i zarodyshei v'iuna, rasshchepliaiushchie insulin.

Insulin-degrading, Ca2+-activated, neutral proteinases of molecular weight about 150 kDa and 70 kDa were purified from plasma membranes of the loach liver and embryo cells. It was shown that dithiothreitol and cysteine enhanced the enzyme activity, whereas p-chloromercuribenzoate and iodoacetic acid inhibited its level. Incubation of isolated plasma membranes with 5'[gamma 32P]ATP resulted in phosphorylation of these proteinases. The intensity of the process increased under the influence of insulin (100 microU/ml), that correlated with a decrease in the activity of proteinase with molecular weight of 150 kDa and an increase in 70 kDa enzyme activity. The data suggest the existence of common regulatory pathways of insulin degradation in plasma membranes of the loach liver and embryo cells.

[Phosphorylation of lactate dehydrogenase by purified plasma membranes of loach liver cells: stimulation with insulin]. / Fosforilirovanie laktatdegidrogenzay ochishchennymi plazmaticheskimi membranami kletok pecheni v'iuna: Stimuliatsiia insulinom.

It is established that insulin enhances the ability of the loach liver plasma membranes to phosphorylate lactate dehydrogenase. In the case of insulin-treated plasma membranes the amount of incorporated 32P is more than 4 times higher than that of the basal level. It is concluded that insulin-stimulated plasma membrane-dependent phosphorylation of the enzyme is one of the possible molecular mechanisms of hormone action on intracellular metabolism.

[Polypeptide growth factors in animal embryogenesis]. / Polipeptidnye faktory rostav v émbriogeneze zhivotnykh.

Data about polypeptide growth factors in animal cells during embryogenesis and about the sensitivity of these cells to regulatory effect of the factors have been systematically reviewed. The conclusion was drawn that they play an important role in molecular mechanisms controlling cell proliferation and differentiation at the early embryonic stages. Information has also been provided about transforming growth factors and about the products of some proto-oncogenes, which are detected in the embryonic cells in amounts comparable with those in the transformed cells. However, this does not lead to malignant growth. Moreover, microenvironment of an intact embryo exerts an antitransforming influence on tumor cells. Studies of the "normalizing" effect of the embryonic cells may help in developing new methods of suppression of the malignant growth.

[Study of membrane glycoproteins of human erythrocytes using lectins]. / Issledovanie membrannykh glikoproteinov éritrotsitov cheloveka s primeneniem lektinov.

A method to study the glycoprotein composition of cell membranes, in particular of human red blood cells, has been developed. It includes the separation of membrane components by the SDS-polyacrylamide gradient slab gel electrophoresis, electroblotting of the phoretograms onto the nitrocellulose sheets and detection of glycoprotein fractions with FITC and peroxidase labeled lectins. PNA detected asialoglycoproteins with O-linked oligosaccharide chains, corresponding to all the PAS-positive bands of the phoretogram. SBA interacted more selectively and revealed only certain PAS-positive bands. Glycoproteins with N-linked carbohydrate chains were PAS-negative and can be identified only by the interaction with WGA, LCL, RCA. Group-specific agglutinins have shown that the ABO antigenic determinants are located in N-linked carbohydrate chains of membrane glycoproteins.

[Polypeptide growth factors from the loach embryo at the blastula stage]. / Polipeptidnye faktory rosta zarodyshei v'iuna na stadii blastuly.

Substances which (depending on their dose) stimulate [3H]thymidine incorporation into the trichloracetic acid insoluble fraction of the in vitro cultured mouse NIH-3T3 cells were isolated from the blastoderm and from the yolk of the teleost fish loach embryos (8-9 hours after the eggs' fertilization) by the acid-ethanol extraction method. In the medium which contained the 0.5% fetal calf serum with addition of 100 micrograms/ml of the above mentioned substances the stimulation of labeled thymidine incorporation into cellular DNA achieved 44-56% of the level which was detected in the presence of 10% serum. The pepsin and trypsin treatment led to a significant decrease in the mitogenic activity of the both investigated factors, that confirmed their polypeptide structure. The factors are sufficiently thermostable as they retain almost completely the activity after 30 min heating at 56 degrees C and lose less than 70% of their activity after boiling for 5 min. Gel-filtration of the growth factors from the loach embryos' yolk on the Biogel P-60 column has shown that the growth stimulating activity is mainly concentrated in the fraction which contains the polypeptides with molecular weight about 5 kDa. The significance of mitogenic factors in the loach embryos at the earliest stages of the embryonic development is discussed.

[Effect of Ca ions on thermoluminescence during lipid peroxidation]. / Vliianie ionov Ca na termoliuminestsentsiiu, soprovozhdaiushchuiu perekisnoe okislenie lipidov.

Addition of CaCl2 in the lipids extracted from oocytes, eggs, embryos and muscles of loach and rat's liver brings about a considerable increase of their thermoinduced peroxidation. In addition a shorter constituent appears in the luminescence spectrum which is not present in lipids unaffected by calcium ions. The kinetics of this luminescence coincides with that of peroxides accumulation in lipids.

[Influence of adrenaline on the transmembrane potential changes in developing loach embryos]. / Vliianie adrenalina na dinamiku transmembrannogo petentsiala razvivaiushchikhsia zarodyshei v'iuna.

Changes of transmembrane potential (TMP) in loach embryos developing for 7--8 hours in the medium supplemented with different adrenalin concentrations and in its mixtures with insulin or actinomycin D are studied. It was shown that value and mode of TMP changes, and its depolarizing effect and removal of periodic TMP oscillations in particular, which coincided with normal cycles of synchronous blastomere divisions depended on adrenalin level in the medium. Actinomycin D gradually eliminated the adrenalin influence on embryos only in the second half of the above mentioned period of their development, whereas insulin exerted its effect already at the early stages of embryogenesis.