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Biomedical subjects

G Acs

Publications and source records attributed to G Acs.

At least 91 records · Page 5Linked to original sources
PubMed1982

Inhibition of DNA methyltransferase and induction of Friend erythroleukemia cell differentiation by 5-azacytidine and 5-aza-2'-deoxycytidine.

Treatment of Friend erythroleukemia cells with the antileukemic drugs 5-azacytidine and 5-aza-2'-deoxycytidine leads to rapid, time-dependent, and dose-dependent decrease of DNA methyltransferase activity and synthesis of markedly undermethylated DNA. Since this DNA is at least partially methylated in vivo and serves as an excellent substrate for methylation in vitro, hypomethylation of DNA in analog-treated cells appears to result from the loss of DNA methyltransferase, rather than from an inherent inability of 5-azacytosine- substituted DNA to serve as a methyl acceptor. Inhibition of DNA synthesis blocks the loss of DNA methyltransferase activity while inhibitors of RNA synthesis do not, suggesting that the analogs must be incorporated into DNA to mediate their effect on the enzyme, and that minor substitution of 5-azacytosine for cytosine in DNA (approximately 0.3%) suffices to inactivate more than 95% of the enzyme in the cell. Several lines of evidence link changes in the pattern of DNA modification with differentiation. In this regard, it is significant that 5-azacytidine and 5-aza-2'-deoxycytidine act as weak inducers of erythroid differentiation of Friend erythroleukemia cells in the same concentration range where they affect DNA methyltransferase activity. For differentiation to proceed, the cells must be washed free of the drugs. Less than 24 h later, normal levels of DNA methyltransferase activity are restored and within 48 h, DNA isolated from the cells is not detectably undermethylated. This may in part explain why 5-azacytidine and 5-aza-2'-deoxycytidine induce differentiation in less than 15% of the population despite their initial profound effect on DNA methylation.

Animals↗
PubMed1982

Amplification of expression of hepatitis B surface antigen in 3T3 cells cotransfected with a dominant-acting gene and cloned viral DNA.

3T3 cells containing hepatitis B virus DNA sequences can be efficiently selected by exposure to methotrexate after cotransfection with cloned viral DNA and DNA coding for a methotrexate-resistant dihydrofolate reductase. More than 75% of methotrexate-resistant cells isolated after cotransfection with a head-to-tail tandem of the hepatitis B virus genome synthesized viral surface antigen. The antigen was released into the culture medium in the form of 22-nm particles with buoyant density of 1.20 g/ml. No other virally coded proteins were detected in the cells or the culture medium. Application of selective pressure by increasing the concentration of methotrexate resulted in an amplification of viral DNA sequences and a concomitant increase in the rate of synthesis and release of hepatitis B surface antigen. The ability to produce large amounts of surface antigen appears to be a stable trait and has been maintained in these cultures through more than 30 passages.

Animals↗
PubMed1980

Expression of cloned hepatitis B virus DNA in human cell cultures.

DNA was isolated from the ayw subtype of hepatitis B virus (HBV) that had been incubated in vitro with all four deoxynucleoside triphosphates in order to complete the circular viral genome by means of the endogenous DNA polymerase. The purified viral DNA was cleaved with EcoRI restriction endonuclease, inserted into the EcoRI site of plasmid pBR322, and cloned in Escherichia coli chi 1776. DNA from a clone, pHBV-1, that contained a 3200-base-pair insert of HBV DNA was cleaved with EcoRI and incubated with phage T4 ligase under conditions favoring intramolecular ligation. HeLa cell cultures exposed to this DNA showed marked cytopathic changes, accompanied by production of hepatitis B core and surface antigens, 11-14 days after subculture. Electron microscopic examination of anti-hepatitis B surface antigen immunoprecipitates from culture media of these cells revealed both 42-nm particles with central cores and 20-nm round particles. Although neither intact circular nor EcoRI-cleaved linear pHBV-1 DNAs evoked these effects in HeLa cells, both cytopathic changes and intranuclear hepatitis B core antigen were detected in HeLa cells infected with Dane particles.

Cloning, Molecular↗
PubMed1980

Hypomethylation of DNA during differentiation of Friend erythroleukemia cells.

DNA from mammalian cells has been shown to contain significant amounts of 5-methyl cytosine resulting from enzymatic transfer of methyl groups from s-adenosylmethionine to cytosine residues in the DNA polymer. The function of this modification is not known. We have found that DNA synthesized during chemically induced differentiation of friend erythroleukemia cells is hypomethylated, as measured by its ability to accept methyl groups transferred by homologous DNA methyltransferases in vitro. The extent of hypomethylation detected by this sensitive method is small, a decrease of less than 1.6 percent in 5-methylcytosine content. Hypomethylated DNA can be isolated from friend erythroleukemia cells grown in the presence of dimethyl sulfoxide, butyrate, hexamethylene-bis- acetamide, pentamethylene-bis acetamide, and ethionine. However, hypomethylated DNA is found only under conditions where differentiation is actually induced. DNA isolated from cells of a dimethyl sulfoxide- resistant subclone grown in the presence of that agent is not hypomethylated, although DNA of these cells becomes hypomethylated after growth in the presence of inducers that can trigger their differentiation. We also find that the DNA of friend erythroleukemia cells does not become hypomethylated when the cells are exposed to inducing agents in the presence of substances that inhibit differentiation. These results suggest a close link between genome modification by methylation and differentiation of friend erythroleukemia cells.

Animals↗
PubMed1980

Effect of tumor promoters on induction of aryl hydrocarbon hydroxylase in human lymphocytes.

The induction of aryl hydrocarbon hydroxylases in lymphocytes is dependent on their activation. The tumor-promoting phorbol esters which induce blast formation and DNA synthesis in lymphocytes enable polycyclic aromatic hydrocarbons to induce aryl hydrocarbon hydroxylases. Melittin, the major constituent of bee venom, acts synergistically with these phorbol esters in enhancing both lymphocyte activation and hydroxylase synthesis. Since aryl hydrocarbon hydroxylases convert procarcinogens to carcinogens these results suggest that tumor promotion by phorbol esters may be associated with their ability to affect the induction of these enzymes. This hypothesis is supported by the finding that phorbol and phorbol esters which lack tumor-promoting activity fail to enhance induction of aryl hydrocarbon hydroxylases in lymphocytes. However mezerein, although rather ineffective in promoting tumors, activates lymphocytes and permits polycyclic aromatic hydrocarbons to induce their hydroxylases effectively.

Aryl Hydrocarbon Hydroxylases↗
PubMed1980

L-Ethionine as an inducer of differentiation in human promyelocytic leukemia cells (HL-60).

The methionine analog, L-ethionine, induces morphological and biochemical changes in cultured HL-60 cells which are indicative of myeloid maturation. After 3 to 5 days of growth in the presence of L-ethionine, the majority of cells have enhanced phagocytic ability. The percentage of cells in the culture which bear complement receptors and which can respond to 12-O-tetradecanoylphorbol-13-acetate with respiratory burst activity increases more than 3-fold. Since the cells fail to become adherent and lose nonspecific esterase activity, we conclude that L-ethionine, like dimethyl sulfoxide, induces granulocytic differentiation of HL-60 cells.

Animals↗
PubMed1979

The effect of 12-O-tetradecanoyl-phorbol 13-acetate on the ribonuclease activity of circulating human lymphocytes.

12-O-Tetradecanoyl-phorbol 13-acetate is a very effective tumor promotor and inflammatory agent and can act as a mitogen for a subset of T lymphocytes. We report here that even short exposure of lymphocytes to 12-O-tetradecanoyl-phorbol 13-acetate changes the balance between the levels of neutral ribonuclease and ribonuclease inhibitor. The most dramatic change occurs in a B-lymphocyte-enriched population. We find that most, if not all, of the neutral ribonuclease activity in circulating lymphocytes is associated with this population and that this activity is lost with exposure to 12-O-tetradecanoyl-phorbol 13-acetate. Both 12-O-tetradecanoyl-phorbol 13-acetate and phytohaemagglutinin increase the level of ribonuclease inhibitor in T cells. However, phytohaemagglutinin has no effect on the ribonuclease or inhibitor level of the B-cell-enriched population.

B-Lymphocytes↗
PubMed1978

Effect of L-ethionine on macromolecular synthesis in mitogen-stimulated lymphocytes.

2--4 mM L-ethionine completely inhibits DNA synthesis in phytohaemagglutinin- or concanavalin A-stimulated lymphocytes even though it does not prevent the morphological changes characteristic of blast formation. Evidence is presented which indicates that complete commitment to DNA synthesis as well as a substantial increase in the rates of RNA and protein synthesis can occur in the presence of ethionine. Ethionine, however, does inhibit methylation of tRNA and prevents mitogen-induced increase in the activity of histone-modifying enzymes. All of these effects of exposure to ethionine are completely reversible. Removal of ethionine after 24 h or more of exposure results in a rapid, synchronous wave of DNA synthesis, an increase in the rate of methylation of RNA and an increase in activity of histone-modifying enzymes.

Acetyltransferases↗
PubMed1978

Co-cultivation of tumorigenic mouse melanoma cells with cells of a non-tumorigenic subclone inhibits plasminogen activator expression by the melanoma cells.

Clone B559 mouse melanoma cells are highly tumorigenic and produce plasminogen activator. Cells of clone C3471, a line obtained by continued growth of B559 cells in medium containing 5-bromodeoxyuridine (1 microgram/ml), have no plasminogen activator and are non-tumorigenic. When B559 cells are co-cultivated with C3471 cells, the ability of B559 cells to activate plasminogen is suppressed. Under these conditions cell fusion occurs. Lack of expression of plasminogen activators is not a consequence of cell fusion, inhibition of cell division or release of soluble inhibitors of either plasminogen activators or plasmin. No inhibitors of plasminogen activators could be demonstrated in association with sub cellular fractions of C3471 cells or with the C-type viral particles released from C3471 cells. Close contact between cells of the two lines is shown to be essential for suppression of plasminogen activation.

Animals↗
PubMed1978

Catabolism of 2-deoxyglucose by phagocytic leukocytes in the presence of 12-O-tetradecanoyl phorbol-13-acetate.

We have found that phagocytic leukocytes exposed to the tumor-promoting agent, 12-O-tetradecanoyl phorbol-13-acetate, efficiently release carbon-1 of 2-deoxyglucose in the form of CO2 with concurrent intracellular accumulation of a phosphorylated 5-carbon intermediate. In the absence of 12-O-tetradecanoyl phorbol-13-acetate, these cells release barely detectable amounts of CO2 from 2-deoxyglucose. 12-O-Tetradecanoyl phorbol-13-acetate, at a concentration of 1 ng/ml, has an immediate effect on CO2 release, which is temperature-dependent and linear with time and cell number. The ability of a number of phorbol ester-like compounds to enhance this catabolic pathway for 2-deoxyglucose correlates with their ability to act as tumor promotors and inflammatory agents. Although this effect of phorbol esters appears to be restricted to granulocytes, monocytes, and macrophages, the possibility arises that other mammalian cells are capable of catabolizing or can be induced to catabolize-2-deoxyglucose. Thus, 2-deoxyglucose decarboxylation should be considered whenever this analog of mannose and glucose is used as an indicator for sugar transport, especially when pharmacodynamic agents are present.

B-Lymphocytes↗
PubMed1977

Correlation between hypomethylation of DNA and expression of globin genes in Friend erythroleukemia cells.

This report identifies L-ethionine as an inducer of differentiation in murine erythroleukemia cells. When Friend erythroleukemia cells are grown in the presence of 4mM L-ethionine, globin mRNA accumulates and in 4-5 days, 25-30% of the cells in the culture contain hemoglobin. Incubation of the cells with bromodeoxyuridine prevents both ethionine-induced accumulation of globin mRNA and erythroide differentiation. At the concentration where L-ethionine acts as an inducer of FL cell differentiation it inhibits methylation of DNA and tRNA in vivo but does not prevent macromolecular synthesis or cell division. To establish whether a link existed between inhibition of a specific methyltransferase and activation of globin synthesis in FL cells, we examined the degree of hypomethylation of DNA and tRNA from FL cells induced to differentiate with dimethylsulfoxide and butyrate. In contrast to the tRNA from ethionine-treated cells, tRNA from cells induced by butyrate or Me2SO cannot be methylated in vitro using homologous enzymes. DNA isolated from cells exposed to any of the three inducers, however, was significantly hypomethylated when compared with DNA from uninduced cells. These data suggest that methylation of DNA may play a role in the regulation of gene expression.

Cell Differentiation↗
PubMed1976

Effect of sodium butyrate on lymphocyte activation.

Butyrate, in relatively low concentrations, has been shown to induce synthesis of enzymes, cause changes in cell morphology, and inhibit growth of a variety of mammalian cells in tissue culture (reviewed in [1]). In this communication, we report our observations on the effect of butyrate on lymphocyte activation. Butyrate completely and reversibly inhibits mitogen-induced blast formation. We present evidence that it does not interfere with the binding of mitogens, that it does not inhibit a number of the "early" reactions involved in activation, and that it does not affect ongoing DNA synthesis for an extended period of time. However, butyrate rapidly inhibits any increase in the rate of DNA synthesis.

Animals↗
PubMed1976

The reovirus replicative cycle.

Soon after entry into their host cells ssRNA viruses form doule-stranded replicative intermediates. Most RNA viruses exist only transiently in this double-stranded form. Double-strand formation occurs irrespective of whether the genome is of the same or complementary polarity to viral mRNA. In contrast, reoviruses contain dsRNAs from the outset. Single, and not double, strands are the intracellular intermediates through which genetic information is transferred from parental to progeny reoviruses. Double strands are the repositories in which the information is stored. We emphasize this distinction because it permits dsRNA-containing viruses to replicate conservatively, a mode of replication that is not shared by any other viruses. One important consequence of the conservative mode of replication is that cellular enzymes never gain access to the reovirus genome but only to its ssRNA precursors.

Binding Sites↗
PubMed1975

Immunological analysis of plasminogen activators from normal and transformed hamster cells. Evidence that the plasminogen activators produced by SV40 virus-transformed hamster embryo cells and normal hamster lung cells are antigenically identical.

Rabbits were immunized against the plasminogen activator released by SV4- virus-transformed hamster embryo cells. The resulting antiplasminogen activator immunoglobulin (APA-IgG) inhibited the enzymatic activity of the plasminogen activator produced by SV40-transformed hamster cells, and the plasmin-catalyzed release of these cells from the tissue culture dish. APA-IgG was not cytotoxic for these cells even in the presence of complement and did not inhibit their release of plasminogen activator. APA-IgG formed a single precipitin line in immunodiffusion plates using highly purified plasminogen activator as antigen. APA-IgG inhibited the plasminogen activator produced by newborn hamster lung cells and by an established diploid line (DON) of hamster lung cells, but did not inhibit plasminogen activators produced by normal or transformed hamster kidney cells or by cells of other species (mouse and human). We derive three major conclusions from these data: (a) There are several immunologically distinguishable forms (isozymes) of plasminogen activators in normal hamster tissues. (b) The plasminogen activators produced by normal hamster lung cells and by SV40 virus-transformed hamster embryo cells share antigenic determinants and are presumably the same isozyme. (c) The plasminogen activators produced by different hamster tumor cells do not share antigenic determinants and are presumably different isozymes.

Animals↗
PubMed1975

Correlated suppression by 5-bromodeoxyuridine of tumorigenicity and plasminogen activator in mouse melanoma cells.

The decrease of tumorigenicity by mouse melanoma clone B559 after growth in the presence of 5-bromodeoxyuridine (BrdU) has been correlated with a decrease in detectable cellular plasminogen activator. Reduction of both activities occurs after one to two cell divisions in the presence of this thymidine analog and is virtually complete within three to four cell cycles. These changes are fully reversible; four to five cell divisions in the absence of BrdU are sufficient to allow both tumorigenicity and plasminogen activator levels to return to normal. These results support the hypotheses that (a) the expression of a cellular plasminogen activator is closely associated with the transformation of normal to malignant cells and that (b) the suppression of tumorigenicity by BrdU reflects the capacity of this base analog to inhibit the expression of specialized functions which accompany the malignant state.

Animals↗