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Biomedical subjects

M A Lea

Publications and source records attributed to M A Lea.

At least 37 records · Page 2Linked to original sources
PubMed1989

Interrelationship between sodium cyanate and pH in the regulation of tumor cell division.

Previous studies have suggested that the selective inhibitory effects of sodium cyanate on tumor metabolism in vivo may be related to a lower interstitial pH in tumors. In the present work, the influence of extracellular pH on the actions of sodium cyanate was studied with one rat hepatoma cell line (HTC) and two human colon tumor cell lines (HT29 and LS174T) and with rat hepatocytes to determine if the effects are accompanied by changes in intracellular pH. With some tumor cells, an inhibition of cell proliferation was observed when the cells were exposed to an acidic medium (pH 6.6). However, the LS174T line of human tumor cells divided at pH 6.6 essentially as fast as at pH 7.4. In the concentration range of 0.02-0.1 mg/ml, a greater inhibitory effect of cyanate on cell proliferation was observed at the lower pH. Intracellular pH was found to be influenced by the sodium ion concentration of the medium to a similar degree in the three tumor lines that were examined. The intracellular pH was found to be significantly affected by cyanate in rat hepatocytes and in two of the tumor cell lines (HT29 and LS174T). The data suggested that not only does extracellular pH influence the inhibitory effect of cyanate on tumor cell proliferation but also that cyanate can affect the regulation of intracellular pH in normal and neoplastic cells.

Animals↗
PubMed1988

pH-related effects of sodium cyanate on macromolecular synthesis and tumor cell division.

In past work, the selective effects of sodium cyanate on macromolecular synthesis in tumors have not been seen with cells in culture. We have explored the possibility that differences in the response of tumor cells to cyanate in vivo and in vitro may be related to the pH in the environment to which cells are exposed. When rat hepatoma (HTC) cells were incubated with sodium cyanate (0.25 mg/ml), there was a greater inhibition of precursor incorporation into RNA and DNA with a decrease in pH from 7.4 to 6.6. At pH 7.4 there was no significant effect of sodium cyanate on the incorporation of [3H]leucine into protein of rat hepatocytes and HTC cells, but at pH 6.6 there were decreases of 50% or greater. The time of response and the reversibility of the inhibitory effects of sodium cyanate were not those anticipated from carbamoylation of amino groups but were compatible with modification of sulfhydryl groups. The uptake of [14C]sodium cyanate in HTC cells and human colon cancer (HT29) cells was greater at pH 6.6 than at 7.4. Over a period of 4 days there was a slower rate of cell division by HTC and HT29 at pH 6.6 than at pH 7.4. The addition of sodium cyanate caused a further reduction in the rate of proliferation, and at a concentration of 0.25 mg sodium cyanate/ml there were decreases in cell numbers. The data suggested that a lower interstitial pH in tumors than normal tissues would result in greater sensitivity to inhibitory effects of sodium cyanate on macromolecular synthesis.

Animals↗
PubMed1988

Selective modulation of nucleotide levels in rat liver and hepatomas by high-orotate or arginine-deficient diets and by carbamoylating agents.

Transplanted Morris hepatomas in Buffalo-strain rats were found to be resistant to the changes in ribonucleotide levels in rat liver caused by a high-orotate diet or an arginine-deficient diet. The increase in UTP levels and decrease in ATP levels seen in the livers of rats on a 1%-orotate diet were less marked in the livers of BUB- and DBA-strain mice on this diet. Although the changes were less than in rat liver, there was a 2-3-fold increase in UTP concentration in the livers of mice on the high-orotate diet. However, there was a similar response in nucleotide levels in the two species when the animals were maintained on an arginine-deficient diet, and there was a greater than 10-fold increase in the UTP level in the livers of both rats and mice. These diets had much less effect on the levels of deoxyribonucleotides than of ribonucleotides. In contrast to the insensitivity of hepatomas to dietary modulation of nucleotide levels, treatment of hepatoma-bearing rats with carbamoylating agents (sodium cyanate and 2-chloroethyl isocyanate) caused decreases in the levels of nucleotides in the tumors which were generally greater than in host livers. For example, 2-chloroethyl isocyanate depressed ATP levels in the Morris hepatomas 5123C and 20 under conditions in which there was no significant effect on host liver ATP. The data revealed selective modulation of nucleotide levels in normal and neoplastic liver which may be achieved by either dietary modification or drug treatment.

Animals↗
PubMed1987

Action of carbamoylating agents on the uptake of metabolites in hepatomas and liver.

The effects of 2-chloroethylisocyanate, ethylisocyanate and sodium cyanate on the uptake of isotope-labeled thymidine, leucine and H2O were compared in rat liver and hepatomas. The data suggested that carbamoylating agents may have a common property of inhibiting uptake of compounds in hepatomas under conditions in which there is a smaller effect or no action in the liver of tumor-bearing rats. The distinction between tissues may have been mediated, in part, through effects on tumor circulation and was less apparent when isolated cells were studied in vitro. Preferential inhibitory effects of carbamoylating agents on the uptake of leucine and H2O were also observed with a murine hepatoma, but they were not as great as with rat hepatomas.

Animals↗
PubMed1987

Chromatin solubilization in rapidly growing hepatomas.

The presence of substantial amounts of DNA in the cytosol fractions prepared from rapidly growing transplanted rat hepatomas was confirmed. Agarose gel electrophoresis of this DNA from the Morris hepatomas 7288CTC, 7777, and 7800 suggested that it corresponds primarily to nucleosome monomers with smaller amounts of nucleosome dimers and trimers. Despite the evidence for solubilization of chromatin in rapidly growing hepatomas the potential DNA endonuclease activity was less in nuclei from these tumors than in nuclei from more slowly growing hepatomas or host livers. It was suggested that focal necrosis in rapidly growing solid tumors may be associated with the influx of divalent cations and the activation of DNA endonuclease activity.

Animals↗
PubMed1987

Orotate uptake and metabolism in normal and neoplastic tissues.

The uptake of [3H]orotate was greater in mouse liver than in hepatoma but the difference was less marked than in the rat. Of the tissues examined, a high uptake of [3H]orotate was restricted to the liver and kidney in rat, mouse and guinea-pig. We confirmed that a high orotate diet greatly increases the ratio of UTP to ATP concentration in rat liver but we observed that there is little change of this nucleotide ratio in kidney. Evidence was obtained for a different pattern of orotate metabolism in rat liver and kidney.

Animals↗
PubMed1987

Effects of carbamoylating agents on tumor metabolism.

Carbamoylation of macromolecules occurs by the displacement of hydrogen on several groups, but the most stable addition at neutral pH is on amino groups. This reaction occurs predominantly with proteins and results from the administration in vivo of inorganic cyanate or organic isocyanates. The latter act more rapidly, but also are more rapidly hydrolyzed in aqueous solution. This instability has been a factor limiting study of the pharmacological properties of organic isocyanates. However, organic isocyanates are released from some nitrosoureas of value in cancer therapy such as 1,3-bis(2-chlorethyl)-1-nitrosourea (BCNU) and 1-(2-chloroethyl)-3-cyclohexyl-1-nitrosourea (CCNU). The carbamoylating activities of BCNU and CCNU are generally considered less significant than their alkylating activity in the action of these drugs on tumors, but carbamoylation may serve to inhibit DNA repair. There is evidence that carbamoylating agents can exert selective inhibitory effects on metabolite uptake and macromolecular synthesis in neoplastic tissues. Such selectivity is much more notable in vivo than in vitro. In the case of cyanate, the selectivity in vivo has been variously attributed to a requirement for metabolic activation, to selective effects on circulation in solid tumors, and to diminished pH in tumors. It is the distinction between such factors and the identification of critical cellular targets which provide major challenges in present studies on the effects of carbamoylating agents on tumor metabolism.

Animals↗
PubMed1987

Effects of carbamoylation with alkyl isocyanates on the assay of proteins by dye binding.

The effect of carbamoylation with alkyl isocyanate was used both to monitor the stability of the isocyanates and to study the influence of charge modification on protein assay. Carbamoylation of poly (L-lysine) with methyl isocyanate, ethyl isocyanate and 2-chloroethyl isocyanate was observed to decrease binding of methyl orange. The data emphasized the lability of alkyl isocyanates and indicated the importance of preparing aqueous solutions at low temperatures for studies on protein carbamoylation. After carbamoylation of several proteins, there was decreased metachromasia on binding to Coomassie Blue G. Poly (L-lysine) and H1 histone showed anomalous behavior in that with low concentrations of Coomassie Blue G the metachromasia was increased by carbamoylation, but at high concentrations of the dye the metachromasia was decreased by carbamoylation. In contrast to some reports in the literature, the data indicated that there is not always a simple relationship between the positive charge on a protein and the interaction with anionic dyes.

Azo Compounds↗
PubMed1987

Relationship of H1(0) histone to differentiation and cancer.

The H1(0) histone was first described by Panyim and Chalkley in 1969 as a new electrophoretic band found with histones of non-replicating tissues. Tissues which are active in DNA replication such as ascites tumor cells or thymus cells were reported to lack this band. In this respect the H1(0) histone differs from the bulk of histones which are generally maintained in a constant ratio with respect to each other and to DNA. An inverse relationship between H1(0) histone levels and growth rate was suggested by the decrease in H1(0) histone concentration during regeneration of the pancreas and liver. The synthesis of H1(0) is unusual but not unique in that, unlike the major histone species, it is not restricted to the S phase of the cell cycle. Although there is a general trend for the levels of H1(0) histone to be lower in neoplastic than normal tissues, exceptions have been observed. Compounds such as sodium butyrate and dimethylsulfoxide, which can induce differentiated properties in neoplastic cells, can bring about the accumulation of increased amounts of H1(0) histones. The relative magnitude of these effects exhibits cell-type specificity. There are two H1(0) histone subtypes (a and b) with ratios which differ according to the tissue examined and whose relative importance is not known. The levels of H1(0) histone appear to be more closely related to the degree of differentiation than to the proliferative activity of cells.

Animals↗
PubMed1986

Effects of sodium cyanate in mice bearing B16 melanoma.

Sodium cyanate injected IP at a dose level of 200 or 250 mg/kg caused a 90% or greater inhibition of the incorporation of [3H]thymidine into DNA of B16 melanoma transplanted SC in mice. Despite the inhibitory effect of sodium cyanate on precursor incorporation into DNA, no significant effect on host survival was observed when sodium cyanate was administered as a single agent in the diet, in drinking water, or by IP injection to mice that had received IP transplants of B16 melanoma. The action of melphalan and 1-(2-chloroethyl)-3-cyclohexyl-1-nitrosourea (CCNU) in prolonging the survival time of melanoma-bearing mice was not enhanced by combined treatment with sodium cyanate. However, combined injections of sodium cyanate and 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) increased the survival of tumor-bearing mice significantly more than injections of BCNU alone at a lower dose than the maximum tolerated one. These data and other studies suggest that B16 melanoma may be less responsive to the action of sodium cyanate than are murine leukemic cells or rat hepatomas.

Animals↗
PubMed1986

Plasma protein alterations in AIDS and hemophilia A.

Plasma proteins of patients with AIDS, ARC, hemophilia A, and some viral infections were studied using various electrophoretic techniques, and compared to healthy control subjects. On isoelectric focusing (IEF) gels the most prominent and consistent finding was a marked increase in a basic protein band (AABP) at pI approximately equal to 9.0 in plasma samples derived from AIDS and hemophilia A patients. Using SDS gels, we noted an increased amount of protein in the 90 KD region in AIDS patients as compared to control subjects. On two dimensional gels (2D gels) basic protein(s) with pI congruent to 8.1-9.0 and molecular weight of approximately 90 KD were noted to be increased. In addition, a basic protein of 27 KD along with two acidic proteins in the low molecular weight region were also elevated in AIDS plasma. Although a limited number of samples were analyzed, it seems probable that a number of proteins are altered in AIDS plasma. AABP stained positive with periodic acid Schiff (PAS) reagent, indicating that it is a glycoprotein. This protein did not bind to an anti-IgG sepharose column, suggesting that it is not an immunoglobulin. The purified protein also did not react with antibody to fibrinogen and hemoglobin beta-chain. There are many changes in the plasma protein pattern of AIDS and hemophilia A patients as compared to normal controls.

Acquired Immunodeficiency Syndrome↗
PubMed1986

Influence of carbamoylation on some analytical properties of basic polypeptides.

The effect of carbamoylation on the assay or identification of histones and polylysine was investigated. Incubation with sodium cyanate decreased the positive charge on these polypeptides as judged by changes in the binding of methyl orange or the electrophoretic mobility. Histones in chromatin appeared less accessible to carbamoylation than isolated histones. Carbamoylation of proteins under conditions in which there was little or no effect on the Lowry procedure could affect their assay by methods utilizing metachromasia with Coomassie Blue G. The Bradford assay has low sensitivity for Hl histone and polylysine but this can be increased by preincubation with sodium cyanate. More extensive carbamoylation of polylysine caused decreased sensitivity which was the only response seen with core nucleosomal histones and bovine serum albumin when preincubated with sodium cyanate. It was concluded that the sensitivity for Hl histone and polylysine in assays dependent on metachromasia with Coomassie Blue G may be changed by factors which decrease the positive charge on these polypeptides.

Animals↗
PubMed1985

Disparity in the effects of two N-methyl nicotinamides on poly(ADP-ribose) synthetase and macromolecular synthesis in hepatomas.

The inhibitory effects of nicotinamide analogs on the activity of poly(ADP-ribose)) synthetase were compared to effects on precursor incorporation into macromolecules in three lines of hepatoma cells (Morris hepatomas 5123C, 7777 and HTC). N'-methylnicotinamide was a less effective inhibitor of poly (ADP-ribose) synthetase than was 1-methylnicotinamide while both these compounds had smaller inhibitory effects on the enzyme than were seen with nicotinamide or 3-aminobenzamide. On the other hand, the incorporation of [3H]thymidine into DNA and of [3H]uridine into RNA were inhibited by N'-methylnicotinamide in the concentration range 2-20 mM but not by 1-methylnicotinamide. Under the conditions examined there were no significant effects on the incorporation of [14C]lysine and [3H]leucine in hepatoma cells. The data indicated that the inhibitory effect of N'-methylnicotinamide on nucleic acid synthesis may be unrelated to action on poly (ADP-ribose) synthetase.

Animals↗
PubMed1984

Factors affecting the assay of histone H1 and polylysine by binding of Coomassie blue G.

Although a mixture of calf thymus histones or the H2B and H4 fractions gave values in the Bradford assay that were similar to those for bovine serum albumin, low values were obtained with histone H1 and polylysine. The assay was found to be more sensitive for histone H1 and polylysine if the phosphoric acid concentration in the reagent was decreased or the polypeptide solutions contained low concentrations of nonionic detergents. Substitution of perchloric acid or hydrochloric acid for phosphoric acid and alcohol in the Coomassie blue G reagent provided an assay that was similar in sensitivity for histone H1 and a total histone mixture. It was suggested that the degree of metachromasia with Coomassie blue G may serve as a probe of conformational changes in histone H1.

Animals↗
PubMed1984

DNA in cytosol fractions obtained by differential centrifugation of homogenates of rapidly growing liver tumors.

Preliminary work revealed that the cytosol fractions obtained by differential centrifugation of homogenates of the rapidly growing Morris hepatomas 7777 and 7288CTC contained substantially more DNA than could be detected with the more slowly growing hepatoma 5123C or the liver of tumor-bearing rats. The amount of DNA in the cytosol fractions of the rapidly growing hepatomas was decreased by concentrations of cations which are known to promote precipitation of solubilized chromatin. Data on the incorporation of [3H]thymidine into DNA of nuclear and cytosol fractions and the time-dependent effects of hydroxyurea were compatible with initial nuclear synthesis and later release of the DNA seen in the cytosol fractions. The amount of this DNA was increased in larger tumors and was less in isolated cells than in solid tumors. Postmortem changes in the liver and hepatoma 5123C, but not in hepatomas 7777 and 7288CTC, resulted in a large increase in DNA appearing in the cytosol fraction. The results indicated that the measurement of DNA in cytosol fractions may serve as a guide to the existence of necrosis in tumors, but is not always proportional to the degree of necrosis.

Animals↗
PubMed1984

Effects of nicotinamide and structural analogs on DNA synthesis and cellular replication of rat hepatoma cells.

The effects of nicotinamide and structural analogs on DNA synthesis were studied in rat hepatoma (HTC) cells. Inhibitory effects of these compounds were observed on DNA synthesis as judged by the incorporation of [3H]-thymidine into DNA. Evidence for a marked effect on DNA integrity after preincubation with 1 mM methyl methanesulfonate was provided by a fluorometric technique with ethidium bromide. There was only a small or insignificant enhancement of this effect when hepatoma cells were incubated with nicotinamide. At concentrations of 2-20 mM, 3-aminobenzamide was observed to cause greater effects than nicotinamide on DNA synthesis and integrity and on cellular proliferation in HTC cells. Comparison of the effects of nicotinamide and 3-aminobenzamide with those of N'-methylnicotinamide suggested that some of the effects on DNA synthesis may not be mediated through inhibition of poly(ADP-ribose) synthetase. Inhibition of HTC cell proliferation was observed at a concentration of 3-aminobenzamide, 2 mM, which has been reported to be nontoxic for other cell types.

Animals↗
PubMed1983

Increased fraction of acid-soluble proteins in 0.35 M NaCl extracts of nuclei from rat liver tumors.

1. The fraction of proteins extracted from nuclei with 0.35 M NaCl and soluble in 2% trichloroacetic acid was examined in five Morris hepatomas and rat liver. 2. This fraction was a much greater percentage of the total 0.35 M NaCl soluble proteins in the tumors than in normal or regenerating liver. 3. In part, this difference was due to proteins with molecular weights greater than high mobility group proteins. 4. The conditions for precipitation of high mobility group proteins 1 and 2 with trichloroacetic acid were found to differ in hepatoma and liver fractions.

Animals↗