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PMID:10063 | Thermothrix thioparus gen. et sp. nov. a facultatively anaerobic facultative chemolithotroph living at neutral pH and high temperature. | Thermothrix thioparus gen. et ep. nov. occurs naturally in a New Mexico hot spring at a temperature of 74 degrees C, a pH of 7.0, and a HS- concentration of 1 mg/litre. The organism is gram-negative, non-motile, 0.5-1.0 X 3-20 mum, and forms cell chains up to 1 cm in length. The resulting filaments do not possess a sheath. Sulfur is deposited extracellularly. The organism was isolated using an autotrophic medium with HS- as the energy source and NO3- as the terminal electron acceptor. Anaerobically either NO2- or NO3- is required, NO2- is formed from NO3-, and no observable gas is evolved. Oxygen can also be used as the terminal electron acceptor, but growth is poor because of the decreased solubility of O2 at temperatures required for growth. Alternate energy sources used aerobically and anaerobically include hexose, HS-, SO3-, and S2O3=. The temperature optimum is 70-73 degrees C and growth occurs from 62 to 77 degrees C. The organism's thermal and physiological characteristics are compared to those of Bacillus stearothermophilus, Methanobacterium thermoautotrophicum, Sulfolobus acidocalderius, Thermus aquaticus, Thermus flavus, as well as Thiobacillus denitrificans, the latter being the only other facultatively anaerobic chemolithotroph which has been isolated and described. | Thermothrix thioparus gen. et sp. nov. a facultatively anaerobic facultative chemolithotroph living at neutral pH and high temperature. Thermothrix thioparus gen. et ep. nov. occurs naturally in a New Mexico hot spring at a temperature of 74 degrees C, a pH of 7.0, and a HS- concentration of 1 mg/litre. The organism is gram-negative, non-motile, 0.5-1.0 X 3-20 mum, and forms cell chains up to 1 cm in length. The resulting filaments do not possess a sheath. Sulfur is deposited extracellularly. The organism was isolated using an autotrophic medium with HS- as the energy source and NO3- as the terminal electron acceptor. Anaerobically either NO2- or NO3- is required, NO2- is formed from NO3-, and no observable gas is evolved. Oxygen can also be used as the terminal electron acceptor, but growth is poor because of the decreased solubility of O2 at temperatures required for growth. Alternate energy sources used aerobically and anaerobically include hexose, HS-, SO3-, and S2O3=. The temperature optimum is 70-73 degrees C and growth occurs from 62 to 77 degrees C. The organism's thermal and physiological characteristics are compared to those of Bacillus stearothermophilus, Methanobacterium thermoautotrophicum, Sulfolobus acidocalderius, Thermus aquaticus, Thermus flavus, as well as Thiobacillus denitrificans, the latter being the only other facultatively anaerobic chemolithotroph which has been isolated and described. | [
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PMID:10064 | Comparison of talc-Celite and polyelectrolyte 60 in virus recovery from sewage: development of technique and experiments with poliovirus (type 1, Sabin)-contaminated multilitre samples. | For virus recovery from sewage, a mixture of talc and Celite was tested as a possible inexpensive substitute for polyelectrolyte 60 (PE 60). After adjustment of pH to 6 and the addition of 45-60 plaque forming units (PFU)/ml of poliovirus type I (Sabin) to the sewage sample under test, 100 ml of it was passed through either a PE 60 (400 mg) or a talc (300 mg)-Celite (100 mg) layer; the layer-adsorbed virus was eluted with 10 ml of 10% fetal calf serum (FCS) in saline (pH 7.2). In these experiments, PE 60 layers recovered 73-80% (mean 76%) of the input virus. In comparison, virus recoveries with the talc-Celite layers were 65-70% (mean 68%). Passage of 5 litres of raw sewage (containing 50 to 1.26 X 10(5) PFU/100 ml of the poliovirus) through the talc (15 g)-Celite (5 g) layers and virus elution with 50 ml of 10% FCS in saline gave virus recoveries of 33-63% (mean 49%). Except for pH adjustment and prefiltration through two layers of gauze to remove large solids, no other sample pretreatment was found to be necessary. Application of this technique to recovery of indigenous viruses from field samples of raw sewage and effluents has been highly satisfactory. | Comparison of talc-Celite and polyelectrolyte 60 in virus recovery from sewage: development of technique and experiments with poliovirus (type 1, Sabin)-contaminated multilitre samples. For virus recovery from sewage, a mixture of talc and Celite was tested as a possible inexpensive substitute for polyelectrolyte 60 (PE 60). After adjustment of pH to 6 and the addition of 45-60 plaque forming units (PFU)/ml of poliovirus type I (Sabin) to the sewage sample under test, 100 ml of it was passed through either a PE 60 (400 mg) or a talc (300 mg)-Celite (100 mg) layer; the layer-adsorbed virus was eluted with 10 ml of 10% fetal calf serum (FCS) in saline (pH 7.2). In these experiments, PE 60 layers recovered 73-80% (mean 76%) of the input virus. In comparison, virus recoveries with the talc-Celite layers were 65-70% (mean 68%). Passage of 5 litres of raw sewage (containing 50 to 1.26 X 10(5) PFU/100 ml of the poliovirus) through the talc (15 g)-Celite (5 g) layers and virus elution with 50 ml of 10% FCS in saline gave virus recoveries of 33-63% (mean 49%). Except for pH adjustment and prefiltration through two layers of gauze to remove large solids, no other sample pretreatment was found to be necessary. Application of this technique to recovery of indigenous viruses from field samples of raw sewage and effluents has been highly satisfactory. | [
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PMID:10065 | The effects of pH and temperature on the assay of superoxide dismutase. | A simple and reliable method for the measurement of superoxide dismutase (EC 1.15.1.1) activity is described. The method is based on a linear inhibition of the reduction of acetylated cytochrome c by superoxide dismutase. | The effects of pH and temperature on the assay of superoxide dismutase. A simple and reliable method for the measurement of superoxide dismutase (EC 1.15.1.1) activity is described. The method is based on a linear inhibition of the reduction of acetylated cytochrome c by superoxide dismutase. | [
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PMID:10066 | Biosynthesis of lysine in Saccharomyces cervisiae: properties and spectrophotometric determination of homocitrate synthase activity. | A rapid assay is described for homocitrate synthase (EC 4.1.3.21) of the lysine biosynthetic pathway of Saccharomyces cerevisiae. The alpha-ketoglutarate-dependent cleavage of acetyl-coA was measured spectrophotometrically as decrease in absorbance at 600 nm in the presence of 2,6-dichlorophenol-indophenol and enzyme from the wild type strain X2180. This activity was also present in citrate synthaseless glutamate auxotroph glu3, and the activity was inhibited by 5 mM L-lysine. Radioactive homocitric acid was obtained from a reaction mixture containing [1-14C]acetyl-coA. Homocitrate synthase activity was dependent upon time, both substrates, and enzyme. The activity exhibited a pH and temperature optimum of 7.5-8.0 and 32 degrees C, respectively, and was inhibited by metal-chelating and sulfhydryl-binding agents. | Biosynthesis of lysine in Saccharomyces cervisiae: properties and spectrophotometric determination of homocitrate synthase activity. A rapid assay is described for homocitrate synthase (EC 4.1.3.21) of the lysine biosynthetic pathway of Saccharomyces cerevisiae. The alpha-ketoglutarate-dependent cleavage of acetyl-coA was measured spectrophotometrically as decrease in absorbance at 600 nm in the presence of 2,6-dichlorophenol-indophenol and enzyme from the wild type strain X2180. This activity was also present in citrate synthaseless glutamate auxotroph glu3, and the activity was inhibited by 5 mM L-lysine. Radioactive homocitric acid was obtained from a reaction mixture containing [1-14C]acetyl-coA. Homocitrate synthase activity was dependent upon time, both substrates, and enzyme. The activity exhibited a pH and temperature optimum of 7.5-8.0 and 32 degrees C, respectively, and was inhibited by metal-chelating and sulfhydryl-binding agents. | [
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PMID:10067 | Dependence of the superficial layers of Spirillum putridiconchylium on Ca2+ or Sr2+. | Chelating agents disrupted the superficial layers on Spirillum putridiconchylium and adsorption of cationized ferritin indicated that both upper and lower surfaces of superficial layer fragments, as well as the outer membrane surface, possessed areas which were negatively charged. Growth of the bacterium in 1% casamino acids (vitamin free) resulted in cells which were devoid of the superficial layers, and negative staining of these cells revealed in amorphous precipitate together with a vesicular outer membrane component extruding from their surfaces into the medium. Addition of either 1 mM Ca2+ or 1 mM Sr2+ to the growth medium produced the typical regularly structured cell surface, whereas addition of equal concentrations of Li+, Na+, K+, Mg2+, Ba2+, Mn2+, Fe3+, or three polyamines produced the structureless surface. | Dependence of the superficial layers of Spirillum putridiconchylium on Ca2+ or Sr2+. Chelating agents disrupted the superficial layers on Spirillum putridiconchylium and adsorption of cationized ferritin indicated that both upper and lower surfaces of superficial layer fragments, as well as the outer membrane surface, possessed areas which were negatively charged. Growth of the bacterium in 1% casamino acids (vitamin free) resulted in cells which were devoid of the superficial layers, and negative staining of these cells revealed in amorphous precipitate together with a vesicular outer membrane component extruding from their surfaces into the medium. Addition of either 1 mM Ca2+ or 1 mM Sr2+ to the growth medium produced the typical regularly structured cell surface, whereas addition of equal concentrations of Li+, Na+, K+, Mg2+, Ba2+, Mn2+, Fe3+, or three polyamines produced the structureless surface. | [
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PMID:10068 | Some properties of p-coumarate decarboxylase from Cladosporium phlei. | The optimal pH and temperature of p-coumarate decarboxylase were 6.0 and 23 degrees C respectively. The enzyme activity was reduced to three quarters by heat treatment at 35 degrees C for 5 min and by half at 25 degrees C in 24 h, but kept almost unchanged at -20 degrees C at least for 10 days. The activity was not inhibited by potassium cyanide, sodium diethyldithiocarbamate, ethylenediaminetetraacetic acid disodium salt, or sodium citrate at 10 mM concentration, but was inhibited by p-chloromercuribenzoate or iodoacetate at 0.1 mM, the inhibition by the former being prevented to a great extent by the presence of reduced glutathione or dithiothreitol. The activity was inhibited by maleic acid cinnamic acid, or p-methoxycinnamic acid, but not by fumaric acid, acrylic acid, p-hydroxystyrene, furcatin p-hydroxyphenylacetic acid, or phloretic acid. An unsubstituted p-hydroxy group on the benzene ring and an acrylic acid side chain were required for the enzyme activity. Km value for trans-p-coumaric acid was about 6.5 X 10(-4) M. | Some properties of p-coumarate decarboxylase from Cladosporium phlei. The optimal pH and temperature of p-coumarate decarboxylase were 6.0 and 23 degrees C respectively. The enzyme activity was reduced to three quarters by heat treatment at 35 degrees C for 5 min and by half at 25 degrees C in 24 h, but kept almost unchanged at -20 degrees C at least for 10 days. The activity was not inhibited by potassium cyanide, sodium diethyldithiocarbamate, ethylenediaminetetraacetic acid disodium salt, or sodium citrate at 10 mM concentration, but was inhibited by p-chloromercuribenzoate or iodoacetate at 0.1 mM, the inhibition by the former being prevented to a great extent by the presence of reduced glutathione or dithiothreitol. The activity was inhibited by maleic acid cinnamic acid, or p-methoxycinnamic acid, but not by fumaric acid, acrylic acid, p-hydroxystyrene, furcatin p-hydroxyphenylacetic acid, or phloretic acid. An unsubstituted p-hydroxy group on the benzene ring and an acrylic acid side chain were required for the enzyme activity. Km value for trans-p-coumaric acid was about 6.5 X 10(-4) M. | [
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PMID:10069 | Production of volatiles from decomposing plant tissues and effect of these volatiles on Rhizoctonia solani in culture. | Volatiles, of which NH3 is a major component, were evolved from decomposing immature corn tissue (c:n9) and affected R. solani in culture two ways: they supplied additional nitrogen to the growth medium so that fungal mycelial growth increased; and they raised substrate pH from 5.5 to 8.2 which induced melanization of mycelium. Volatiles increased fungus growth and pigmentation within 2 weeks of amendment addition to soil. Increases were concomitant with NH3 production from corn tissue. More NH3 evolved from decomposing corn tissues of C:N9 and 17 than from those of C-N 33 and 81. More growth and pigmentation occurred in flasks through which volatiles from decomposing corn (C:N9) were passed than in flasks through which volatiles from nonamended soil or decomposing corn (C:N81) were passed. Carbon dioxide from decomposing tissues did not affect growth or pigmentation. Twice as much NH3 evolved from corn tissue (C:N9) which decomposed in saturated soil than from tissue which decomposed in soil at 50% of its water-holding capacity. Pigment production doubled under saturated conditions. | Production of volatiles from decomposing plant tissues and effect of these volatiles on Rhizoctonia solani in culture. Volatiles, of which NH3 is a major component, were evolved from decomposing immature corn tissue (c:n9) and affected R. solani in culture two ways: they supplied additional nitrogen to the growth medium so that fungal mycelial growth increased; and they raised substrate pH from 5.5 to 8.2 which induced melanization of mycelium. Volatiles increased fungus growth and pigmentation within 2 weeks of amendment addition to soil. Increases were concomitant with NH3 production from corn tissue. More NH3 evolved from decomposing corn tissues of C:N9 and 17 than from those of C-N 33 and 81. More growth and pigmentation occurred in flasks through which volatiles from decomposing corn (C:N9) were passed than in flasks through which volatiles from nonamended soil or decomposing corn (C:N81) were passed. Carbon dioxide from decomposing tissues did not affect growth or pigmentation. Twice as much NH3 evolved from corn tissue (C:N9) which decomposed in saturated soil than from tissue which decomposed in soil at 50% of its water-holding capacity. Pigment production doubled under saturated conditions. | [
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PMID:10070 | Factors affecting rate of methane formation from acetic acid by enriched methanogenic cultures. | A stable enrichment culture converting acetic acid to methane was successfully obtained from a pear waste digester, using a synthetic substrate solution with acetic acid as the main carbon source. This enrichment culture converted up to 10 mmol of acetic acid per litre per day at 35 degrees C and did not use hydrogen or formic acid in appreciable amounts as substrate for methane production instead of, or in addition to, acetic acid. The rate of conversion of acetic acid to methane was maximum at temperature of 40-45 degrees C, at a pH of 6.5 to 7.1, and was adversely affected by exposure to air, reducing agents, and high salt concentrations. The rate of conversion was independent of acetic acid concentration between 0.2 and 100 mM, but dropped markedly at concentrations below 0.2 mM. | Factors affecting rate of methane formation from acetic acid by enriched methanogenic cultures. A stable enrichment culture converting acetic acid to methane was successfully obtained from a pear waste digester, using a synthetic substrate solution with acetic acid as the main carbon source. This enrichment culture converted up to 10 mmol of acetic acid per litre per day at 35 degrees C and did not use hydrogen or formic acid in appreciable amounts as substrate for methane production instead of, or in addition to, acetic acid. The rate of conversion of acetic acid to methane was maximum at temperature of 40-45 degrees C, at a pH of 6.5 to 7.1, and was adversely affected by exposure to air, reducing agents, and high salt concentrations. The rate of conversion was independent of acetic acid concentration between 0.2 and 100 mM, but dropped markedly at concentrations below 0.2 mM. | [
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PMID:10071 | Roles of low pH, carbon and inorganic nitrogen source use in chlamydospore formation by Fusarium solani. | Citrate and malate were poorer sources of exogenous carbon than several hexose, pentose, or disaccharide sugars for supporting macroconidial germination by Fusarium solani at high conidial density (1 X 10(5) condia/ml). Only citrate, however, failed to block chlamydospore morphogenesis to a degree comparable to glucose or other readily used sugars. Mostly immature chlamydospores were formed in the presence of citrate. At low conidial density (5 X 10(3) conidia/ml), exogenous carbon-independent macroconidial germination and subsequent rapid chalmydospore formation on germ tubes was not inhibited by ammonium or nitrate nitrogen. The citrate-phosphate buffered, low pH (4.0) medium of Cochrane induced more immature chlamydospore formation by F. solani than a pH 6.0 medium, but few mature chlamydospores were formed in either medium. Condensation of hyphal cytoplasm into developing chlamydospores, a character typical of chlamydospore formation, did not occur extensively and macroconidia, hyphae, and immature chlamydospores stained deeply with Sudan III, suggesting lipid biosynthesis. This inhibition of chlamydospore maturation may be due partly to nitrogen deficiency imposed by the high C:N ratio of the medium and to the presence of citrate. Only vesiculate hyphal cells were formed by F. solani f. sp. phaseoli in both media. Field soils to which the clone of F. solani used is indigenous had mean pH values ranging from 5.2 to 6.0. | Roles of low pH, carbon and inorganic nitrogen source use in chlamydospore formation by Fusarium solani. Citrate and malate were poorer sources of exogenous carbon than several hexose, pentose, or disaccharide sugars for supporting macroconidial germination by Fusarium solani at high conidial density (1 X 10(5) condia/ml). Only citrate, however, failed to block chlamydospore morphogenesis to a degree comparable to glucose or other readily used sugars. Mostly immature chlamydospores were formed in the presence of citrate. At low conidial density (5 X 10(3) conidia/ml), exogenous carbon-independent macroconidial germination and subsequent rapid chalmydospore formation on germ tubes was not inhibited by ammonium or nitrate nitrogen. The citrate-phosphate buffered, low pH (4.0) medium of Cochrane induced more immature chlamydospore formation by F. solani than a pH 6.0 medium, but few mature chlamydospores were formed in either medium. Condensation of hyphal cytoplasm into developing chlamydospores, a character typical of chlamydospore formation, did not occur extensively and macroconidia, hyphae, and immature chlamydospores stained deeply with Sudan III, suggesting lipid biosynthesis. This inhibition of chlamydospore maturation may be due partly to nitrogen deficiency imposed by the high C:N ratio of the medium and to the presence of citrate. Only vesiculate hyphal cells were formed by F. solani f. sp. phaseoli in both media. Field soils to which the clone of F. solani used is indigenous had mean pH values ranging from 5.2 to 6.0. | [
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PMID:10072 | Transport of phenylalanine by conidia of Fusarium sulphureum. | L-Phenylalanine was actively transported by conidia of Fusarium sulphurenum Schlect (isolate 1). Uptake was optimal at pH 7, 30 degrees C; respiration-dependent; and was unaffected by relatively high internal concentrations of phenylalanine. The Km for transport was 1-3 X 10(-5) M and the Vmax was 2.5-4 nmol/min per milligram dry weight. Phenylalanine is transported by a general transport system for basic and neutral amino acids. Sucrose repressed uptake of phenylalanine and this repression was largely negated by cycloheximide. Efflux of accumulated phenylalanine was influx-dependent; this transport system deteriorated slowly with aging of the conidial culture. | Transport of phenylalanine by conidia of Fusarium sulphureum. L-Phenylalanine was actively transported by conidia of Fusarium sulphurenum Schlect (isolate 1). Uptake was optimal at pH 7, 30 degrees C; respiration-dependent; and was unaffected by relatively high internal concentrations of phenylalanine. The Km for transport was 1-3 X 10(-5) M and the Vmax was 2.5-4 nmol/min per milligram dry weight. Phenylalanine is transported by a general transport system for basic and neutral amino acids. Sucrose repressed uptake of phenylalanine and this repression was largely negated by cycloheximide. Efflux of accumulated phenylalanine was influx-dependent; this transport system deteriorated slowly with aging of the conidial culture. | [
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PMID:10074 | Characterization of a strain of Methanospirillum hungatti. | The results of morphological, base ratio, nutritional, temperature, and pH studies on a strain of Methanospirillum hungatii, isolated from an anaerobic pear waste digester, are described. The isolate, designated as strain GP 1, was compared with some of the characteristics of type-strain M. hungatii JF 1. Strain GP 1 is Gram-negative, weakly motile, and a strict anaerobe with a guanine plus cytosine (G +C) content of 46.5 mol%. The preferred substrates for methane production are hydrogen, carbon dioxide, and formate. Acetate is used under certain conditions but its specific contribution to cell carbon and (or) methane formation was not established. The optimum temperature for both growth and methane production is 35 degrees C, but growth and methane production occur over the range 25-45 degrees C. Methane production is optimal at pH 7.0. | Characterization of a strain of Methanospirillum hungatti. The results of morphological, base ratio, nutritional, temperature, and pH studies on a strain of Methanospirillum hungatii, isolated from an anaerobic pear waste digester, are described. The isolate, designated as strain GP 1, was compared with some of the characteristics of type-strain M. hungatii JF 1. Strain GP 1 is Gram-negative, weakly motile, and a strict anaerobe with a guanine plus cytosine (G +C) content of 46.5 mol%. The preferred substrates for methane production are hydrogen, carbon dioxide, and formate. Acetate is used under certain conditions but its specific contribution to cell carbon and (or) methane formation was not established. The optimum temperature for both growth and methane production is 35 degrees C, but growth and methane production occur over the range 25-45 degrees C. Methane production is optimal at pH 7.0. | [
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PMID:10075 | Psychotropic drug use among women. | The consistent 2:1 ratio of women to men in the receipt of prescriptions for psychotropic drugs is reflected in the higher rates for women of neurotic illness, symptoms of both physical and mental discomfort, and help-seeking and drug-taking behaviour. Physicians' perceptions of the problems presented by their male and female patients influence their prescribing of these drugs. Recent statistics in Ontario indicate that greater use of physicians' services by women is an inadequate explanation of the higher rate of prescribing of psychotropic drugs to women. A longitudinal study of a large insured population in Ontario showed that almost twice the proportion of females, compared with males, received a prescription for psychotropic drugs in 1970-71 and in 1973-74, a higher proportion of females received multiple prescriptions for each drug class, and males were more likely than females to have received only one prescription in a year. | Psychotropic drug use among women. The consistent 2:1 ratio of women to men in the receipt of prescriptions for psychotropic drugs is reflected in the higher rates for women of neurotic illness, symptoms of both physical and mental discomfort, and help-seeking and drug-taking behaviour. Physicians' perceptions of the problems presented by their male and female patients influence their prescribing of these drugs. Recent statistics in Ontario indicate that greater use of physicians' services by women is an inadequate explanation of the higher rate of prescribing of psychotropic drugs to women. A longitudinal study of a large insured population in Ontario showed that almost twice the proportion of females, compared with males, received a prescription for psychotropic drugs in 1970-71 and in 1973-74, a higher proportion of females received multiple prescriptions for each drug class, and males were more likely than females to have received only one prescription in a year. | [
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PMID:10076 | Adoptive immunotherapy of a Gross virus producing lymphoma and a methylcholanthrene-induced fibrosarcoma in tolerant rats. | Immunological tolerance to Gross virus-specific transplantation antigens in rats given neonatae transfer of donor lymphoid cells beneath the kidney capsule of syngeneic recipient rats. Immune or normal donor cells invariably developed a cell-mediated immune reaction in kidneys of GV-tolerant recipients, presumably against GV antigens present on the surface of recipient lymphoid cells in the kidney. Spleen and lymph node cells from tolerant rats failed to develop a reaction in tolerant recipients, but developed a strong reaction to histoincompatible antigens in the kidneys of semisyngeneic tolerant rats. The immunologically tolerant state in the rats could be broken by adoptive transfer of spleen and lymph node cells from syngeneic rats immunized with GV-induced lymphoma cells. Immunotherapy of a GV-induced and also a GV-infected methylcholanthrene-induced fibrosarcoma growing in tolerant rats was successful when immune spleen and lymph node cells were administered i.p. 3 days after s.c. inoculation of 2 X 10(7) tumor cells in the case of the lymphoma, and 1 day after inoculation of 5 X 10(6) tumor cells in the case of the fibrosarcoma. | Adoptive immunotherapy of a Gross virus producing lymphoma and a methylcholanthrene-induced fibrosarcoma in tolerant rats. Immunological tolerance to Gross virus-specific transplantation antigens in rats given neonatae transfer of donor lymphoid cells beneath the kidney capsule of syngeneic recipient rats. Immune or normal donor cells invariably developed a cell-mediated immune reaction in kidneys of GV-tolerant recipients, presumably against GV antigens present on the surface of recipient lymphoid cells in the kidney. Spleen and lymph node cells from tolerant rats failed to develop a reaction in tolerant recipients, but developed a strong reaction to histoincompatible antigens in the kidneys of semisyngeneic tolerant rats. The immunologically tolerant state in the rats could be broken by adoptive transfer of spleen and lymph node cells from syngeneic rats immunized with GV-induced lymphoma cells. Immunotherapy of a GV-induced and also a GV-infected methylcholanthrene-induced fibrosarcoma growing in tolerant rats was successful when immune spleen and lymph node cells were administered i.p. 3 days after s.c. inoculation of 2 X 10(7) tumor cells in the case of the lymphoma, and 1 day after inoculation of 5 X 10(6) tumor cells in the case of the fibrosarcoma. | [
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PMID:10077 | Formation in isolated rat liver microsomes and nuclei of benzo(a)pyrene metabolites that bind to DNA. | The hepatic nuclear fraction isolated from 3-methylcholanthrene (MC)-treated rats contained enhanced levels of cytochrome P-450 and aryl hydrocarbon hydroxylase [benzo(a)pyrene (BP) monooxygenase], whereas the activities of epoxide hydrase and reduced nicotinamide adenine dinucleotide phosphate-cytochrome c reductase and the concentration of cytochrome b5 were not altered. The metabolite pattern of BP was investigated by using high-pressure liquid chromatography and was found to be similar in nuclei and microsomes from MC-treated rats. After incubation of the nuclear fraction with [3H]BP and reduced nicotinamide adenine dinculeotide phosphate, radioactivity was found to be associated with nuclear DNA and the extent of binding was markedly enhanced by pretreatment of the animals with MC. Binding was strongly inhibited by a-napthoflavone but was not influenced by 1,1,1-trichloropropene-2,3-oxide, an inhibitor of epoxide hydrase. In the presence of microsomes from MC-treated rats, increased binding of BP to DNA was observed in nuclei from both control and MC-treated rats; moreover, when the nuclear DNA was replaced by a corresponding amount of calf thymus DNA, the extent of binding was severalfold enhanced. In contrast to nuclei from control rats, the nuclear fraction from MC-treated rats showed an increase in bound radioactivity when incubated with a microsome-free supernatant, obtained by incubating microsomes from MC-treated rats with [3H]BP. The increase in extent of binding was eliminated in the presence of menadione or alpha-naphthoflavone. It is suggested that under the conditions used here the following different processes may have contributed to the total incorporation of BP products into nuclear DNA: (a) formation of DNA-binding products derived from BP by nuclear aryl hydrocarbon hydroxylase; (b) formation of DNA-binding products from microsomal BP metabolites by nuclear aryl hydrocarbon hydroxylase; and (c) direct transfer of reactive microsomal metabolites to nuclear DNA. | Formation in isolated rat liver microsomes and nuclei of benzo(a)pyrene metabolites that bind to DNA. The hepatic nuclear fraction isolated from 3-methylcholanthrene (MC)-treated rats contained enhanced levels of cytochrome P-450 and aryl hydrocarbon hydroxylase [benzo(a)pyrene (BP) monooxygenase], whereas the activities of epoxide hydrase and reduced nicotinamide adenine dinucleotide phosphate-cytochrome c reductase and the concentration of cytochrome b5 were not altered. The metabolite pattern of BP was investigated by using high-pressure liquid chromatography and was found to be similar in nuclei and microsomes from MC-treated rats. After incubation of the nuclear fraction with [3H]BP and reduced nicotinamide adenine dinculeotide phosphate, radioactivity was found to be associated with nuclear DNA and the extent of binding was markedly enhanced by pretreatment of the animals with MC. Binding was strongly inhibited by a-napthoflavone but was not influenced by 1,1,1-trichloropropene-2,3-oxide, an inhibitor of epoxide hydrase. In the presence of microsomes from MC-treated rats, increased binding of BP to DNA was observed in nuclei from both control and MC-treated rats; moreover, when the nuclear DNA was replaced by a corresponding amount of calf thymus DNA, the extent of binding was severalfold enhanced. In contrast to nuclei from control rats, the nuclear fraction from MC-treated rats showed an increase in bound radioactivity when incubated with a microsome-free supernatant, obtained by incubating microsomes from MC-treated rats with [3H]BP. The increase in extent of binding was eliminated in the presence of menadione or alpha-naphthoflavone. It is suggested that under the conditions used here the following different processes may have contributed to the total incorporation of BP products into nuclear DNA: (a) formation of DNA-binding products derived from BP by nuclear aryl hydrocarbon hydroxylase; (b) formation of DNA-binding products from microsomal BP metabolites by nuclear aryl hydrocarbon hydroxylase; and (c) direct transfer of reactive microsomal metabolites to nuclear DNA. | [
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PMID:10078 | Nonenzymatic reduction of benzo(a)pyrene diol-epoxides to trihydroxypentahydrobenzo(a)pyrenes by reduced nicotinamide adenine dinucleotide phosphate. | The diol-epoxide r-7,t-8-dihydroxy-t-9,10-oxy-7,8,9,10-tetrahydrobenzo(a)pyrene is a potent mutagen and possibly the ultimate carcinogenic form of benzo(a)pyrene. A (7/8,9)-trihydroxy-7,8,9,10,10-pentahydrobenzo(a)pyrene is formed from the diol-epoxide r-7,t-8-dihydroxy-t-9,10-oxy-7,8,9,10-tetrahydroxybenzo(a)pyrene by reduction with reduced nicotinamide adenine dinucleotide phosphate. Its formation is linear with reduced nicotinamide adenine dinucleotide phosphate concentration and does not require the presence of enzyme. A (7,9/8)-trihydroxy-7,8,9,10,10-pentahydrobenzo(a)pyrene is similarly formed from the diol-epoxide r-7,t-8-dihydroxy-c-9,10-oxy-7,8,9,10-tetrahydrobenzo(a)pyrene by reduction with reduced nicotinamide adenine dinucleotide phosphate. The structures of the trihydroxypentahydrobenzo(a)pyrenes were established by their ultraviolet absorption and mass spectra and their reaction with potassium triacetylosmate. | Nonenzymatic reduction of benzo(a)pyrene diol-epoxides to trihydroxypentahydrobenzo(a)pyrenes by reduced nicotinamide adenine dinucleotide phosphate. The diol-epoxide r-7,t-8-dihydroxy-t-9,10-oxy-7,8,9,10-tetrahydrobenzo(a)pyrene is a potent mutagen and possibly the ultimate carcinogenic form of benzo(a)pyrene. A (7/8,9)-trihydroxy-7,8,9,10,10-pentahydrobenzo(a)pyrene is formed from the diol-epoxide r-7,t-8-dihydroxy-t-9,10-oxy-7,8,9,10-tetrahydroxybenzo(a)pyrene by reduction with reduced nicotinamide adenine dinucleotide phosphate. Its formation is linear with reduced nicotinamide adenine dinucleotide phosphate concentration and does not require the presence of enzyme. A (7,9/8)-trihydroxy-7,8,9,10,10-pentahydrobenzo(a)pyrene is similarly formed from the diol-epoxide r-7,t-8-dihydroxy-c-9,10-oxy-7,8,9,10-tetrahydrobenzo(a)pyrene by reduction with reduced nicotinamide adenine dinucleotide phosphate. The structures of the trihydroxypentahydrobenzo(a)pyrenes were established by their ultraviolet absorption and mass spectra and their reaction with potassium triacetylosmate. | [
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PMID:10079 | Establishment and characterization of human neuroblastoma cell lines. | Three new tissue culture cell lines, CHP-100, CHP-126, and CHP-134, have been established from explant cultures of human neuroblastoma. The cell lines have been characterized with respect to morphology, chromosomes constitution, growth, neural enzyme content, and their ability to grow in nude mice. The cells grow as dense masses comprised of fibroblast-or neuroblast-like cells with small processes. The cell lines differ in their neural enzyme acitivity. The chromosomal content of the 3 cell lines is near diploid, and all are capable of forming tumors in nude mice. The morphological findings indicate that the cells in culture resemble those found in the tumor, and the enzyme activities are consistent with those of nervous tissue. This the morphological, biochemical, and tumorigenic properties confirm that the 3 cell lines are neoplastic cells of neural origin. | Establishment and characterization of human neuroblastoma cell lines. Three new tissue culture cell lines, CHP-100, CHP-126, and CHP-134, have been established from explant cultures of human neuroblastoma. The cell lines have been characterized with respect to morphology, chromosomes constitution, growth, neural enzyme content, and their ability to grow in nude mice. The cells grow as dense masses comprised of fibroblast-or neuroblast-like cells with small processes. The cell lines differ in their neural enzyme acitivity. The chromosomal content of the 3 cell lines is near diploid, and all are capable of forming tumors in nude mice. The morphological findings indicate that the cells in culture resemble those found in the tumor, and the enzyme activities are consistent with those of nervous tissue. This the morphological, biochemical, and tumorigenic properties confirm that the 3 cell lines are neoplastic cells of neural origin. | [
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] |
PMID:10080 | Behavior of transaldolase (EC 2.2.1.2) and transketolase (EC 2.2.1.1) Activities in normal, neoplastic, differentiating, and regenerating liver. | The objective of this investigation was to throw light on the biological behavior and metabolic regulation of hepatic enzymes of the nonoxidative branch of the pentose phosphate pathway. The activities of transaldolase (EC 2.2.1.2) and trasketolase (EC 2.2.1.1) Were compared in biological conditions that involve modulation of gene expression such as in starvation, in differentiation, after partial hepatectomy, and in a spectrum of hepatomas of different growth rates. The enzyme activities were determined under optimal kinetic conditions by spectrophotometric methods in the 100,000 X g supernatant fluids prepared from tissue homogenates. The kinetic properties of transaldolase and transketolase were similar in normal liver and in rapidly growing hepatoma 3924A. For transaldolase, apparent Km values of 0.13 mM (normal liver) and 0.17 mM (hepatoma) were observed for erythrose 4-phosphate and of 0.30 to 0.35 mM for fructose 6-phosphate. The pH optima in liver and hepatoma were at approximately 6.9 to 7.2. For the transketolase substrates, ribose 5-phosphate and xylulose 5-phosphate, the apparent Km values were 0.3 and 0.5 mM, respectively, in both liver and hepatoma. A broad pH optimum around 7.6 was observed in both tissues. In organ distribution studies, enzyme activities were measured in liver, intestinal mucosa, thymus, kidney, spleen, brain, adipose tissue, lung, heart, and skeletal muscle. Taking the specific activity of liver as 100%, transaldolase activity was the highest in intestinal mucosa (316%) and in thymus (219%); it was the lowest in heart (53%) and in skeletal muscle (21%). Transketolase activity was highest in kidney (155%) and lowest in heart (26%) and skeletal muscle (23%). Starvation decreased transaldolase and transketolase activities in 6 days to 69 and 74%, respectively, of those of the liver of the normal, fed rat. This was in the same range as the decrease in the protein concentration (66%y. In the liver tumors, transaldolase activity was increased 1.5- to 3.4-fold over the activities observed in normal control rat liver. Transketolase activity showed no relationship to tumor proliferation rate. In the regenerating liver at 24 hr after partial hepatectomy, the activity of both pentose phosphate pathway enzymes was in the same range as that of the sham-operated controls. In differentiation at the postnatal age of 5, 12, 23, and 32 days, hepatic transaldolase activities were 33, 44, 55, and 72%, respectively, of the activities observed in the 60-day-old, adult male rat. During the same period, transketolase activ-ties were 18, 21, 26, and 55% of the activities observed in liver of adult rat. The demonstration of increased transaldolase activity in hepatomas, irrespective of the degree of tumor malignancy, differentiation, or growth rate, suggests that the reprogramming of gene expression in malignant transformation is linked with an increase in the expression of this pentose phosphate pathway enzyme... | Behavior of transaldolase (EC 2.2.1.2) and transketolase (EC 2.2.1.1) Activities in normal, neoplastic, differentiating, and regenerating liver. The objective of this investigation was to throw light on the biological behavior and metabolic regulation of hepatic enzymes of the nonoxidative branch of the pentose phosphate pathway. The activities of transaldolase (EC 2.2.1.2) and trasketolase (EC 2.2.1.1) Were compared in biological conditions that involve modulation of gene expression such as in starvation, in differentiation, after partial hepatectomy, and in a spectrum of hepatomas of different growth rates. The enzyme activities were determined under optimal kinetic conditions by spectrophotometric methods in the 100,000 X g supernatant fluids prepared from tissue homogenates. The kinetic properties of transaldolase and transketolase were similar in normal liver and in rapidly growing hepatoma 3924A. For transaldolase, apparent Km values of 0.13 mM (normal liver) and 0.17 mM (hepatoma) were observed for erythrose 4-phosphate and of 0.30 to 0.35 mM for fructose 6-phosphate. The pH optima in liver and hepatoma were at approximately 6.9 to 7.2. For the transketolase substrates, ribose 5-phosphate and xylulose 5-phosphate, the apparent Km values were 0.3 and 0.5 mM, respectively, in both liver and hepatoma. A broad pH optimum around 7.6 was observed in both tissues. In organ distribution studies, enzyme activities were measured in liver, intestinal mucosa, thymus, kidney, spleen, brain, adipose tissue, lung, heart, and skeletal muscle. Taking the specific activity of liver as 100%, transaldolase activity was the highest in intestinal mucosa (316%) and in thymus (219%); it was the lowest in heart (53%) and in skeletal muscle (21%). Transketolase activity was highest in kidney (155%) and lowest in heart (26%) and skeletal muscle (23%). Starvation decreased transaldolase and transketolase activities in 6 days to 69 and 74%, respectively, of those of the liver of the normal, fed rat. This was in the same range as the decrease in the protein concentration (66%y. In the liver tumors, transaldolase activity was increased 1.5- to 3.4-fold over the activities observed in normal control rat liver. Transketolase activity showed no relationship to tumor proliferation rate. In the regenerating liver at 24 hr after partial hepatectomy, the activity of both pentose phosphate pathway enzymes was in the same range as that of the sham-operated controls. In differentiation at the postnatal age of 5, 12, 23, and 32 days, hepatic transaldolase activities were 33, 44, 55, and 72%, respectively, of the activities observed in the 60-day-old, adult male rat. During the same period, transketolase activ-ties were 18, 21, 26, and 55% of the activities observed in liver of adult rat. The demonstration of increased transaldolase activity in hepatomas, irrespective of the degree of tumor malignancy, differentiation, or growth rate, suggests that the reprogramming of gene expression in malignant transformation is linked with an increase in the expression of this pentose phosphate pathway enzyme... | [
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PMID:10081 | L-Asparagine synthetase in serum as a marker for neoplasia. | L-Asparagine synthetase appears in serum approximately 7 days after the s.c. implantation of 1 X 10(5) cells of Leukemia 5178Y/AR (resistant to L-asparaginase) and increases in activity as the neoplasm grows and metastasizes. The principal source of the enzyme is the primary tumor. After intravranial inoculation of tumor, the rate of leakage of the enzyme is more pronounced than when the subcutaneous, intramuscular, or intraperitoneal routes are used. 1-(2-Chloroethyl)-3-cyclohexyl-1-nitrosourea (NSC 79037), a nitro-sourea effective in the palliation of L5178Y/AR, temporarily halts the influx of enzyme into the blood stream, as does surgical excision of the s.c. tumor nodules. Treatment of mice with L-asparaginase within 24 hr of inoculation of the tumor markedly augments both tumor growth and the rate of penetration of L-asparagine synthetase into the circulation. Several other L-asparagine synthetase into the circulation. Several other L-asparaginase-resistant tumors also were found to spill L-asparagine synthetase into the serum, but the correlation between this phenomenon and the specific activity of the enzyme in homogenates of the tumor was imperfect. | L-Asparagine synthetase in serum as a marker for neoplasia. L-Asparagine synthetase appears in serum approximately 7 days after the s.c. implantation of 1 X 10(5) cells of Leukemia 5178Y/AR (resistant to L-asparaginase) and increases in activity as the neoplasm grows and metastasizes. The principal source of the enzyme is the primary tumor. After intravranial inoculation of tumor, the rate of leakage of the enzyme is more pronounced than when the subcutaneous, intramuscular, or intraperitoneal routes are used. 1-(2-Chloroethyl)-3-cyclohexyl-1-nitrosourea (NSC 79037), a nitro-sourea effective in the palliation of L5178Y/AR, temporarily halts the influx of enzyme into the blood stream, as does surgical excision of the s.c. tumor nodules. Treatment of mice with L-asparaginase within 24 hr of inoculation of the tumor markedly augments both tumor growth and the rate of penetration of L-asparagine synthetase into the circulation. Several other L-asparagine synthetase into the circulation. Several other L-asparaginase-resistant tumors also were found to spill L-asparagine synthetase into the serum, but the correlation between this phenomenon and the specific activity of the enzyme in homogenates of the tumor was imperfect. | [
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PMID:10082 | The isolation of lectins on acid-treated agarose. | The ability of several D-galactose and N-acetyl-D-galactosamine-binding lectins to bind to Sepharose was investigated. Lectins from soybean, Wistaria floribunda, Bauhinia purpurea alba, and Sophora japonica could be isolated by affinity chromatography on acid-treated Sepharose 6B. These lectins would not bind to untreated Sepharose 2B, 4B, and 6B. The binding of B. purpurea alba and S. japonica was temperature-dependent. The S. japonica lectin would bind only at high pH. Ricinus communis toxin also showed a temperature-dependence of binding; acid-treated Sepharose 6B was a better affinity support for the toxin than was untreated Sepharose 4B Lectins from lima bean, Dolichos biflorus, and kidney-bean phytohemagglutinin did not bind to Sepharose under any of the conditions studied. | The isolation of lectins on acid-treated agarose. The ability of several D-galactose and N-acetyl-D-galactosamine-binding lectins to bind to Sepharose was investigated. Lectins from soybean, Wistaria floribunda, Bauhinia purpurea alba, and Sophora japonica could be isolated by affinity chromatography on acid-treated Sepharose 6B. These lectins would not bind to untreated Sepharose 2B, 4B, and 6B. The binding of B. purpurea alba and S. japonica was temperature-dependent. The S. japonica lectin would bind only at high pH. Ricinus communis toxin also showed a temperature-dependence of binding; acid-treated Sepharose 6B was a better affinity support for the toxin than was untreated Sepharose 4B Lectins from lima bean, Dolichos biflorus, and kidney-bean phytohemagglutinin did not bind to Sepharose under any of the conditions studied. | [
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PMID:10085 | Identification of a ribonuclease P-like activity from human KB cells. | An endoribonuclease which cleaves tRNA precursor molecules has been partially purified from human KB tissue culture cells. This activity is found in cytoplasmic fractions but is not detectable in the nucleoplasm. tRNA precursor molecules from both E. coli and KB cells are cleaved by this novel activity to produce 5' phosphate-terminated oligonucleotides. E coli RNAase P and the KB cell nuclease both make a single endonucleolytic scission in E. coli tRNATyr precursor, thereby separating the 41 extra nucleotides on the 5' end of the precursor molecule from the 5' terminal sequence of the mature tRNATyr molecule. The cleavage products generated from other E. coli tRNA precursors by the KB cell activity are identical in size to those produced by RNAase P. The KB cell endoribonuclease requires Mg2+ and a monovalent cation (Na+, K+, or NH4+) for function. The enzymatic activity has a broad pH optimum, centered near pH 8.0, and the activity is inhibited by tRNA. Several KB cell RNAs with long half-lives in vivo, including 5S and bulk 4S RNA, are not cleaved by this nuclease. The KB cell endoribonuclease resembles E. coli RNAase P in its substrate specificity, pH optimum, ion requirements, and sensitivity to tRNA. These properties and the cytoplasmic localization of the novel endoribonuclease indicate its involvement in the biosynthesis of KB cell tRNA. | Identification of a ribonuclease P-like activity from human KB cells. An endoribonuclease which cleaves tRNA precursor molecules has been partially purified from human KB tissue culture cells. This activity is found in cytoplasmic fractions but is not detectable in the nucleoplasm. tRNA precursor molecules from both E. coli and KB cells are cleaved by this novel activity to produce 5' phosphate-terminated oligonucleotides. E coli RNAase P and the KB cell nuclease both make a single endonucleolytic scission in E. coli tRNATyr precursor, thereby separating the 41 extra nucleotides on the 5' end of the precursor molecule from the 5' terminal sequence of the mature tRNATyr molecule. The cleavage products generated from other E. coli tRNA precursors by the KB cell activity are identical in size to those produced by RNAase P. The KB cell endoribonuclease requires Mg2+ and a monovalent cation (Na+, K+, or NH4+) for function. The enzymatic activity has a broad pH optimum, centered near pH 8.0, and the activity is inhibited by tRNA. Several KB cell RNAs with long half-lives in vivo, including 5S and bulk 4S RNA, are not cleaved by this nuclease. The KB cell endoribonuclease resembles E. coli RNAase P in its substrate specificity, pH optimum, ion requirements, and sensitivity to tRNA. These properties and the cytoplasmic localization of the novel endoribonuclease indicate its involvement in the biosynthesis of KB cell tRNA. | [
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PMID:10090 | Changes in extracellular potassium activity in response to decreased pH in rabbit atrial muscle. | The extracellular and intracellular potassium (K+) activities of isolated superfused rabbit atrial muscle were measured using K+-sensitive liquid ion exchanger microelectrodes. When the pH of the bathing medium was decreased from 7.5 to 6.8, intracellular K+ activity fell and extracellular K+ activity rose from a mean control level of 3.6 mM to a new steady state level of 3.9 mM after 1 hour. When the pH was further decreased to 6.1, extracellular K+ activity increased to a mean of 4.9 mM. Following the change in pH, the increase in extracellular K+ activity occurred over a period of 30-40 minutes at which time a stable value was reached and maintained for the next hour. On return to normal pH the extracellular K+ activity returned to control with a time constant of 20 minutes or less. Measurements of intracellular K+ activity over 1 hour showed a mean loss of 3 mM at pH 6.8 and a mean loss of 8 mM at pH 6.1. The loss was reversible within 20 minutes of return to control pH. The increase in extracellular K+ activity was accompanied by a decrease in resting membrane potential as well as decreases in maximum dv/dt and overshoot of the action potential. The action potential contour underwent complex changes consisting of decrease in the plateau and a prolongation of the time to full repolarization. | Changes in extracellular potassium activity in response to decreased pH in rabbit atrial muscle. The extracellular and intracellular potassium (K+) activities of isolated superfused rabbit atrial muscle were measured using K+-sensitive liquid ion exchanger microelectrodes. When the pH of the bathing medium was decreased from 7.5 to 6.8, intracellular K+ activity fell and extracellular K+ activity rose from a mean control level of 3.6 mM to a new steady state level of 3.9 mM after 1 hour. When the pH was further decreased to 6.1, extracellular K+ activity increased to a mean of 4.9 mM. Following the change in pH, the increase in extracellular K+ activity occurred over a period of 30-40 minutes at which time a stable value was reached and maintained for the next hour. On return to normal pH the extracellular K+ activity returned to control with a time constant of 20 minutes or less. Measurements of intracellular K+ activity over 1 hour showed a mean loss of 3 mM at pH 6.8 and a mean loss of 8 mM at pH 6.1. The loss was reversible within 20 minutes of return to control pH. The increase in extracellular K+ activity was accompanied by a decrease in resting membrane potential as well as decreases in maximum dv/dt and overshoot of the action potential. The action potential contour underwent complex changes consisting of decrease in the plateau and a prolongation of the time to full repolarization. | [
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PMID:10091 | The use of gamma-glutamyl transpeptidase in differentiating liver from bone isoenzymes of alkaline phosphatase. | Sixty-one patients with elevated alkaline phosphatase activity due to liver or bone diseases were studied. An attempt was made to identify the origin of the increased alkaline phosphatase by chemical inhibition, by inactivation by heat and urea, and by electrophoretic separation. The results obtained from these procedures were correlated with the gamma-glutamyl transpeptidase activities performed on each patient. We concluded from this study that gamma-glutamyl transpeptidase determination, together with alkaline phosphatase electrophoretic separations, are useful laboratory procedures for accurately identifying the origin of elevated alkaline phosphatase activity. | The use of gamma-glutamyl transpeptidase in differentiating liver from bone isoenzymes of alkaline phosphatase. Sixty-one patients with elevated alkaline phosphatase activity due to liver or bone diseases were studied. An attempt was made to identify the origin of the increased alkaline phosphatase by chemical inhibition, by inactivation by heat and urea, and by electrophoretic separation. The results obtained from these procedures were correlated with the gamma-glutamyl transpeptidase activities performed on each patient. We concluded from this study that gamma-glutamyl transpeptidase determination, together with alkaline phosphatase electrophoretic separations, are useful laboratory procedures for accurately identifying the origin of elevated alkaline phosphatase activity. | [
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PMID:10092 | Normal limits of urinary excretion of eleven enzymes. | Urinary excretion of lactate dehydrogenase, hydroxybutyrate dehydrogenase, gamma-glutamyltransferase, alkaline phosphatase, arylsulphatase A, alpha-glucosidase, beta-galactosidase, trehalase, N-acetyl-beta-glucosaminidase, beta-glucuronidase, and leucinearylamidase was studies in a carefully selected group of 100 healthy subjects, 50 women and 50 men. Enzyme activities were assayed in 3-h morning samples after gel filtration of the urine. Activities were related to time volume, and to urinary creatinine concentration. Several transforming functions had to be applied to enzyme output data to obtain an approximation to gaussian frequency distribution. Men showed a significantly higher excretion of gamma-glutamyltransferase, alpha-glucosidase, trehalase, N-acetyl-beta-glucosaminidase,beta-glucuronidase, and leucine arylamidase activity than did women if enzyme activity was related to urinary time volume. Women excreted more lactate dehydrogenase, hydroxybutyrate dehydrogenase, gamma-glutamyltransferase, alkaline phosphatase, alpha-glucosidase, trehalase, and N-acetyl-beta-glucosaminidase activity than did men, if urinary creatinine was used as the basis of reference. Reference intervals were calculated as 2.5 and 97.5 percentiles for both sexes. | Normal limits of urinary excretion of eleven enzymes. Urinary excretion of lactate dehydrogenase, hydroxybutyrate dehydrogenase, gamma-glutamyltransferase, alkaline phosphatase, arylsulphatase A, alpha-glucosidase, beta-galactosidase, trehalase, N-acetyl-beta-glucosaminidase, beta-glucuronidase, and leucinearylamidase was studies in a carefully selected group of 100 healthy subjects, 50 women and 50 men. Enzyme activities were assayed in 3-h morning samples after gel filtration of the urine. Activities were related to time volume, and to urinary creatinine concentration. Several transforming functions had to be applied to enzyme output data to obtain an approximation to gaussian frequency distribution. Men showed a significantly higher excretion of gamma-glutamyltransferase, alpha-glucosidase, trehalase, N-acetyl-beta-glucosaminidase,beta-glucuronidase, and leucine arylamidase activity than did women if enzyme activity was related to urinary time volume. Women excreted more lactate dehydrogenase, hydroxybutyrate dehydrogenase, gamma-glutamyltransferase, alkaline phosphatase, alpha-glucosidase, trehalase, and N-acetyl-beta-glucosaminidase activity than did men, if urinary creatinine was used as the basis of reference. Reference intervals were calculated as 2.5 and 97.5 percentiles for both sexes. | [
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PMID:10093 | Isolation and identification of benzodiazepine drugs and their metabolites in urine by use of Amberlite XAD-2 resin and thin-layer chromatography. | We used the method described here to detect and identify seven benzodiazepine derivatives--diazepam, chlorodiazepoxide, nitrazepam, cloxazolam, oxazolam, oxazepam, and medazepam--and their metabolites in the urine of rabbits given the seven drugs orally. We column-chromatographed 25-ml samples of urine on Amberlite XAD-2. The drugs and their metabolites in the urine were adsorbed by the resin, irrespective of urinary pH, and upon successive elution with methanol and ethyl acetate/methanol/acetic acid (90/10/0.1 by vol) they could be separated and extracted from the normal components of urine with satisfactory analytical recovery. The conjugated metabolites were then enzymatically hydrolyzed and the hydrolysate was extracted into ethyl acetate and the extract thin-layer chromatographed to detect and identify each drug and each of its metabolites. In an experiment in which the urine of human subjects given 5 mg of nitrazepam orally was analyzed by this method, the metabolites of nitrazepam in the 24-h urine could be identified satisfactorily. | Isolation and identification of benzodiazepine drugs and their metabolites in urine by use of Amberlite XAD-2 resin and thin-layer chromatography. We used the method described here to detect and identify seven benzodiazepine derivatives--diazepam, chlorodiazepoxide, nitrazepam, cloxazolam, oxazolam, oxazepam, and medazepam--and their metabolites in the urine of rabbits given the seven drugs orally. We column-chromatographed 25-ml samples of urine on Amberlite XAD-2. The drugs and their metabolites in the urine were adsorbed by the resin, irrespective of urinary pH, and upon successive elution with methanol and ethyl acetate/methanol/acetic acid (90/10/0.1 by vol) they could be separated and extracted from the normal components of urine with satisfactory analytical recovery. The conjugated metabolites were then enzymatically hydrolyzed and the hydrolysate was extracted into ethyl acetate and the extract thin-layer chromatographed to detect and identify each drug and each of its metabolites. In an experiment in which the urine of human subjects given 5 mg of nitrazepam orally was analyzed by this method, the metabolites of nitrazepam in the 24-h urine could be identified satisfactorily. | [
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PMID:10094 | Factors contributing to intra-individual variation of serum constituents: Physiological day-to-day variation in concentrations of 10 specific proteins in sera of healthy subjects. | Using an automated immunoprecipitin method, we assayed human sera for 10 proteins: haptoglobin, orosomucoid, transferrin, alpha1 antitrypsin, alpha2-macroglobulin, IgG, IGa, IgM, complement C3, and complement C4. Blood from 14 healthy subjects (25-40y) was sampled on six separate days. From each venipuncture serum was divided into four eliquots; two were assayed on the day of venipuncture and two were frozen and kept until the end of the study, when all of the frozen samples were analyzed in one batch. With this experimental design, batch-to-batch analytical variation could be estimated, and we avoided confounding it with the biological variation. Data analysis was based on the analysis of variance technique. The average physiological intra-individual coefficient of variation ranged from 2.5% for transferrin to 11.1% for orosomucoid. THe interindividual variation ranged from 9.5% for transferrin to 70.5% for haptoglobin and the ratio between intra-individual variation and interindividual variation ranged from 0.66 for IgM to 0.26 for orosomucoid and transferrin. | Factors contributing to intra-individual variation of serum constituents: Physiological day-to-day variation in concentrations of 10 specific proteins in sera of healthy subjects. Using an automated immunoprecipitin method, we assayed human sera for 10 proteins: haptoglobin, orosomucoid, transferrin, alpha1 antitrypsin, alpha2-macroglobulin, IgG, IGa, IgM, complement C3, and complement C4. Blood from 14 healthy subjects (25-40y) was sampled on six separate days. From each venipuncture serum was divided into four eliquots; two were assayed on the day of venipuncture and two were frozen and kept until the end of the study, when all of the frozen samples were analyzed in one batch. With this experimental design, batch-to-batch analytical variation could be estimated, and we avoided confounding it with the biological variation. Data analysis was based on the analysis of variance technique. The average physiological intra-individual coefficient of variation ranged from 2.5% for transferrin to 11.1% for orosomucoid. THe interindividual variation ranged from 9.5% for transferrin to 70.5% for haptoglobin and the ratio between intra-individual variation and interindividual variation ranged from 0.66 for IgM to 0.26 for orosomucoid and transferrin. | [
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PMID:10095 | Reaction of alkaline sodium picrate with creatinine: I. Kinetics and mechanism of formation of the mono-creatinine picric acid complex. | Spectrophotometric, kinetic, and nuclear magnetic resonance studies indicate that alkaline sodium picrate and creatinine react to form a 1/1 aduct between picric and creatinine, with a stability constant of log K= 4.26. Kinetic studies indicate that the forward reaction is first order with respect to picric acid, hydroxide, and creatinine concentration. The reverse reaction, the dissociation of the 1/1 complex, shows a complex dependence on hydroxide concentration. The expression for the observed pseudo-first-order rate constant in the presence of excess picric acid is: Kobsd = K1K0[P][OH] +[K2[OH]x. A value of K1K0 = 5.0 (mol/liter)-2s-1 is obtained. For accurate analytical results with this reaction, hydroxide concentration must be maintained at a constant value for both samples and standards. | Reaction of alkaline sodium picrate with creatinine: I. Kinetics and mechanism of formation of the mono-creatinine picric acid complex. Spectrophotometric, kinetic, and nuclear magnetic resonance studies indicate that alkaline sodium picrate and creatinine react to form a 1/1 aduct between picric and creatinine, with a stability constant of log K= 4.26. Kinetic studies indicate that the forward reaction is first order with respect to picric acid, hydroxide, and creatinine concentration. The reverse reaction, the dissociation of the 1/1 complex, shows a complex dependence on hydroxide concentration. The expression for the observed pseudo-first-order rate constant in the presence of excess picric acid is: Kobsd = K1K0[P][OH] +[K2[OH]x. A value of K1K0 = 5.0 (mol/liter)-2s-1 is obtained. For accurate analytical results with this reaction, hydroxide concentration must be maintained at a constant value for both samples and standards. | [
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PMID:10098 | Long-term reproducibility of a new pH/blood-gas quality-control system compared to two other procedures. | The long-term precision and stability of a new quality-control system for blood pH and gas measurements are compared to that of tonometered bicarbonate solutions and serum-based preparations. The new system, consisting of gas-equilibrated bicarbonate solutions in glass ampuls, is shown to be as stable as the serum-based preparation, and as reproducible as either of the other methods. The new system, offering three discrete sets of control values, has certain advantages in the simultaneous quality of pH, carbon dioxide tension, and oxygen tension measurements. | Long-term reproducibility of a new pH/blood-gas quality-control system compared to two other procedures. The long-term precision and stability of a new quality-control system for blood pH and gas measurements are compared to that of tonometered bicarbonate solutions and serum-based preparations. The new system, consisting of gas-equilibrated bicarbonate solutions in glass ampuls, is shown to be as stable as the serum-based preparation, and as reproducible as either of the other methods. The new system, offering three discrete sets of control values, has certain advantages in the simultaneous quality of pH, carbon dioxide tension, and oxygen tension measurements. | [
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PMID:10099 | Application of a vidicon spectrometer for simultaneous multicomponent drug determinations. | A rapid scanning vidicon spectrometer has been evaluated for the simultaneous determination of drugs in mixtures, without a separation step. Spectral data in the ultraviolet region are collected on-line with a small computer at repetition rates of 100 scans per second. Absorbance data at several wavelengths are processed by matrix equations to resolve them into the concentration of each component in each mixture. Results are reported for two-component mixtures of procainamide and N-acetylprocainamide in serum, for two-component aqueous mixtures of butabarbital and seconbarbital, and for seven-component aqueous mixtures of phenobarbital, diphenylhydantoin, aminophylline, acetaminophen, salicylamide, phenylbutazone, and secobarbital. We conclude that the computer-inter-faced vidicon spectrometer is a viable tool for simultaneous multicomponent determinations. | Application of a vidicon spectrometer for simultaneous multicomponent drug determinations. A rapid scanning vidicon spectrometer has been evaluated for the simultaneous determination of drugs in mixtures, without a separation step. Spectral data in the ultraviolet region are collected on-line with a small computer at repetition rates of 100 scans per second. Absorbance data at several wavelengths are processed by matrix equations to resolve them into the concentration of each component in each mixture. Results are reported for two-component mixtures of procainamide and N-acetylprocainamide in serum, for two-component aqueous mixtures of butabarbital and seconbarbital, and for seven-component aqueous mixtures of phenobarbital, diphenylhydantoin, aminophylline, acetaminophen, salicylamide, phenylbutazone, and secobarbital. We conclude that the computer-inter-faced vidicon spectrometer is a viable tool for simultaneous multicomponent determinations. | [
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PMID:10100 | A search for the best buffer to use in assaying human lactate dehydrogenase with the lactate-to-pyruvate reaction. | Highly purified human lactate dehydrogenases I and V were assayed in 17 different buffers, at a variety of reaction pH's. Diethanolamine and 2-amino-2-methyl-1,3-propanediol provided the best measurements of the enzyme, assayed lactate-to-pyruvate. However, the commercial preparation of 2-amino-2-methyl-1,3-propanediol contained insoluble matter and was relatively expensive. All of the four buffers nowmost commonly used were found to present difficulties. Glycine and pyrophosphate were inhibotory tolactate dehydrogenase activity with increasing buffer concentration. 2-Amino-2-methyl-1-propanol had three major disadvantages: it is chemically unstable during reagent preparation; activity is dependent on buffer concentration; and the pH optima for isoenzymes I and V are vastly different. The pKa of tris(hydroxymethyl)aminomethane is 8.0 at 30 degrees C, whereas to measure total activity the reaction pH should be greater than 8.5; thus tris(hydroxymethyl)aminomethane has limited buffering capacity at the reaction pH. | A search for the best buffer to use in assaying human lactate dehydrogenase with the lactate-to-pyruvate reaction. Highly purified human lactate dehydrogenases I and V were assayed in 17 different buffers, at a variety of reaction pH's. Diethanolamine and 2-amino-2-methyl-1,3-propanediol provided the best measurements of the enzyme, assayed lactate-to-pyruvate. However, the commercial preparation of 2-amino-2-methyl-1,3-propanediol contained insoluble matter and was relatively expensive. All of the four buffers nowmost commonly used were found to present difficulties. Glycine and pyrophosphate were inhibotory tolactate dehydrogenase activity with increasing buffer concentration. 2-Amino-2-methyl-1-propanol had three major disadvantages: it is chemically unstable during reagent preparation; activity is dependent on buffer concentration; and the pH optima for isoenzymes I and V are vastly different. The pKa of tris(hydroxymethyl)aminomethane is 8.0 at 30 degrees C, whereas to measure total activity the reaction pH should be greater than 8.5; thus tris(hydroxymethyl)aminomethane has limited buffering capacity at the reaction pH. | [
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PMID:10101 | Negative anion gap in a young adult with multiple myeloma. | A young patient with multiple myeloma was found to have a negative anion gap, with marked asymptomatic hyponatremia. The cause for his negative anion gap is thought to be the myeloma protein, which acts as a cation at physiological pH. Such a hyponatremia responds to reduction in serum concentration of paraprotein and should not be treated by sodium replacement. | Negative anion gap in a young adult with multiple myeloma. A young patient with multiple myeloma was found to have a negative anion gap, with marked asymptomatic hyponatremia. The cause for his negative anion gap is thought to be the myeloma protein, which acts as a cation at physiological pH. Such a hyponatremia responds to reduction in serum concentration of paraprotein and should not be treated by sodium replacement. | [
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PMID:10102 | Characterization of adenosine deaminase isozymes from normal human erythrocytes. | Adenosine deaminase of phenotype ADA was partially purified by chromatography on CM-Sephadex C-50 and ammonium sulphate precipitation. With DEAE-Sephadex A-50 three isozymes could be detected. a. The KM values for the substrate adenosine were found to be 30 muM for each isozyme. b. pH optimum was 7.0 and the molecular weight estimated by gel filtration was found to be 30 000 for each isozyme. c. The heat stability of RBC-ADA type 1-1 was greater than type 1-2. The isozyme in type 2-1 representing the electrophoretic band of phenotype ADA2-2 is the most labile. d. ATP, ADP, AMP and cyclic AMP, PCMB and 6-methylmercaptopurine riboside were found to be competitive inhibitors with ADA in all three isozymes. | Characterization of adenosine deaminase isozymes from normal human erythrocytes. Adenosine deaminase of phenotype ADA was partially purified by chromatography on CM-Sephadex C-50 and ammonium sulphate precipitation. With DEAE-Sephadex A-50 three isozymes could be detected. a. The KM values for the substrate adenosine were found to be 30 muM for each isozyme. b. pH optimum was 7.0 and the molecular weight estimated by gel filtration was found to be 30 000 for each isozyme. c. The heat stability of RBC-ADA type 1-1 was greater than type 1-2. The isozyme in type 2-1 representing the electrophoretic band of phenotype ADA2-2 is the most labile. d. ATP, ADP, AMP and cyclic AMP, PCMB and 6-methylmercaptopurine riboside were found to be competitive inhibitors with ADA in all three isozymes. | [
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PMID:10103 | An abnormal form of purine nucleoside phosphorylase in a family with a child with severe defective T-cell-and normal B-cell immunity. | 1. Purine nucleoside phosphorylase and adenosine deaminase (ADA) were studied in normal red blood cells and lymphocytes and in the cells of a family with a child with a defective T-cell-and normal B-cell immunity. 2. In the propositus no purine nucleoside phosphorylase (NP) activity could be detected in her red cells and lymphocytes, while the ADA activity was somewhat increased. The NP activities of the father, mother and brother of the propositus are in the heterozygote range. The decreased activity of NP was not only found for the substrate inosine but also when guanosine or xanthosine were used as substrate. The mode of inheritance is autosomal recessive. 3. With starch gel electrophoresis no NP activity could be detected in the patient's haemolysate. The electrophoretic patterns of NP from the father, mother and brother of the patient seem to be the same as for normal NP with six bands of NP activity. 4. The nucleoside phosphorylases of the father, mother and brother of the patient were characterized by an increased KM for the substrate inosine, normal pH optimum and a decreased heat stability. | An abnormal form of purine nucleoside phosphorylase in a family with a child with severe defective T-cell-and normal B-cell immunity. 1. Purine nucleoside phosphorylase and adenosine deaminase (ADA) were studied in normal red blood cells and lymphocytes and in the cells of a family with a child with a defective T-cell-and normal B-cell immunity. 2. In the propositus no purine nucleoside phosphorylase (NP) activity could be detected in her red cells and lymphocytes, while the ADA activity was somewhat increased. The NP activities of the father, mother and brother of the propositus are in the heterozygote range. The decreased activity of NP was not only found for the substrate inosine but also when guanosine or xanthosine were used as substrate. The mode of inheritance is autosomal recessive. 3. With starch gel electrophoresis no NP activity could be detected in the patient's haemolysate. The electrophoretic patterns of NP from the father, mother and brother of the patient seem to be the same as for normal NP with six bands of NP activity. 4. The nucleoside phosphorylases of the father, mother and brother of the patient were characterized by an increased KM for the substrate inosine, normal pH optimum and a decreased heat stability. | [
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] |
PMID:10104 | [Sialyltransferase in human malignant melanoma]. | A glycosyltransferase, CMP-N-acetylneuraminic acid : glycoprotein sialyltransferase was found in human malignant melanoma. Activities were measured with desialized glycoprotein as an exogenous acceptor. The enzyme was characterized by means of its pH optimum, 5.5, temperature optimum, 30 degrees C, KM values, 10 muM for the sugar nucleotide and 0.3 mM for desialized glycoprotein. It did not require exogenously added metal ions but was slightly stimulated by Mg2+. It required detergent for optimal activity. The effect of nucleotides and sugar nucleotides on enzyme activity has been investigated. | [Sialyltransferase in human malignant melanoma]. A glycosyltransferase, CMP-N-acetylneuraminic acid : glycoprotein sialyltransferase was found in human malignant melanoma. Activities were measured with desialized glycoprotein as an exogenous acceptor. The enzyme was characterized by means of its pH optimum, 5.5, temperature optimum, 30 degrees C, KM values, 10 muM for the sugar nucleotide and 0.3 mM for desialized glycoprotein. It did not require exogenously added metal ions but was slightly stimulated by Mg2+. It required detergent for optimal activity. The effect of nucleotides and sugar nucleotides on enzyme activity has been investigated. | [
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PMID:10105 | Gamma-Glutamyl-3-carboxy-14-nitroanilide: the substrate of choice for routine determinations of gamma-glutamyl-transferase activity in serum? | Gamma-Glutamyl-3-carboxy-4-nitroanilide has been tested as donor substrate in the assay of gamma-glutamyltransferase activity in serum, glycylglycine being used as acceptor substrate. This donor substrate is highly solube even in neutral solutions, in contrast to the commonly used gamma-glutamyl-4-nitroanilide. The enzyme which apparently acts accordingly to a ping-pong bi bi kinetic mechanism, shows an absolute KM value for gamma-glutamyl-3-carboxy-4-nitroanilide of about 0.64 mmol/l, and for glycylglycine of about 13.4 mmol/l. The former KM value is significantly lower than that previously found for gamma-glutamyl-4-nitroanilide. The carboxyl derivative exhibits a marked competitive inhibitory effect on the gamma-glutamyltransferase. This effect is more pronounced than that of gamma-glutamyl-4-nitroanilide. The carboxyl derivative has somewhat higher absorbance in the range of wave length (400-420 nm) used to monitor the formation of the product. It is concluded that as donor substrate in the assay of gamma-glutamyltransferase activity of serum, the new derivative is not substantially superior to the gamma-glutamyl-4-nitroanilide conventionally used. | Gamma-Glutamyl-3-carboxy-14-nitroanilide: the substrate of choice for routine determinations of gamma-glutamyl-transferase activity in serum? Gamma-Glutamyl-3-carboxy-4-nitroanilide has been tested as donor substrate in the assay of gamma-glutamyltransferase activity in serum, glycylglycine being used as acceptor substrate. This donor substrate is highly solube even in neutral solutions, in contrast to the commonly used gamma-glutamyl-4-nitroanilide. The enzyme which apparently acts accordingly to a ping-pong bi bi kinetic mechanism, shows an absolute KM value for gamma-glutamyl-3-carboxy-4-nitroanilide of about 0.64 mmol/l, and for glycylglycine of about 13.4 mmol/l. The former KM value is significantly lower than that previously found for gamma-glutamyl-4-nitroanilide. The carboxyl derivative exhibits a marked competitive inhibitory effect on the gamma-glutamyltransferase. This effect is more pronounced than that of gamma-glutamyl-4-nitroanilide. The carboxyl derivative has somewhat higher absorbance in the range of wave length (400-420 nm) used to monitor the formation of the product. It is concluded that as donor substrate in the assay of gamma-glutamyltransferase activity of serum, the new derivative is not substantially superior to the gamma-glutamyl-4-nitroanilide conventionally used. | [
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PMID:10106 | Serum gamma glutamyl transferase and alkaline phosphatase activities in epileptics receiving anticonvulsant therapy. | 1. Serum gamma glutamyl transferase (gammaGT) and alkaline phosphatase (ALP) activities have been estimated in 49 epileptic patients taking anticonvulsant drugs. 2. Serum gammaGT activity was clearly elevated in 12 of the patients and borderline in 8, giving a 40% frequency of abnormal values. 3. Total serum ALP activity was elevated in 7 out of 33 adult patients and 2 of 16 juveniles. 4. Electrophoresis on polyacrylamide gel showed that the bone isoenzyme was responsible for most of the increased serum ALP activity and that even when the total was not elevated, the contribution of the bone isoenzyme was greater than normal. 5. There was no correlation between total serum ALP and gammaGT activities but elevations of serum gammaGT were often accompanied by an increase in the proportion of the bone enzyme in the total serum ALP activity. | Serum gamma glutamyl transferase and alkaline phosphatase activities in epileptics receiving anticonvulsant therapy. 1. Serum gamma glutamyl transferase (gammaGT) and alkaline phosphatase (ALP) activities have been estimated in 49 epileptic patients taking anticonvulsant drugs. 2. Serum gammaGT activity was clearly elevated in 12 of the patients and borderline in 8, giving a 40% frequency of abnormal values. 3. Total serum ALP activity was elevated in 7 out of 33 adult patients and 2 of 16 juveniles. 4. Electrophoresis on polyacrylamide gel showed that the bone isoenzyme was responsible for most of the increased serum ALP activity and that even when the total was not elevated, the contribution of the bone isoenzyme was greater than normal. 5. There was no correlation between total serum ALP and gammaGT activities but elevations of serum gammaGT were often accompanied by an increase in the proportion of the bone enzyme in the total serum ALP activity. | [
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PMID:10108 | Determination of isoenzyme contents of lactic dehydrogenase activity and 2-hydroxybutyric dehydrogenase activity in lactic dehydrogenase preparations. | In this inestigation, a determination of the isoenzyme contents of LDH and HBD activities in lactate dehydrogenase preparations and the differences in the interaction of these preparations with NAD analogues were examined. The results obtained were as follows. 1. The activity ratio between oxidation and reduction in LDH reaction is shown to be similar in both H4 and M4 preparations, whereas for HBD activity, the ratio seems to be lower in the M4 preparation than in H4(H4/M = 1/2). 2. NAD and its analogues (NXD, TNAD, and TNXD) are shown to be useful coenzymes for the LDH reaction, while 3-acetyl derivatives appear to be unsuitable for this purpose because of their lower activity. HBD activity with 3-acetyl NXD is shown to be higher than with TNAD or TNXD. among these NAD analogues, 3-acetyl NXD gives the highest HBD activity, especially in the M4 preparation. 3. The LDH activity of H4 relative to M4 preparations has been shown to be maximal when 450 mM lactic acid with NAD or 15 mM lactic acid with TNXD are used. Under these conditions, the contents of LDH subunits can be estimated with considerable reliability. As to HBD activity, the content of LDH subunit having HBD activity has been estimated by determing the enzyme activity under conditions in which either 300 mM 2-hydroxybutyrate with 3-acetyl NXD or 15 mM 2-hydroxybutyrate with NAD are employed. | Determination of isoenzyme contents of lactic dehydrogenase activity and 2-hydroxybutyric dehydrogenase activity in lactic dehydrogenase preparations. In this inestigation, a determination of the isoenzyme contents of LDH and HBD activities in lactate dehydrogenase preparations and the differences in the interaction of these preparations with NAD analogues were examined. The results obtained were as follows. 1. The activity ratio between oxidation and reduction in LDH reaction is shown to be similar in both H4 and M4 preparations, whereas for HBD activity, the ratio seems to be lower in the M4 preparation than in H4(H4/M = 1/2). 2. NAD and its analogues (NXD, TNAD, and TNXD) are shown to be useful coenzymes for the LDH reaction, while 3-acetyl derivatives appear to be unsuitable for this purpose because of their lower activity. HBD activity with 3-acetyl NXD is shown to be higher than with TNAD or TNXD. among these NAD analogues, 3-acetyl NXD gives the highest HBD activity, especially in the M4 preparation. 3. The LDH activity of H4 relative to M4 preparations has been shown to be maximal when 450 mM lactic acid with NAD or 15 mM lactic acid with TNXD are used. Under these conditions, the contents of LDH subunits can be estimated with considerable reliability. As to HBD activity, the content of LDH subunit having HBD activity has been estimated by determing the enzyme activity under conditions in which either 300 mM 2-hydroxybutyrate with 3-acetyl NXD or 15 mM 2-hydroxybutyrate with NAD are employed. | [
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PMID:10109 | beta-Glucuronidase and oestrogens in hydatidiform mole. | The activity of beta-glucuronidase (beta-D-glucuronide glycuronohydrolase, EC 3.2.1.31) in placental and hydatidiform mole tissue and in sera from non-pregnant, normal and molar pregnant women was determined. The oestrogen concentrations in the two solid tissues were also assayed. Significant differences were found in the activities of the enzyme between placental and molar tissues and among the various sera. The lower activity in molar serum corresponds to a lower concentration of oestrogens in molar tissues and may be regarded as a response to the alleviation of conjugation requirements. | beta-Glucuronidase and oestrogens in hydatidiform mole. The activity of beta-glucuronidase (beta-D-glucuronide glycuronohydrolase, EC 3.2.1.31) in placental and hydatidiform mole tissue and in sera from non-pregnant, normal and molar pregnant women was determined. The oestrogen concentrations in the two solid tissues were also assayed. Significant differences were found in the activities of the enzyme between placental and molar tissues and among the various sera. The lower activity in molar serum corresponds to a lower concentration of oestrogens in molar tissues and may be regarded as a response to the alleviation of conjugation requirements. | [
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PMID:10110 | Serum gamma-glutamyl transpeptidase and physical exercise. | The immediate and delayed influence of exercise of variable type, duration and intensity on the serum gamma-glutamyl transpeptidase activity (gamma-GT) has been examined in athletes and untrained persons. The possible effects of exercise-induced changes in other parameters (plasma free glutamate, serum triglyceride, haemoconcentration) on the measured postexercise serum gamma-GT have been discussed, partly on the basis of our own experimental data. It appears that neither exercise itself, nor any one of the above mentioned factors (excepting a slight, transient effect of haemoconcentration after brief, intensive exertion) have noticeable influence on serum gamma-GT. Determinations of this enzyme for diagnostic purposes are therefore likely to be used without regard to acute and chronic exercise history in man. | Serum gamma-glutamyl transpeptidase and physical exercise. The immediate and delayed influence of exercise of variable type, duration and intensity on the serum gamma-glutamyl transpeptidase activity (gamma-GT) has been examined in athletes and untrained persons. The possible effects of exercise-induced changes in other parameters (plasma free glutamate, serum triglyceride, haemoconcentration) on the measured postexercise serum gamma-GT have been discussed, partly on the basis of our own experimental data. It appears that neither exercise itself, nor any one of the above mentioned factors (excepting a slight, transient effect of haemoconcentration after brief, intensive exertion) have noticeable influence on serum gamma-GT. Determinations of this enzyme for diagnostic purposes are therefore likely to be used without regard to acute and chronic exercise history in man. | [
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PMID:10112 | A simple, rapid method for prenatal detection of defects in propionate metabolism. | Incorporation of radiolabel from propionate-1(-14)C into protein (TCA insoluble material) in fibroblasts or amniotic fluid cells, provides a rapid, simple means of detecting fetuses with inborn errors of propionate metabolism using small numbers of cells. Controls were easily differentiated from mutant lines over differing media pH conditions. This method was successfully used to diagnose correctly a normal fetus at risk for methylmalonic acidemia. This method can be used as an adjunct in diagnosis, but cannot replace direct enzyme analysis. | A simple, rapid method for prenatal detection of defects in propionate metabolism. Incorporation of radiolabel from propionate-1(-14)C into protein (TCA insoluble material) in fibroblasts or amniotic fluid cells, provides a rapid, simple means of detecting fetuses with inborn errors of propionate metabolism using small numbers of cells. Controls were easily differentiated from mutant lines over differing media pH conditions. This method was successfully used to diagnose correctly a normal fetus at risk for methylmalonic acidemia. This method can be used as an adjunct in diagnosis, but cannot replace direct enzyme analysis. | [
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PMID:10113 | Effects of morphine on brain histamine, antinociception and activity in mice. | 1. The effect of morphine on the histamine content of the mouse brain has been investigated. The changes in the brain histamine level have been related to morphine-induced analgesia and morphine-induced changes in locomotor activity. 2. With doses of morphine between 1 and 5 mg/kg there was a significant increase in histamine levels. The time required to produce a maximal rise in the brain histamine level with 5 mg/kg of morphine was 15 min. 3. There was a significant decrease in brain histamine levels with doses of morphine between 7-5 and 100 mg/kg. The time at which the greatest decrease was produced with 50 mg/kg was 30 min. 4. The time couse of the alteration of brain histamine by morphine did not correlate with its antinociceptive activity. Both the 5 and 50 mg/kg doses of morphine produced analgesia in mice whereas brain histamine levels were increased and decreased, respectively. 5. Pretreating mice with compounds which modify histaminergic function did not modify the antinociceptive action of morphine. 6. Morphine produced a biphasic effect on locomotor activity when the dose was increased from 0-5 through to 100 mg/kg. Doses up to 2-5 mg/kg caused a reduction of activity and doses above this produced significant increases. 7. There appears to be an inverse relationship between the morphine-induced changes of brain histamine and the morphine-induced changes in locomotor activity. | Effects of morphine on brain histamine, antinociception and activity in mice. 1. The effect of morphine on the histamine content of the mouse brain has been investigated. The changes in the brain histamine level have been related to morphine-induced analgesia and morphine-induced changes in locomotor activity. 2. With doses of morphine between 1 and 5 mg/kg there was a significant increase in histamine levels. The time required to produce a maximal rise in the brain histamine level with 5 mg/kg of morphine was 15 min. 3. There was a significant decrease in brain histamine levels with doses of morphine between 7-5 and 100 mg/kg. The time at which the greatest decrease was produced with 50 mg/kg was 30 min. 4. The time couse of the alteration of brain histamine by morphine did not correlate with its antinociceptive activity. Both the 5 and 50 mg/kg doses of morphine produced analgesia in mice whereas brain histamine levels were increased and decreased, respectively. 5. Pretreating mice with compounds which modify histaminergic function did not modify the antinociceptive action of morphine. 6. Morphine produced a biphasic effect on locomotor activity when the dose was increased from 0-5 through to 100 mg/kg. Doses up to 2-5 mg/kg caused a reduction of activity and doses above this produced significant increases. 7. There appears to be an inverse relationship between the morphine-induced changes of brain histamine and the morphine-induced changes in locomotor activity. | [
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PMID:10115 | The influence of histamine on epithelial cell proliferation in the jejunum of the rat. | 1. The influence of histamine and histamine receptor blockade on the mitotic rate in epithelial cell proliferation in epithelial cells lining the crypts of Lieberkühn in rat jejunum was studied. 2. Histamine injection resulted in an increase in the mitotic rate. This increase in mitotic rate was blocked by metiamide but not by mepyramine. 3. Prevention of histamine synthesis by alpha-methylhistidine administration did not alter the mitotic rate. 4. The mechanism by which histamine may influence crypt cell proliferation and possible role of cyclic GMP in this mediation are discussed.¿ | The influence of histamine on epithelial cell proliferation in the jejunum of the rat. 1. The influence of histamine and histamine receptor blockade on the mitotic rate in epithelial cell proliferation in epithelial cells lining the crypts of Lieberkühn in rat jejunum was studied. 2. Histamine injection resulted in an increase in the mitotic rate. This increase in mitotic rate was blocked by metiamide but not by mepyramine. 3. Prevention of histamine synthesis by alpha-methylhistidine administration did not alter the mitotic rate. 4. The mechanism by which histamine may influence crypt cell proliferation and possible role of cyclic GMP in this mediation are discussed.¿ | [
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PMID:10116 | Comparison of the bromureide sedative-hypnotic drugs, bromvaletone (bromisoval) and carbromal, and their chloro analogues in mice. | 1. The central depressant effects of bromvaletone, carbromal and six non-bromo analogues were compared in mice. 2. The chloro analogues of bromvaletone and carbromal were slightly less potent as central depressant agents than the bromo compounds. 3. The chloro analogue of bromvaletone had the greatest margin between central depressant and lethal doses. 4. Lipophilicity (octanol-water partition coefficient) did not provide a unifying relationship for potency within this group of eight acylureas. However, within each of the two subsets of compounds, a linear relationship was found between relative potency and lipophilicity. | Comparison of the bromureide sedative-hypnotic drugs, bromvaletone (bromisoval) and carbromal, and their chloro analogues in mice. 1. The central depressant effects of bromvaletone, carbromal and six non-bromo analogues were compared in mice. 2. The chloro analogues of bromvaletone and carbromal were slightly less potent as central depressant agents than the bromo compounds. 3. The chloro analogue of bromvaletone had the greatest margin between central depressant and lethal doses. 4. Lipophilicity (octanol-water partition coefficient) did not provide a unifying relationship for potency within this group of eight acylureas. However, within each of the two subsets of compounds, a linear relationship was found between relative potency and lipophilicity. | [
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PMID:10117 | Saturable metabolic pathways for ethotoin in man. | 1. The urinary excretion pattern of ethotoin and five metabolites were examined in three patients receiving continuous treatment with ethotoin at two dose levels, in order to investigate the mechanism behind the dose-dependent kinetics of this anticonvulsant drug. 2. The results suggest a partial saturation in the dealkylation process at high dose levels in three patients. 3. A rough approximation of the Michaelis-Menten constants for different enzymatic processes was attempted. On the basis of the results obtained, the p-hydroxylation may be a saturable process. 4. The dose-dependent kinetics of ethotoin in man seem to be explicable by the existence of partly saturable enzymatic pathways. | Saturable metabolic pathways for ethotoin in man. 1. The urinary excretion pattern of ethotoin and five metabolites were examined in three patients receiving continuous treatment with ethotoin at two dose levels, in order to investigate the mechanism behind the dose-dependent kinetics of this anticonvulsant drug. 2. The results suggest a partial saturation in the dealkylation process at high dose levels in three patients. 3. A rough approximation of the Michaelis-Menten constants for different enzymatic processes was attempted. On the basis of the results obtained, the p-hydroxylation may be a saturable process. 4. The dose-dependent kinetics of ethotoin in man seem to be explicable by the existence of partly saturable enzymatic pathways. | [
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PMID:10114 | Adrenergic factors influencing the mitotic rate in stratified squamous epithelium of the buccal mucosa of the rat. | 1. An in vivo stathmokinetic technique was used to determine the immediate effect of various adrenoceptor agonists and antagonists on the mitotic rate in the stratified squamous epithelium of the rat buccal mucosa. 2. The mitotic rate increased significantly in rats treated with propranolol and with practolol, whereas the mitotic rate decreased significantly in rats treated with metaraminol. In animals treated with isoprenaline and phentolamine the mitotic rate did not differ significantly from the control value. 3. The nature of the involvement of adrenergic mechanisms with cell proliferation is still uncertain but both alpha- and beta-adrenergic mechanisms appear to be associated with the control of cell proliferation. | Adrenergic factors influencing the mitotic rate in stratified squamous epithelium of the buccal mucosa of the rat. 1. An in vivo stathmokinetic technique was used to determine the immediate effect of various adrenoceptor agonists and antagonists on the mitotic rate in the stratified squamous epithelium of the rat buccal mucosa. 2. The mitotic rate increased significantly in rats treated with propranolol and with practolol, whereas the mitotic rate decreased significantly in rats treated with metaraminol. In animals treated with isoprenaline and phentolamine the mitotic rate did not differ significantly from the control value. 3. The nature of the involvement of adrenergic mechanisms with cell proliferation is still uncertain but both alpha- and beta-adrenergic mechanisms appear to be associated with the control of cell proliferation. | [
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PMID:10121 | Absorption and malabsorption of folates. | Folic acid is one of the 'younger' vitamins, yet it has attracted intensive study in the thirty years since the identification of pteroylglutamic acid and its polyglutamyl conjugates. The absorption and malabsorption of folates, natural, purified and synthetic, in disease has been studied more than any other vitamin and indeed folate absorption has become one clinical test of intestinal function. We know little about the release of folate from protein complexes, but we have learned, with the help of synthetic radiolabelled pteroylpolyglutamates that polyglutamyl folates are hydrolysed at or near the luminal border of the intestine and the released folate is efficiently absorbed. The rate limiting stage of folate absorption appears to be the transport of the monoglutamyl folate. In disease, and with drugs, folate malabsorption occurs primarily when monoglutamyl transport is depressed. The specific components of the folate transport system, listed in Table 4, are receiving increased attention. The mechanism of uptake is still a topic of controversy but a dual system including both a saturable and a diffusion component would explain most of the data. Reduction and methyl or formyl addition occur in the intestine but such metabolism is not obligatory for transport. The nature of folate binding within the cell and the function of specific folate binding proteins requires further study. At present we have little or no information about the mechanism of folate release from the epithelial cell to the circulation but this step also could influence the rate and specificity of overall process. The tools are now at hand to complete our understanding of the steps in folate absorption and metabolism. Such an understanding should facilitate the management of folate deficiency whenever it complicates gastrointestinal disease or drug therapy. | Absorption and malabsorption of folates. Folic acid is one of the 'younger' vitamins, yet it has attracted intensive study in the thirty years since the identification of pteroylglutamic acid and its polyglutamyl conjugates. The absorption and malabsorption of folates, natural, purified and synthetic, in disease has been studied more than any other vitamin and indeed folate absorption has become one clinical test of intestinal function. We know little about the release of folate from protein complexes, but we have learned, with the help of synthetic radiolabelled pteroylpolyglutamates that polyglutamyl folates are hydrolysed at or near the luminal border of the intestine and the released folate is efficiently absorbed. The rate limiting stage of folate absorption appears to be the transport of the monoglutamyl folate. In disease, and with drugs, folate malabsorption occurs primarily when monoglutamyl transport is depressed. The specific components of the folate transport system, listed in Table 4, are receiving increased attention. The mechanism of uptake is still a topic of controversy but a dual system including both a saturable and a diffusion component would explain most of the data. Reduction and methyl or formyl addition occur in the intestine but such metabolism is not obligatory for transport. The nature of folate binding within the cell and the function of specific folate binding proteins requires further study. At present we have little or no information about the mechanism of folate release from the epithelial cell to the circulation but this step also could influence the rate and specificity of overall process. The tools are now at hand to complete our understanding of the steps in folate absorption and metabolism. Such an understanding should facilitate the management of folate deficiency whenever it complicates gastrointestinal disease or drug therapy. | [
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PMID:10122 | Vitamin B12--folate interrelations. | Megaloblastic anaemia is due to a derangement of DNA synthesis caused by insufficient supply of one or other of the four deoxyribonucleoside triphosphate (dNTP) precursors of DNA synthesis or by direct inhibition of one or other DNA polymerase. Reduced supply of the pyrimidine deoxythymidine triphosphate (dTTP) may be caused by folate or vitamin B12 deficiencies or by the action of dihydrofolate reductase inhibitors (e.g. methotrexate, pyrimethamine or trimethoprim), all of which cause reduced supply of the coenzyme 5, 10 methylene tetrahydrofolate (pentaglutamate) needed for thymidylate synthetase. Reduced dTTP supply may also be caused by direct inhibition of thymidylate synthetase by 5-fluorouracil. Reduced supply of both purines, deoxyadenosine triphosphate (dATP) and deoxyguanosine triphosphate (dGTP), may be caused by hydroxyurea, 6-mercaptopurine (and probably by another purine antagonist azaserine), whilst reduced supply of both pyrimidine DNA precursors, dTTP and dCTP (deoxycytidine triphosphate) may be due to inherited orotic aciduria or to treatment with azauridine. Cytosine arabinoside directly inhibits DNA polymerase. DNA replication is a discontinuous process and a number of enzymes are concerned with different aspects of the process. The parental strands partly unwind and a large number of initiation points or origins are activated on both strands. A primer RNA is first synthesised using the parental strand of DNA as template. Fragments of new DNA are then synthesised on the parental DNA template, starting at the RNA primer, under the action of one or other DNA polymerase (probably gamma). The RNA primer is then removed and the gap left is filled by further DNA synthesis under the action of a different DNA polymerase (probably alpha). The fragments of new DNA are joined to give newly synthesised stretches of DNA (replicons) which are then liigated together to form bulk DNA of enormous molecular weight. It is suggested here that reduced supply of one or other of the four deoxyribonucleoside triphosphate (dNTP) during the 'S' phase of the cell cycle (due to vitamin B12 or folate deficiency, drug treatment or other congenital or acquired abnormality in synthesis of the dNTP) impairs the cell's ability to elongate newly initiated DNA fragments by preventing gap-filling, the polymerase needed for gap-filling requiring substantially greater concentrations of the deoxyribonucleoside triphosphates than the polymerase involved in chain initiation. Cytosine arabinoside, which also may cause megaloblastosis, may affect principally the synthesis of new DNA fragments. Since active protein synthesis is needed for the cell to enter the S phase and RNA synthesis is needed to prime new DNA synthesis, megaloblastic anaemia may be expected to occur only when DNA synthesis is inhibited but protein and RNA synthesis are relatively unimpaired... | Vitamin B12--folate interrelations. Megaloblastic anaemia is due to a derangement of DNA synthesis caused by insufficient supply of one or other of the four deoxyribonucleoside triphosphate (dNTP) precursors of DNA synthesis or by direct inhibition of one or other DNA polymerase. Reduced supply of the pyrimidine deoxythymidine triphosphate (dTTP) may be caused by folate or vitamin B12 deficiencies or by the action of dihydrofolate reductase inhibitors (e.g. methotrexate, pyrimethamine or trimethoprim), all of which cause reduced supply of the coenzyme 5, 10 methylene tetrahydrofolate (pentaglutamate) needed for thymidylate synthetase. Reduced dTTP supply may also be caused by direct inhibition of thymidylate synthetase by 5-fluorouracil. Reduced supply of both purines, deoxyadenosine triphosphate (dATP) and deoxyguanosine triphosphate (dGTP), may be caused by hydroxyurea, 6-mercaptopurine (and probably by another purine antagonist azaserine), whilst reduced supply of both pyrimidine DNA precursors, dTTP and dCTP (deoxycytidine triphosphate) may be due to inherited orotic aciduria or to treatment with azauridine. Cytosine arabinoside directly inhibits DNA polymerase. DNA replication is a discontinuous process and a number of enzymes are concerned with different aspects of the process. The parental strands partly unwind and a large number of initiation points or origins are activated on both strands. A primer RNA is first synthesised using the parental strand of DNA as template. Fragments of new DNA are then synthesised on the parental DNA template, starting at the RNA primer, under the action of one or other DNA polymerase (probably gamma). The RNA primer is then removed and the gap left is filled by further DNA synthesis under the action of a different DNA polymerase (probably alpha). The fragments of new DNA are joined to give newly synthesised stretches of DNA (replicons) which are then liigated together to form bulk DNA of enormous molecular weight. It is suggested here that reduced supply of one or other of the four deoxyribonucleoside triphosphate (dNTP) during the 'S' phase of the cell cycle (due to vitamin B12 or folate deficiency, drug treatment or other congenital or acquired abnormality in synthesis of the dNTP) impairs the cell's ability to elongate newly initiated DNA fragments by preventing gap-filling, the polymerase needed for gap-filling requiring substantially greater concentrations of the deoxyribonucleoside triphosphates than the polymerase involved in chain initiation. Cytosine arabinoside, which also may cause megaloblastosis, may affect principally the synthesis of new DNA fragments. Since active protein synthesis is needed for the cell to enter the S phase and RNA synthesis is needed to prime new DNA synthesis, megaloblastic anaemia may be expected to occur only when DNA synthesis is inhibited but protein and RNA synthesis are relatively unimpaired... | [
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PMID:10123 | Effect of sulfasalazine on digoxin bioavailability. | Low levels of digoxin were noted in a patient receiving digoxin and sulfasalazine (SSA). Discontinuation of SSA resulted in a significant increase in serum digoxin levels. To determine whether or not SSA consistently interfered with the therapeutic effect of digoxin, both drugs were administered to 10 normal subjects in a crossover study. Each received 2 doses of digoxin (0.5 mg, elixir): one dose given alone, and a second dose after 6 days of treatment with SSA. When digoxin was given with SSA, the average area under the serum digoxin curve fell from the control value of 8.79 ng-hr-ml(-1) to 6.66 ng-hr-ml(-1) (p less than 0.05), fell and total urinary excretion decreased from 278 mcg/10 days to 228 mcg/10 days (p less than 0.025). These changes suggest interference with the bioavailability of digoxin by SSA. Studies were conducted to determine whether SSA inhibited digoxin absorption by physically absorbing the glycoside from solution. In vitro tests failed to reveal any significant adsorptive properties for SSA. | Effect of sulfasalazine on digoxin bioavailability. Low levels of digoxin were noted in a patient receiving digoxin and sulfasalazine (SSA). Discontinuation of SSA resulted in a significant increase in serum digoxin levels. To determine whether or not SSA consistently interfered with the therapeutic effect of digoxin, both drugs were administered to 10 normal subjects in a crossover study. Each received 2 doses of digoxin (0.5 mg, elixir): one dose given alone, and a second dose after 6 days of treatment with SSA. When digoxin was given with SSA, the average area under the serum digoxin curve fell from the control value of 8.79 ng-hr-ml(-1) to 6.66 ng-hr-ml(-1) (p less than 0.05), fell and total urinary excretion decreased from 278 mcg/10 days to 228 mcg/10 days (p less than 0.025). These changes suggest interference with the bioavailability of digoxin by SSA. Studies were conducted to determine whether SSA inhibited digoxin absorption by physically absorbing the glycoside from solution. In vitro tests failed to reveal any significant adsorptive properties for SSA. | [
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PMID:10118 | The influence of beta-adrenoceptor antagonists on accommodation of the lens. | 1. A study of the influence of single oral doses of the adrenergic-beta-receptor anatagonists alprenolol (100 mg) and propranolol (50 mg) on the accommodation of the lens was made in six healthy male subjects. 2. There was no significant influence of either drug on lens accommodation. | The influence of beta-adrenoceptor antagonists on accommodation of the lens. 1. A study of the influence of single oral doses of the adrenergic-beta-receptor anatagonists alprenolol (100 mg) and propranolol (50 mg) on the accommodation of the lens was made in six healthy male subjects. 2. There was no significant influence of either drug on lens accommodation. | [
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PMID:10124 | Pharmacokinetics of oxprenolol in normal subjects. | The effect of oxprenolol administered intravenously (10 and 20 mg) and orally (20, 40, 80, and 160 mg) on plasma concentrations of the drug, resting heart rate, exercise-induced tachycardia, and arterial blood pressure was assessed as a function of time in 6 healthy subjects. The pharmacokinetics of oxprenolol following intravenous administration are best described as 2-compartnent open model with dose-dependent parameters. The mean (+/-SD) plasma half-life for oral doses is 1.94 +/- 0.37 and for intravenous doses is 2.31 +/- 0.64 hr. After oral administration, peak plasma concentrations are reached within 30 to 90 min, and the area under the plasma concentration-time curve varies linearly with the dose. Comparison of oral and intravenous data reveals the variation in bioavailabilty of orally administered oxprenolol to range from 19% to 74%. Unlike propranolol, oxprenolol does not show a saturable "first-pass" elimination effect. Blockade of beta-receptors occurs at plasma levels in excess of 60 ng/ml as evidenced by significant reductions in resting heart rate and exercise-induced tachycardia. Higher plasma concentrations of oxprenolol are required to lower blood pressure compared to those necessary to slow heart rate. These data suggest significant pharmacokinetic differences between oxprenolol and other beta-adrenergic receptor antagonists. | Pharmacokinetics of oxprenolol in normal subjects. The effect of oxprenolol administered intravenously (10 and 20 mg) and orally (20, 40, 80, and 160 mg) on plasma concentrations of the drug, resting heart rate, exercise-induced tachycardia, and arterial blood pressure was assessed as a function of time in 6 healthy subjects. The pharmacokinetics of oxprenolol following intravenous administration are best described as 2-compartnent open model with dose-dependent parameters. The mean (+/-SD) plasma half-life for oral doses is 1.94 +/- 0.37 and for intravenous doses is 2.31 +/- 0.64 hr. After oral administration, peak plasma concentrations are reached within 30 to 90 min, and the area under the plasma concentration-time curve varies linearly with the dose. Comparison of oral and intravenous data reveals the variation in bioavailabilty of orally administered oxprenolol to range from 19% to 74%. Unlike propranolol, oxprenolol does not show a saturable "first-pass" elimination effect. Blockade of beta-receptors occurs at plasma levels in excess of 60 ng/ml as evidenced by significant reductions in resting heart rate and exercise-induced tachycardia. Higher plasma concentrations of oxprenolol are required to lower blood pressure compared to those necessary to slow heart rate. These data suggest significant pharmacokinetic differences between oxprenolol and other beta-adrenergic receptor antagonists. | [
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PMID:10125 | Clinical pharmacologic observations on atenolol, a beta-adrenoceptor blocker. | The effects of oral and intravenous administration of atenolol were studied in healthy volunteers. The oral administration of a series of single doses of atenolol reduced an exercise tachycardia. After a 200-mg dose, the effect on an exercise tachycardia was maximal at 3 hr and declined linearly with time at a rate of approximately 10% per 24 hr. The peak plasma atenolol concentration occurred at 3 hr and thereafter declined exponentially with time with an elimination half-life of 6.36 +/- 0.55 hr: 43 +/- 3.9% of the dose was excreted in the urine within 72 hr. There was a correlation between the reduction in an exercise tachycardia and the logarithm of the corresponding plasma concentration. The intravenous administration of atenolol reduced exercise tachycardia with a significant correlation between effect and plasma concentration. After 50 mg intravenously, 100% of the dose was recovered from the urine, and the clearance was 97.3 ml/min. Comparison of AUC O leads to chi after oral and intravenous administration of 50 mg showed the bioavailability to be 63% after oral drug. Repeated oral administration of atenolol 200 mg daily either as a single dose or in divided 12 hourly doses for 8 days maintained reduction of an exercise tachycardia of at least 24% during the period of drug administration. The plasma elimination half-life, area under the plasma concentration-time curve, and peak plasma concentration after 200 mg atenolol were not changed by chronic dosing for 8 days. | Clinical pharmacologic observations on atenolol, a beta-adrenoceptor blocker. The effects of oral and intravenous administration of atenolol were studied in healthy volunteers. The oral administration of a series of single doses of atenolol reduced an exercise tachycardia. After a 200-mg dose, the effect on an exercise tachycardia was maximal at 3 hr and declined linearly with time at a rate of approximately 10% per 24 hr. The peak plasma atenolol concentration occurred at 3 hr and thereafter declined exponentially with time with an elimination half-life of 6.36 +/- 0.55 hr: 43 +/- 3.9% of the dose was excreted in the urine within 72 hr. There was a correlation between the reduction in an exercise tachycardia and the logarithm of the corresponding plasma concentration. The intravenous administration of atenolol reduced exercise tachycardia with a significant correlation between effect and plasma concentration. After 50 mg intravenously, 100% of the dose was recovered from the urine, and the clearance was 97.3 ml/min. Comparison of AUC O leads to chi after oral and intravenous administration of 50 mg showed the bioavailability to be 63% after oral drug. Repeated oral administration of atenolol 200 mg daily either as a single dose or in divided 12 hourly doses for 8 days maintained reduction of an exercise tachycardia of at least 24% during the period of drug administration. The plasma elimination half-life, area under the plasma concentration-time curve, and peak plasma concentration after 200 mg atenolol were not changed by chronic dosing for 8 days. | [
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PMID:10126 | Psychiatric sequelae of phencyclidine abuse. | Phencyclidine use has been noted to produce a psychosis of several week's duration in a small fraction of users. Descriptions of the premorbid personalities of those who became psychotic resemble descriptions of LSD and marijuana users who experienced prolonged psychiatric difficulty. In addition, the psychosis produced can often be recognized as a "hallucinogen" psychosis. Certain features of the phencyclidine psychosis, namely the neurologic abnormalities, dose-related severity of symptoms, and regularity of the length of illness, are not noted with other psychedelic drugs, leading to the conclusion that PCP psychosis is a drug effect rather than a brief functional psychosis precipitated by the disintegrating PCP experience. However, the infrequent occurrence of psychosis in the (apparently) large exposed population still suggests that this is a combination of drug effect and vulnerable, pathologic personality. | Psychiatric sequelae of phencyclidine abuse. Phencyclidine use has been noted to produce a psychosis of several week's duration in a small fraction of users. Descriptions of the premorbid personalities of those who became psychotic resemble descriptions of LSD and marijuana users who experienced prolonged psychiatric difficulty. In addition, the psychosis produced can often be recognized as a "hallucinogen" psychosis. Certain features of the phencyclidine psychosis, namely the neurologic abnormalities, dose-related severity of symptoms, and regularity of the length of illness, are not noted with other psychedelic drugs, leading to the conclusion that PCP psychosis is a drug effect rather than a brief functional psychosis precipitated by the disintegrating PCP experience. However, the infrequent occurrence of psychosis in the (apparently) large exposed population still suggests that this is a combination of drug effect and vulnerable, pathologic personality. | [
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PMID:10134 | Intrarenal arterial aneurysms. | Intraparenchymal renal aneurysms have been reported with increasing frequency; yet, to our knowledge, this subject has not been reviewed in radiologic literature. The spectrum of such aneurysms includes congenital aneurysms, those secondary to disease usually affecting the main renal arteries, those associated with renal masses, microaneurysms, and false or pseudo aneurysms. Seemingly unrelated conditions, such as atherosclerosis, bacterial endocarditis, and trauma, can all produce similar radiographic appearance of aneurysmal dilatation within the kidney, albeit through differing mechanisms. In addition, there are several etiologies for renal microaneurysms, even though this finding has been considered specific for polyarteritis in the past. Although there were a few guidelines for recognizing certain specific etiologies based solely on the angiographic appearance, it must be appreciated that many of these conditions may be indistinguishable. The possibility of hemorrhage from such intrarenal aneurysms, and the question of whether such lesions are responsible for renovascular hypertension are also discussed. | Intrarenal arterial aneurysms. Intraparenchymal renal aneurysms have been reported with increasing frequency; yet, to our knowledge, this subject has not been reviewed in radiologic literature. The spectrum of such aneurysms includes congenital aneurysms, those secondary to disease usually affecting the main renal arteries, those associated with renal masses, microaneurysms, and false or pseudo aneurysms. Seemingly unrelated conditions, such as atherosclerosis, bacterial endocarditis, and trauma, can all produce similar radiographic appearance of aneurysmal dilatation within the kidney, albeit through differing mechanisms. In addition, there are several etiologies for renal microaneurysms, even though this finding has been considered specific for polyarteritis in the past. Although there were a few guidelines for recognizing certain specific etiologies based solely on the angiographic appearance, it must be appreciated that many of these conditions may be indistinguishable. The possibility of hemorrhage from such intrarenal aneurysms, and the question of whether such lesions are responsible for renovascular hypertension are also discussed. | [
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PMID:10136 | Relationship between pulmonary hemodynamics and arterial pH and carbon dioxide tension in critically ill patients. | To ascertain the clinical significance of derangements in arterial pH and arterial carbon dioxide tension (PaCO2) in modifying pulmonary arterial pressures and pulmonary vascular resistance in critically ill patients, the relationship between these two sets of variables was evaluated in 75 patients. No significant differences in pulmonary hemodynamic values were found among patients with acidemia, a normal pH, or alkalemia, even at extreme pH values; and there was no consistent relationship between PaCO2 and each of the pulmonary hemodynamic measurements. In patients who initially had a normal pH but subsequently developed acidemia or alkalemia, there was also no significant correlation between changes in pH and pulmonary hemodynamic values. We conclude that abnormalities of pulmonary hemodynamic values in seriously ill patients are usually due to factors other than acid-base derangements. Of practical importance is the observation that the predictability of the pulmonary arterial wedge pressure from the pulmonary arterial diastolic pressure is not invalidated by acid-base disturbances. | Relationship between pulmonary hemodynamics and arterial pH and carbon dioxide tension in critically ill patients. To ascertain the clinical significance of derangements in arterial pH and arterial carbon dioxide tension (PaCO2) in modifying pulmonary arterial pressures and pulmonary vascular resistance in critically ill patients, the relationship between these two sets of variables was evaluated in 75 patients. No significant differences in pulmonary hemodynamic values were found among patients with acidemia, a normal pH, or alkalemia, even at extreme pH values; and there was no consistent relationship between PaCO2 and each of the pulmonary hemodynamic measurements. In patients who initially had a normal pH but subsequently developed acidemia or alkalemia, there was also no significant correlation between changes in pH and pulmonary hemodynamic values. We conclude that abnormalities of pulmonary hemodynamic values in seriously ill patients are usually due to factors other than acid-base derangements. Of practical importance is the observation that the predictability of the pulmonary arterial wedge pressure from the pulmonary arterial diastolic pressure is not invalidated by acid-base disturbances. | [
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PMID:10139 | Polyamines and drug oxidations. | The addition of spermine or of spermidine to rat liver assay systems produced marked changes in a number of microsomal drug oxidations. The hydroxylation of aniline and the N-demethylation of ethylmorphine were both enhanced with concentrations of 1-10 mM spermine or of spermidine. The results with putrescine on ethylmorphine metabolism were less dramatic, and no effect could be observed with putrescine in studies with other drug substrates. In contrast to the enhancing effects, inhibition was observed when spermine or spermidine was added to p-nitroanisole O-demethylation assay mixtures, and no effect was observed in assays for acetanilide hydroxylation. The inhibiting and enhancing effects of the polyamines can be observed in assays containing liver preparations from both male and female rats, and those from rats pretreated with phenobarbital or 3-methylcholanthrene. In all studies, the alterations were kinetically noncompetitive. The effects were shown to be independent of the NADPH-generating system and the cation requirements, and were not mediated through an interaction with NADPH-cytochrome c reductase. The possibility is considered that the enhancing and inhibiting effects may be related to the ability of these polycations to bind to microsomal membranes and cause alterations at different sites of substrate interaction. | Polyamines and drug oxidations. The addition of spermine or of spermidine to rat liver assay systems produced marked changes in a number of microsomal drug oxidations. The hydroxylation of aniline and the N-demethylation of ethylmorphine were both enhanced with concentrations of 1-10 mM spermine or of spermidine. The results with putrescine on ethylmorphine metabolism were less dramatic, and no effect could be observed with putrescine in studies with other drug substrates. In contrast to the enhancing effects, inhibition was observed when spermine or spermidine was added to p-nitroanisole O-demethylation assay mixtures, and no effect was observed in assays for acetanilide hydroxylation. The inhibiting and enhancing effects of the polyamines can be observed in assays containing liver preparations from both male and female rats, and those from rats pretreated with phenobarbital or 3-methylcholanthrene. In all studies, the alterations were kinetically noncompetitive. The effects were shown to be independent of the NADPH-generating system and the cation requirements, and were not mediated through an interaction with NADPH-cytochrome c reductase. The possibility is considered that the enhancing and inhibiting effects may be related to the ability of these polycations to bind to microsomal membranes and cause alterations at different sites of substrate interaction. | [
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PMID:10140 | Drug biotransformation in microsomes from the fetal stumptailed macaque, Macaca arctoides: hepatic N-demethylation. | The kinetics of the N-demethylation of benzphetamine, ethylmorphine, meperidine, and methadone have been studied in microsomes isolated from livers of the fetal stumptailed macaque (Macaca arctoides) during the last third of gestation. The apparent KM for each substrate did not change during this time period. Values were similar to those from livers of adult African green monkeys. The Vmax for each substrate, when expressed per mg of microsomal protein, did not change during the last third of gestation. N-demethylase activity (Vmax) per g of liver increased during the last third of gestation, as did the content of microsomal protein, cytochrome P-450 concentration, and liver weight. The amount of cytochrome P-450 per g of liver was greater in whole homogenates of the left physiological lobe than in those of the right physiological lobe of fetal liver obtained near term; no differences occurred in whole homogenates of the separate lobes of adult liver. This observation suggests that a differential capacity for drug (and possibly steroid) metabolism may exist between the two physiological lobes of fetal liver. | Drug biotransformation in microsomes from the fetal stumptailed macaque, Macaca arctoides: hepatic N-demethylation. The kinetics of the N-demethylation of benzphetamine, ethylmorphine, meperidine, and methadone have been studied in microsomes isolated from livers of the fetal stumptailed macaque (Macaca arctoides) during the last third of gestation. The apparent KM for each substrate did not change during this time period. Values were similar to those from livers of adult African green monkeys. The Vmax for each substrate, when expressed per mg of microsomal protein, did not change during the last third of gestation. N-demethylase activity (Vmax) per g of liver increased during the last third of gestation, as did the content of microsomal protein, cytochrome P-450 concentration, and liver weight. The amount of cytochrome P-450 per g of liver was greater in whole homogenates of the left physiological lobe than in those of the right physiological lobe of fetal liver obtained near term; no differences occurred in whole homogenates of the separate lobes of adult liver. This observation suggests that a differential capacity for drug (and possibly steroid) metabolism may exist between the two physiological lobes of fetal liver. | [
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PMID:10141 | N-Hydroxylation of phenacetin by hamster liver microsomes. | Hamster liver microsomes have been shown to catalyze the N-hydroxylation of phenacetin. The reaction, which requires oxygen and NADPH, is inhibited by a carbon monoxide/oxygen atmosphere, indicating that it is catalyzed by a cytochrome P-450-dependent mixed-function oxidase. The N-hydroxyphenacetin can be further metabolized by the microsomes, and the reaction is inhibited by phenacetin. | N-Hydroxylation of phenacetin by hamster liver microsomes. Hamster liver microsomes have been shown to catalyze the N-hydroxylation of phenacetin. The reaction, which requires oxygen and NADPH, is inhibited by a carbon monoxide/oxygen atmosphere, indicating that it is catalyzed by a cytochrome P-450-dependent mixed-function oxidase. The N-hydroxyphenacetin can be further metabolized by the microsomes, and the reaction is inhibited by phenacetin. | [
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PMID:10142 | Properties of microsomal enzyme systems that reduce N-hydroxyphentermine. | The reduction of N-hydroxyphentermine was studied in liver microsomes isolated from rat, guinea pig, and rabbit. The reduction requires a NADPH-generating system and was inhibited by oxygen and carbon monoxide. In the rat, the reduction was increased by phenobarbital pretreatment. Kinetic analysis of the reductase activity in rat liver microsomes suggests that the reduction of the hydroxylamine is mediated by at least two enzyme systems, one of which is a CO-sensitive system inducible by phenobarbital. | Properties of microsomal enzyme systems that reduce N-hydroxyphentermine. The reduction of N-hydroxyphentermine was studied in liver microsomes isolated from rat, guinea pig, and rabbit. The reduction requires a NADPH-generating system and was inhibited by oxygen and carbon monoxide. In the rat, the reduction was increased by phenobarbital pretreatment. Kinetic analysis of the reductase activity in rat liver microsomes suggests that the reduction of the hydroxylamine is mediated by at least two enzyme systems, one of which is a CO-sensitive system inducible by phenobarbital. | [
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PMID:10143 | Cytochrome P-450 content and mixed-function oxidase activity in microsomes isolated from mouse skin. | A microsomal fraction, prepared from mouse skin, catalyzed the hydroxylation of benzpyrene and aniline and the deethylation of 7-ethoxycoumarin. Contamination of the preparation by cytochrome oxidase and cytochrome P-420 was determined by spectral analysis. The enzyme activities studied in mouse skin (Swiss-Webster CD-1) did not respond to topical application of 3-MC. Twenty-four hours after topical application of TCDD to mice, microsomes from skin had 50% greater benzpyrene hydroxylase and 7-ethoxycoumarin deethylase activity, and 4- to 8-fold greater activity of these enzymes was seen after 72 hr. Increases in cytochrome P-450 content of skin microsomes could be demonstrated 24 and 72 hr after topical TCDD treatment of mice. Cholate treatment (solubilization) of skin microsomes, followed by centrifugation, removed the contaminating cytochrome oxidase. Quantitative and qualitative analyses of cytochrome P-450 difference spectra were made from the solubilized preparations. | Cytochrome P-450 content and mixed-function oxidase activity in microsomes isolated from mouse skin. A microsomal fraction, prepared from mouse skin, catalyzed the hydroxylation of benzpyrene and aniline and the deethylation of 7-ethoxycoumarin. Contamination of the preparation by cytochrome oxidase and cytochrome P-420 was determined by spectral analysis. The enzyme activities studied in mouse skin (Swiss-Webster CD-1) did not respond to topical application of 3-MC. Twenty-four hours after topical application of TCDD to mice, microsomes from skin had 50% greater benzpyrene hydroxylase and 7-ethoxycoumarin deethylase activity, and 4- to 8-fold greater activity of these enzymes was seen after 72 hr. Increases in cytochrome P-450 content of skin microsomes could be demonstrated 24 and 72 hr after topical TCDD treatment of mice. Cholate treatment (solubilization) of skin microsomes, followed by centrifugation, removed the contaminating cytochrome oxidase. Quantitative and qualitative analyses of cytochrome P-450 difference spectra were made from the solubilized preparations. | [
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PMID:10144 | Preservation of various microsomal drug metabolizing components in tissue preparations from the livers, lungs, and small intestines of rodents. | The livers, lungs, and small intestines of untreated rabbits and the livers of control rats were stored intact, or as microsomal suspensions, under liquid nitrogen at -196 degrees C. Aniline hydroxylase, aminopyrine demethylase, benzpyrene hydroxylase, biphenyl hydroxylase, NADPH-cytochrome c reductase, UDP-glucuronyltransferase activities, the microsomal content of cytochrome P-450, and the aniline- and benzphetamine-induced spectral changes were compared in fresh and stored preparations. Few significant changes in any of the above parameters resulted from storage of rabbit tissue preparations in liquid nitrogen for periods of up to 28 days. Pretreatment of rabbits with phenobarbital did not affect the stability of their stored microsomal preparations. Enzyme activities in the livers of untreated or 3-methylcholanthrene-pretreated rats were less stable to storage than in tissue preparations from rabbits stored under identical conditions. However, when rat hepatic microsomes were resuspended in KCl-HEPES supplemented with 1 mM EDTA before storage, enzyme activities were largely unaffected by freezing in liquid nitrogen. | Preservation of various microsomal drug metabolizing components in tissue preparations from the livers, lungs, and small intestines of rodents. The livers, lungs, and small intestines of untreated rabbits and the livers of control rats were stored intact, or as microsomal suspensions, under liquid nitrogen at -196 degrees C. Aniline hydroxylase, aminopyrine demethylase, benzpyrene hydroxylase, biphenyl hydroxylase, NADPH-cytochrome c reductase, UDP-glucuronyltransferase activities, the microsomal content of cytochrome P-450, and the aniline- and benzphetamine-induced spectral changes were compared in fresh and stored preparations. Few significant changes in any of the above parameters resulted from storage of rabbit tissue preparations in liquid nitrogen for periods of up to 28 days. Pretreatment of rabbits with phenobarbital did not affect the stability of their stored microsomal preparations. Enzyme activities in the livers of untreated or 3-methylcholanthrene-pretreated rats were less stable to storage than in tissue preparations from rabbits stored under identical conditions. However, when rat hepatic microsomes were resuspended in KCl-HEPES supplemented with 1 mM EDTA before storage, enzyme activities were largely unaffected by freezing in liquid nitrogen. | [
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PMID:10145 | Role of blood flow in carbon monoxide- and hypoxic hypoxia-induced alterations in hexobarbital metabolism in rats. | The flow dependency of hepatic hexobarbital metabolism was examined in the isolated perfused rat liver. At low flow rates (0.5-1.0 ml/min/g of liver) hexobarbital clearance was found to depend on perfusion fluid flow, whereas at higher flow rates drug clearnace approached flow independence. Calculation of the in vivo hepatic blood flow rate suggested that hexobarbital metabolism in vivo should be highly dependent upon flow. Blood flow in the conscious rat was measured by use of radiolabeled microspheres during acute exposure to levels of hypoxic hypoxia (lowered pO2) or carbon monoxide which resulted in equal alterations in arterial oxyhemoglobin content (approximately 65% oxyhemoglobin). Hypoxic hypoxia (8% O2) caused a massive redistribution of flow away from the splanchnic area, resulting in a 45% decrease in hepatic blood flow. Carbon monoxide (500 ppm) was without significant effect on hepatic blood flow. These data would appear to explain the relatively greater inhibitory potency of hypoxic hypoxia on drug metabolism in vivo, since drug delivery to the liver is depressed by hypoxic hypoxia but unaffected by carbon monoxide exposure. | Role of blood flow in carbon monoxide- and hypoxic hypoxia-induced alterations in hexobarbital metabolism in rats. The flow dependency of hepatic hexobarbital metabolism was examined in the isolated perfused rat liver. At low flow rates (0.5-1.0 ml/min/g of liver) hexobarbital clearance was found to depend on perfusion fluid flow, whereas at higher flow rates drug clearnace approached flow independence. Calculation of the in vivo hepatic blood flow rate suggested that hexobarbital metabolism in vivo should be highly dependent upon flow. Blood flow in the conscious rat was measured by use of radiolabeled microspheres during acute exposure to levels of hypoxic hypoxia (lowered pO2) or carbon monoxide which resulted in equal alterations in arterial oxyhemoglobin content (approximately 65% oxyhemoglobin). Hypoxic hypoxia (8% O2) caused a massive redistribution of flow away from the splanchnic area, resulting in a 45% decrease in hepatic blood flow. Carbon monoxide (500 ppm) was without significant effect on hepatic blood flow. These data would appear to explain the relatively greater inhibitory potency of hypoxic hypoxia on drug metabolism in vivo, since drug delivery to the liver is depressed by hypoxic hypoxia but unaffected by carbon monoxide exposure. | [
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PMID:10146 | The time course of tolmetin and its metabolites in the plasma of individual rats and mice. | Tolmetin, 1-methyl-(5-p-toluoyl)pyrrole-2-acetic acid, is a new, nonsteroidal, anti-inflammatory agent. After oral administration of tolmetin-14C to rats and mice, sequential microsamples of blood were obtained from the ophthalmic venous plexus via the orbital sinus. Plasma was collected after centrifugation, and microaliquots of each plasma sample were analyzed. The total radioactivity and thin-layer chromatographic assays used permitted quantitation of tolmetin, its dicarboxylic acid metabolite, and (by difference) all other metabolites collectively for each sample. Time-course data on plasma levels were obtained for individual rats and mice. The plasma elimination half-life of tolmetin was estimated at 0.67 +/- 0.13 hr (mean +/- SD) in male rats, 1.4 +/- 0.6 hr in female rats, 1.2 +/- 0.3 hr in male mice, and 1.0 +/- 0.0 hr in female mice. A one-compartment open model was used. | The time course of tolmetin and its metabolites in the plasma of individual rats and mice. Tolmetin, 1-methyl-(5-p-toluoyl)pyrrole-2-acetic acid, is a new, nonsteroidal, anti-inflammatory agent. After oral administration of tolmetin-14C to rats and mice, sequential microsamples of blood were obtained from the ophthalmic venous plexus via the orbital sinus. Plasma was collected after centrifugation, and microaliquots of each plasma sample were analyzed. The total radioactivity and thin-layer chromatographic assays used permitted quantitation of tolmetin, its dicarboxylic acid metabolite, and (by difference) all other metabolites collectively for each sample. Time-course data on plasma levels were obtained for individual rats and mice. The plasma elimination half-life of tolmetin was estimated at 0.67 +/- 0.13 hr (mean +/- SD) in male rats, 1.4 +/- 0.6 hr in female rats, 1.2 +/- 0.3 hr in male mice, and 1.0 +/- 0.0 hr in female mice. A one-compartment open model was used. | [
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PMID:10147 | The urinary excretion profiles of naltrexone in man, monkey, rabbit, and rat. | A gas-chromatographic method has been developed for the simultaneous determination of naltrexone, alpha-naltrexol, and beta-naltrexol as trimethylsiyl derivatives. Analysis of urine from rabbit, monkey, and rat demonstrated that, like man, these species reduce naltrexone primarily to beta-naltrexol. In naltrexone maintenance patients receiving 125 mg po three times per week, an average of 37% of the dose was recovered in 48-hr urine as free naltrexone (0.8%), conjugated naltrexone (7.6%), free beta-naltrexol (16.8%), and conjugated beta-naltrexol (11.8%). Thirty-four percent of the dose appeared in 0-24 hr and 3% during 24-48 hr. The ratio of beta-naltrexol to naltrexone rose from 2 at 0-4 hr to 34-48 hr. Monkeys receiving a daily dose of 12 mg/kg po, chronically, excreted very little free beta-naltrexol and exhibited an apparent sex-related difference in excretion patterns, with females excreting more than twice as much total base as males. Rabbits given a dose of 30 mg/kg ip for 4 days excreted conjugated naltrexone as the predominant urinary metabolite, accounting for 80% of total base recovered in 24 hr. In rats receiving 100 mg/kg po, less than 1% of the administered dose could be accounted for in the 24-hr urine, indicating that although the beta-naltrexol is produced as a urinary metabolite, other means of disposition of the drug must exist. Thus, in man and the monkey, beta-naltrexol is the predominant and persistent urinary metabolite. Urinary excretion profiles of naltrexone differ greatly between species commonly examined for chronic toxicity studies. | The urinary excretion profiles of naltrexone in man, monkey, rabbit, and rat. A gas-chromatographic method has been developed for the simultaneous determination of naltrexone, alpha-naltrexol, and beta-naltrexol as trimethylsiyl derivatives. Analysis of urine from rabbit, monkey, and rat demonstrated that, like man, these species reduce naltrexone primarily to beta-naltrexol. In naltrexone maintenance patients receiving 125 mg po three times per week, an average of 37% of the dose was recovered in 48-hr urine as free naltrexone (0.8%), conjugated naltrexone (7.6%), free beta-naltrexol (16.8%), and conjugated beta-naltrexol (11.8%). Thirty-four percent of the dose appeared in 0-24 hr and 3% during 24-48 hr. The ratio of beta-naltrexol to naltrexone rose from 2 at 0-4 hr to 34-48 hr. Monkeys receiving a daily dose of 12 mg/kg po, chronically, excreted very little free beta-naltrexol and exhibited an apparent sex-related difference in excretion patterns, with females excreting more than twice as much total base as males. Rabbits given a dose of 30 mg/kg ip for 4 days excreted conjugated naltrexone as the predominant urinary metabolite, accounting for 80% of total base recovered in 24 hr. In rats receiving 100 mg/kg po, less than 1% of the administered dose could be accounted for in the 24-hr urine, indicating that although the beta-naltrexol is produced as a urinary metabolite, other means of disposition of the drug must exist. Thus, in man and the monkey, beta-naltrexol is the predominant and persistent urinary metabolite. Urinary excretion profiles of naltrexone differ greatly between species commonly examined for chronic toxicity studies. | [
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PMID:10148 | The biotransformation of (6,7-dichloro-2-methyl-1-oxo-2-phenyl-5-indanyloxy) acetic acid (MK-196) in the chimpanzee. | The metabolism of a novel polyvalent saluretic agent (6,7-dichloro-2-methyl-1-oxo-2-phenyl-5-indanyloxy)acetic acid (MK-196) was studied in the chimpanzee. Following oral administration, 50% of the radioactive dose was recovered in the urine in four days; 8-14% of the dose was excreted as unchanged drug. The fecal specimens accounted for 5-9% of the dose. Following intravenous administration an initial rapid elimination of drug from the plasma was observed [(t1/2)alpha approximately 0.4 hr, (t1/2)beta approximately 4 hr]. The data are consistent with the rapid elimination of radioactivity, approximately 30% of dose, in the urine during the first 24 hr, followed by a much slower rate of excretion of drug and metabolites. These findings are congruous with the high affinity (greater than 98%) of MK-196 and the major metabolite with plasma proteins. The urinary metabolites were isolated and identified by the following techniques: solvent extraction, column, thin-layer, and gas-liquid chromatography, derivatization, and mass and nuclear magnetic resonance spectroscopy. The major metabolite, which resulted from para-hydroxylation of the 2-phenyl substitutent, accounted for about 40% of the urinary radioactivity. Reduction of the ketone group, methylation of the p-hydroxy group, and additional phenyl ring hydroxylation were also shown to occur. There was no evidence for glucuronide formation nor did SKF-525-A inhibit the metabolism of the drug in the chimpanzee. Under conditions of induced metabolic alkalosis, the urinary levels of MK-196 increased from 11 to 40%. Probenecid and p-aminohippurate administered during metabolic alkalosis decreased the clearance of drug (40 to 15%). | The biotransformation of (6,7-dichloro-2-methyl-1-oxo-2-phenyl-5-indanyloxy) acetic acid (MK-196) in the chimpanzee. The metabolism of a novel polyvalent saluretic agent (6,7-dichloro-2-methyl-1-oxo-2-phenyl-5-indanyloxy)acetic acid (MK-196) was studied in the chimpanzee. Following oral administration, 50% of the radioactive dose was recovered in the urine in four days; 8-14% of the dose was excreted as unchanged drug. The fecal specimens accounted for 5-9% of the dose. Following intravenous administration an initial rapid elimination of drug from the plasma was observed [(t1/2)alpha approximately 0.4 hr, (t1/2)beta approximately 4 hr]. The data are consistent with the rapid elimination of radioactivity, approximately 30% of dose, in the urine during the first 24 hr, followed by a much slower rate of excretion of drug and metabolites. These findings are congruous with the high affinity (greater than 98%) of MK-196 and the major metabolite with plasma proteins. The urinary metabolites were isolated and identified by the following techniques: solvent extraction, column, thin-layer, and gas-liquid chromatography, derivatization, and mass and nuclear magnetic resonance spectroscopy. The major metabolite, which resulted from para-hydroxylation of the 2-phenyl substitutent, accounted for about 40% of the urinary radioactivity. Reduction of the ketone group, methylation of the p-hydroxy group, and additional phenyl ring hydroxylation were also shown to occur. There was no evidence for glucuronide formation nor did SKF-525-A inhibit the metabolism of the drug in the chimpanzee. Under conditions of induced metabolic alkalosis, the urinary levels of MK-196 increased from 11 to 40%. Probenecid and p-aminohippurate administered during metabolic alkalosis decreased the clearance of drug (40 to 15%). | [
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PMID:10152 | [A new immunochemical tube test for pregnancy, using the principle of the latex-agglutination inhibition reaction. I. Basic studies (author's transl)]. | In a joint study of five laboratories the immunochemical tube test for pregnancy 'Roche', which uses the principle of the latex-agglutination inhibition test, was examined. A total of 1117 samples, complemented by a number of laboratory studies, proved that the test is specific, reproducible and low in interference with a sensitivity of 900-1000 U/l HCG. It is a simple test, the results can be easily read after 90 minutes. pH of the urine sample, protein, urea or calcium content, presence of oral contraceptives, detergents, disinfectants or organic solvents cause no or very little interference. | [A new immunochemical tube test for pregnancy, using the principle of the latex-agglutination inhibition reaction. I. Basic studies (author's transl)]. In a joint study of five laboratories the immunochemical tube test for pregnancy 'Roche', which uses the principle of the latex-agglutination inhibition test, was examined. A total of 1117 samples, complemented by a number of laboratory studies, proved that the test is specific, reproducible and low in interference with a sensitivity of 900-1000 U/l HCG. It is a simple test, the results can be easily read after 90 minutes. pH of the urine sample, protein, urea or calcium content, presence of oral contraceptives, detergents, disinfectants or organic solvents cause no or very little interference. | [
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PMID:10149 | The physiological disposition of the uricosuric-saluretic agent (6,7-dichloro-2-methyl-1-oxo-2-phenyl-5-indanyloxy)acetic acid (MK-196) in the rat, dog, and monkey. | The physiological disposition of a new saluretic-uricosuric agent, (6,7-dichloro-2-methyl-1-oxo-2-phenyl-5-indanyloxy)acetic acid (MK-196), was studied in the rat, dog, and monkey. MK-196 was well absorbed and showed minimal metabolism in these species. Peak plasma levels of radioactivity and drug occurred 0.5-2 hr after oral administration at a dose of 2.5 mg/kg. Essentially all of the radioactivity present in the plasma during the first day was intact MK-196. Following a single dose, a long terminal half-life for plasma radioactivity was observed in the dog (approximately 68 hr) and monkey (approximately 105 hr). The chronic administration of MK-196 to dogs resulted in a dose-related plasma profile and showed no tendency to increase or decrease with dosing. However, upon repeated drug administration to monkeys, the plasma levels of drug increased and then decreased, possibly due to hypochloremia and secondary metabolic alkalosis. Fecal excretion was the predominant route of tracer elimination in the dog (approximately 80%) and rat (approximately 94%), whereas the monkey eliminated the majority of the dose (approximately 60%) via the urine. Minimal metabolism was noted in the three lower species; most of the urinary, plasma, and fecal radioactivity was accounted for as intact drug and its glucuronide conjugate. Three minor metabolites, which were present in dog bile, plasma, and urine, were characterized as: (l,7-dichloro-1alpha-hydroxy-2-methyl-2-phenyl-5-indanyloxy)acetic acid, I; (6,7-dichloro-2-(4-hydroxyphenyl)-2-methyl-2-oxo-5-indanyloxy)acetic acid, II; and 2-methyl-2-phenyl-5-hydroxy-6,7-dichloro-1-indanone, III. The monkey urine and plasma also contained small amounts of II. | The physiological disposition of the uricosuric-saluretic agent (6,7-dichloro-2-methyl-1-oxo-2-phenyl-5-indanyloxy)acetic acid (MK-196) in the rat, dog, and monkey. The physiological disposition of a new saluretic-uricosuric agent, (6,7-dichloro-2-methyl-1-oxo-2-phenyl-5-indanyloxy)acetic acid (MK-196), was studied in the rat, dog, and monkey. MK-196 was well absorbed and showed minimal metabolism in these species. Peak plasma levels of radioactivity and drug occurred 0.5-2 hr after oral administration at a dose of 2.5 mg/kg. Essentially all of the radioactivity present in the plasma during the first day was intact MK-196. Following a single dose, a long terminal half-life for plasma radioactivity was observed in the dog (approximately 68 hr) and monkey (approximately 105 hr). The chronic administration of MK-196 to dogs resulted in a dose-related plasma profile and showed no tendency to increase or decrease with dosing. However, upon repeated drug administration to monkeys, the plasma levels of drug increased and then decreased, possibly due to hypochloremia and secondary metabolic alkalosis. Fecal excretion was the predominant route of tracer elimination in the dog (approximately 80%) and rat (approximately 94%), whereas the monkey eliminated the majority of the dose (approximately 60%) via the urine. Minimal metabolism was noted in the three lower species; most of the urinary, plasma, and fecal radioactivity was accounted for as intact drug and its glucuronide conjugate. Three minor metabolites, which were present in dog bile, plasma, and urine, were characterized as: (l,7-dichloro-1alpha-hydroxy-2-methyl-2-phenyl-5-indanyloxy)acetic acid, I; (6,7-dichloro-2-(4-hydroxyphenyl)-2-methyl-2-oxo-5-indanyloxy)acetic acid, II; and 2-methyl-2-phenyl-5-hydroxy-6,7-dichloro-1-indanone, III. The monkey urine and plasma also contained small amounts of II. | [
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PMID:10153 | Antianxiety drugs: clinical pharmacology and therapeutic use. | It is difficult to choose among the many drugs advocated for treating anxiety symptoms. The barbiturates were the most commonly used antianxiety agents until recently but are being superseded by the benzodiazepines. The latter are more effective than the barbiturates as shown in comparative clinical trials, they are safer in overdosage (deliberate or accidental), and they are somewhat less likely to induce dependence. The barbiturates have the additional drawback of interfering with the action of other drugs by inducing liver microsomal (oxidising) drug metabolising enzymes. The major tranquilisers (neuroleptics or antipsychotics) are often of value in low dosage in patients with a previous history of dependence on alcohol, the barbiturates or the benzodiazepines. Tricyclic antidepressants are the treatment of choice in anxious and depressed patients and monoamine oxidase inhibitors may be helpful in phobic patients. The beta-adrenoreceptor blocking agents such as propranolol often ameliorate somatic symptoms such as palpitations and tremor. In the treatment of anxious patients it is important to remove causes for anxiety and to limit any course of drug treatment to a finite period. Both dosage level and dosage interval should be flexible. Benzodiazepines remain the drug treatment of choice. | Antianxiety drugs: clinical pharmacology and therapeutic use. It is difficult to choose among the many drugs advocated for treating anxiety symptoms. The barbiturates were the most commonly used antianxiety agents until recently but are being superseded by the benzodiazepines. The latter are more effective than the barbiturates as shown in comparative clinical trials, they are safer in overdosage (deliberate or accidental), and they are somewhat less likely to induce dependence. The barbiturates have the additional drawback of interfering with the action of other drugs by inducing liver microsomal (oxidising) drug metabolising enzymes. The major tranquilisers (neuroleptics or antipsychotics) are often of value in low dosage in patients with a previous history of dependence on alcohol, the barbiturates or the benzodiazepines. Tricyclic antidepressants are the treatment of choice in anxious and depressed patients and monoamine oxidase inhibitors may be helpful in phobic patients. The beta-adrenoreceptor blocking agents such as propranolol often ameliorate somatic symptoms such as palpitations and tremor. In the treatment of anxious patients it is important to remove causes for anxiety and to limit any course of drug treatment to a finite period. Both dosage level and dosage interval should be flexible. Benzodiazepines remain the drug treatment of choice. | [
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PMID:10150 | Biliary copper excretion in the rat is enhanced by spironolactone. | Tissue distribution and excretion (urinary, fecal, and biliary) of an intravenous bolus of 64Cu(NO3)2 were measured in rats pretreated with spironolactone and in controls. Intact animals pretreated with spironolactone excreted 10% more of a standard intravenous injection of copper in the first 24 hr than did controls. At the end of that time, kidney, red blood cell, and serum copper levels all were similar for the two groups, but liver copper concentrations were higher in controls. During the 1st hr after copper injection, plasma copper levels tended to fall more rapidly in pretreated animals, whereas liver copper concentrations increased more rapidly; red blood cell copper concentrations were not higher in animals given spironolactone. Pretreated animals excreted significantly more copper in the bile during the first 2 hr after 64Cu(NO3)2 injection, and had higher hepatic copper levels at 3 hr. | Biliary copper excretion in the rat is enhanced by spironolactone. Tissue distribution and excretion (urinary, fecal, and biliary) of an intravenous bolus of 64Cu(NO3)2 were measured in rats pretreated with spironolactone and in controls. Intact animals pretreated with spironolactone excreted 10% more of a standard intravenous injection of copper in the first 24 hr than did controls. At the end of that time, kidney, red blood cell, and serum copper levels all were similar for the two groups, but liver copper concentrations were higher in controls. During the 1st hr after copper injection, plasma copper levels tended to fall more rapidly in pretreated animals, whereas liver copper concentrations increased more rapidly; red blood cell copper concentrations were not higher in animals given spironolactone. Pretreated animals excreted significantly more copper in the bile during the first 2 hr after 64Cu(NO3)2 injection, and had higher hepatic copper levels at 3 hr. | [
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PMID:10155 | An equine cryptorchid with testicular and ovarian tissues. | Cytogenetic and histological studies were carried out on an intersex horse which was diagnosed clinically as a cryptorchid. Surgery confirmed the horse to be a bilateral abdominal cryptorchid and histological examination revealed ovarian tissue associated with the left epididymis. Chromosome analysis of cultured cells from testicular tissue, ovarian tissue and skin revealed 64-XX and 64-XY make-up, the left gonad containing a greater preponderance of XX cells over XY cells. The external characteristics and behaviour of the horse were indistinguishable from that of a "routine" cryptorchid. Other cases of equine intersexes are reviewed and theories for the discrepancies between genetic, gonadal and phenotypic sex are discussed. | An equine cryptorchid with testicular and ovarian tissues. Cytogenetic and histological studies were carried out on an intersex horse which was diagnosed clinically as a cryptorchid. Surgery confirmed the horse to be a bilateral abdominal cryptorchid and histological examination revealed ovarian tissue associated with the left epididymis. Chromosome analysis of cultured cells from testicular tissue, ovarian tissue and skin revealed 64-XX and 64-XY make-up, the left gonad containing a greater preponderance of XX cells over XY cells. The external characteristics and behaviour of the horse were indistinguishable from that of a "routine" cryptorchid. Other cases of equine intersexes are reviewed and theories for the discrepancies between genetic, gonadal and phenotypic sex are discussed. | [
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PMID:10151 | The secretion of methadone and its major metabolite in the gastric juice of humans: comparison with blood and salivary concentrations. | Four healthy subjects and four addicts on high daily maintenance doses of methadone each received a parenteral dose of methadone hydrochloride following an overnight fast. The concentration of methadone in blood was compared with that in the gastric juice obtained over 8 hr by continuous low-pressure suction via a nasogastric tube. The concentration in the gastric juice was 25-200 times that measured at the same time in the blood. Thus, 8 hr after the injection mean blood concentrations of 28 and 210 ng of methadone per ml were recorded in the normal subjects and the addicts, respectively. The corresponding concentrations in gastric juice were 2,200 ng/ml and 18,000 ng/ml, respectively. In the normal subjects about 2% of the administered dose was recovered in the gastric juice in 8 hr, whereas in addicts about 7% was recovered. The greater recovery of methadone from the addicts appears to be the result of the larger volume of gastric juice recovered from the latter subjects. Methadone was also excreted in the saliva of both groups of subjects. In addicts, salivary concentrations were often 10 times those recorded in the blood. The N-monodemethylated metabolite of methadone was identified in the gastric juice of addicts by gas chromatography and mass spectrometry. | The secretion of methadone and its major metabolite in the gastric juice of humans: comparison with blood and salivary concentrations. Four healthy subjects and four addicts on high daily maintenance doses of methadone each received a parenteral dose of methadone hydrochloride following an overnight fast. The concentration of methadone in blood was compared with that in the gastric juice obtained over 8 hr by continuous low-pressure suction via a nasogastric tube. The concentration in the gastric juice was 25-200 times that measured at the same time in the blood. Thus, 8 hr after the injection mean blood concentrations of 28 and 210 ng of methadone per ml were recorded in the normal subjects and the addicts, respectively. The corresponding concentrations in gastric juice were 2,200 ng/ml and 18,000 ng/ml, respectively. In the normal subjects about 2% of the administered dose was recovered in the gastric juice in 8 hr, whereas in addicts about 7% was recovered. The greater recovery of methadone from the addicts appears to be the result of the larger volume of gastric juice recovered from the latter subjects. Methadone was also excreted in the saliva of both groups of subjects. In addicts, salivary concentrations were often 10 times those recorded in the blood. The N-monodemethylated metabolite of methadone was identified in the gastric juice of addicts by gas chromatography and mass spectrometry. | [
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PMID:10157 | Red cell hemoglobin, hydrogen ion and electrolyte concentrations during exercise in trained and untrained subjects. | Red cell concentrations of hemoglobin (MCHC), H+, Na+, K+, Mg++, cl- were measured in femoral venous blood of six untrained (UT), six endurance trained (TR) and three semitrained (ST) subjects during graded increasing work (4, 8, 12, 18 and 24 mkp/s, 10-15 min on each step) on a bicycle ergometer. Before exercise no significant differences were detected for the measured variables when comparing UT and TR. During exercise MCHC, [Na+], [K+] and [Mg++] remained constant indicating lack of water shift into the erythrocytes in spite of a marked acidosis (lowest pH Blood value 7.225). This lack resulted from an elevated extracellular osmolality. [H+]Ery and [Cl-]Ery maximally increased by 2.0 X 10(-8) eq/kg H2O and 10 meq/l, respectively. The change was markedly greater in UT than in TR at equal load. However, if [H+] Ery and [Cl-] Ery were related to pH of whole blood, differences between groups, almost disappeared and the ions were distributed as predictable from in vitro experiments (Fitzsimmons and Sendroy, 1961). Behaviour of H+ and Cl- may be of importance for oxygen dissociation under in vivo conditions. | Red cell hemoglobin, hydrogen ion and electrolyte concentrations during exercise in trained and untrained subjects. Red cell concentrations of hemoglobin (MCHC), H+, Na+, K+, Mg++, cl- were measured in femoral venous blood of six untrained (UT), six endurance trained (TR) and three semitrained (ST) subjects during graded increasing work (4, 8, 12, 18 and 24 mkp/s, 10-15 min on each step) on a bicycle ergometer. Before exercise no significant differences were detected for the measured variables when comparing UT and TR. During exercise MCHC, [Na+], [K+] and [Mg++] remained constant indicating lack of water shift into the erythrocytes in spite of a marked acidosis (lowest pH Blood value 7.225). This lack resulted from an elevated extracellular osmolality. [H+]Ery and [Cl-]Ery maximally increased by 2.0 X 10(-8) eq/kg H2O and 10 meq/l, respectively. The change was markedly greater in UT than in TR at equal load. However, if [H+] Ery and [Cl-] Ery were related to pH of whole blood, differences between groups, almost disappeared and the ions were distributed as predictable from in vitro experiments (Fitzsimmons and Sendroy, 1961). Behaviour of H+ and Cl- may be of importance for oxygen dissociation under in vivo conditions. | [
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PMID:10158 | Binding of carbon monoxide to alpha-hemocyanin and beta-hemocyanin from Helix pomatia. | The binding of carbon monoxide to alpha and beta-hemocyanin from the snail Helix pomatia was studied under equilibrium conditions. Homotropic interactions upon carbon monoxide binding were much weaker than upon the binding of oxygen. Heterotropic interactions (Bohr effect and calcium-ion effect), however, were just as strong as in the case of the binding of oxygen. For alpha-hemocyanin a linkage has been observed between the binding of carbon monoxide and a change in quaternary structure of the protein. | Binding of carbon monoxide to alpha-hemocyanin and beta-hemocyanin from Helix pomatia. The binding of carbon monoxide to alpha and beta-hemocyanin from the snail Helix pomatia was studied under equilibrium conditions. Homotropic interactions upon carbon monoxide binding were much weaker than upon the binding of oxygen. Heterotropic interactions (Bohr effect and calcium-ion effect), however, were just as strong as in the case of the binding of oxygen. For alpha-hemocyanin a linkage has been observed between the binding of carbon monoxide and a change in quaternary structure of the protein. | [
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PMID:10159 | Studies on the kinetic effects of adenosine-3':5'-monophosphate-dependent phosphorylation of purified pig-liver pyruvate kinase type L. | The effect of cyclic-AMP-dependent phosphorylation on the activity of isolated pig liver pyruvate kinase was studied. It was found that the major kinetic effect of the phosphorylation was to reduce the affinity for the substrate phosphoenolpyruvate, K0.5 for this substrate increasing from 0.3 to 0.9 mM upon phosphorylation. The cooperative effect with phosphoenolpyruvate was enhanced, the Hill constant nH increasing concomitantly from 1.1 to 1.5. V was unaltered. The change in activity occurred in parallel with the phosphate incorporation, except during the initial part of the reaction, when inactivation was correspondingly slower. The affinity for the second substrate ADP was unchanged, with an apparent Km of 0.3 mM at saturating concentration of phosphoenolpyruvate. Likewise, the requirement for potassium was unaffected, whereas the phosphoenzyme required a higher concentration of magnesium ions for maximal activity, compared with the control enzyme. The inhibitory effect of the phosphorylation was counteracted by positive effectors, fructose 1,6-biphosphate in micromolar concentrations completely activated the phosphoenzyme, resulting in an enzyme with properties similar to the fructose 1,6-biphosphate-activated unphosphorylated enzyme, with K0.5 for phosphoenolpyruvate about 0.025 mM and with a Hill constant of 1.1. Hydrogen ions were also effective in activating the phosphoenzyme. Thus, when pH was lowered from 8 to 6.5 the inhibition due to phosphorylation was abolished. The phosphoenzyme was sensitive to further inhibition by negative effectors such as ATP and alanine. 2 mM ATP increased K0.5 for phosphoenolpyruvate to 1.5 mM and nH to 2.3. The corresponding values with alanine were 1.3 mM and 1.9. Phosphorylation is thought to be an additional mechanism of inhibition of the enzyme under gluconeogenetic conditions. | Studies on the kinetic effects of adenosine-3':5'-monophosphate-dependent phosphorylation of purified pig-liver pyruvate kinase type L. The effect of cyclic-AMP-dependent phosphorylation on the activity of isolated pig liver pyruvate kinase was studied. It was found that the major kinetic effect of the phosphorylation was to reduce the affinity for the substrate phosphoenolpyruvate, K0.5 for this substrate increasing from 0.3 to 0.9 mM upon phosphorylation. The cooperative effect with phosphoenolpyruvate was enhanced, the Hill constant nH increasing concomitantly from 1.1 to 1.5. V was unaltered. The change in activity occurred in parallel with the phosphate incorporation, except during the initial part of the reaction, when inactivation was correspondingly slower. The affinity for the second substrate ADP was unchanged, with an apparent Km of 0.3 mM at saturating concentration of phosphoenolpyruvate. Likewise, the requirement for potassium was unaffected, whereas the phosphoenzyme required a higher concentration of magnesium ions for maximal activity, compared with the control enzyme. The inhibitory effect of the phosphorylation was counteracted by positive effectors, fructose 1,6-biphosphate in micromolar concentrations completely activated the phosphoenzyme, resulting in an enzyme with properties similar to the fructose 1,6-biphosphate-activated unphosphorylated enzyme, with K0.5 for phosphoenolpyruvate about 0.025 mM and with a Hill constant of 1.1. Hydrogen ions were also effective in activating the phosphoenzyme. Thus, when pH was lowered from 8 to 6.5 the inhibition due to phosphorylation was abolished. The phosphoenzyme was sensitive to further inhibition by negative effectors such as ATP and alanine. 2 mM ATP increased K0.5 for phosphoenolpyruvate to 1.5 mM and nH to 2.3. The corresponding values with alanine were 1.3 mM and 1.9. Phosphorylation is thought to be an additional mechanism of inhibition of the enzyme under gluconeogenetic conditions. | [
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PMID:10160 | Efficiency of oxidative phosphorylation and energy dissipation by H+ ion recycling in rat-liver mitochondrial metabolizing pyruvate. | A method was developed for the calculation of metabolic fluxes through individual enzymatic reactions of pyruvate metabolism including the citric acid cycle in rat liver mitochondrial incubated at metabolic states between state 4 and state 3. This method is based on the measurement of the specific radioactivities of the products formed from [2-14C]pyruvate. With this procedure the energy balance of mitochondria incubated in the presence of [2-14C]pyruvate, ATP, bicarbonate and phosphate at different ATP/ADP ratios in the medium was calculated. The ATP/ADP ratios were maintained at a steady state with creatine kinase plus creatine as a phosphoryl acceptor. The calculations revealed that by adding increasing concentrations of creatine up to 20 mM the energy dissipated by the mitochondria decreased but showed a local maximum at 13mM creatine. Omission of bicarbonate from the medium led to a shift of this maximum. When energy dissipation was minimal the overall P/O ratio was maximal. The amount of energy dissipated was paralleled by the magnitude of the pH gradient across the inner membrane. From these results it was concluded that the recycling of H+ ions which consists of a passive leakage of H+ ions into the matrix and an active extrusion of these ions out of this compartment, is an important energy dissipating process. The H+ ion recycling is thus one of the processes which give rise to the state 4 respiration in mitochondria. | Efficiency of oxidative phosphorylation and energy dissipation by H+ ion recycling in rat-liver mitochondrial metabolizing pyruvate. A method was developed for the calculation of metabolic fluxes through individual enzymatic reactions of pyruvate metabolism including the citric acid cycle in rat liver mitochondrial incubated at metabolic states between state 4 and state 3. This method is based on the measurement of the specific radioactivities of the products formed from [2-14C]pyruvate. With this procedure the energy balance of mitochondria incubated in the presence of [2-14C]pyruvate, ATP, bicarbonate and phosphate at different ATP/ADP ratios in the medium was calculated. The ATP/ADP ratios were maintained at a steady state with creatine kinase plus creatine as a phosphoryl acceptor. The calculations revealed that by adding increasing concentrations of creatine up to 20 mM the energy dissipated by the mitochondria decreased but showed a local maximum at 13mM creatine. Omission of bicarbonate from the medium led to a shift of this maximum. When energy dissipation was minimal the overall P/O ratio was maximal. The amount of energy dissipated was paralleled by the magnitude of the pH gradient across the inner membrane. From these results it was concluded that the recycling of H+ ions which consists of a passive leakage of H+ ions into the matrix and an active extrusion of these ions out of this compartment, is an important energy dissipating process. The H+ ion recycling is thus one of the processes which give rise to the state 4 respiration in mitochondria. | [
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PMID:10161 | Membrane-bound DD-carboxypeptidase and transpeptidase activities from Bacillus megaterium KM at pH 7. General properties, substrate specificity and inhibition by beta-lactam antibiotics. | 1. The membranes from Bacillus megaterium KM contained a DD-carboxypeptidase with optimum activity under the following conditions: pH 7; ionic strength, 1.3 M; temperature, 40 degrees C and below 20 degrees C. It did not require any divalent cation, but was inactivated by Cu2+ and Hg2+. It was stimulated by 2-mercaptoethanol and low concentrations of p-chloromercuribenzoate. 2. The membrane preparation also catalyzed a simple transpeptidation reaction using as carboxyl acceptors D-alanine or glycine. 3. The conditions for optimum activity, temperature-inactivation, temperature-dependence of the activity, carboxyl donor specificity, sensitivity to beta-lactam antibiotics, and insensitivity to potential peptide inhibitors of both enzyme activities, was identical. The DD-carboxypeptidase showed inhibition by D-alanine and Ac2-L-Lys-D-Ala. 4. The inhibition by beta-lactam antibiotic was reversible for both enzymic activities and the time-dependence for their recovery was identical. 5. The DD-carboxypeptidase was very sensitive to changes in the configuration and size of the side-chains of the C-terminal dipeptide of the substrate. Amino acid residues at the C-terminus that precluded the peptide from being a DD-carboxypeptidase substrate were not acceptors in the transpeptidation reaction. Dipeptides were not acceptors for the 'model transpeptidase'. 6. It is suggested that both activities are catalysed by the same enzyme molecule, whose physiological role is not the formation of peptide crosslinks during peptidoglycan biosynthesis. | Membrane-bound DD-carboxypeptidase and transpeptidase activities from Bacillus megaterium KM at pH 7. General properties, substrate specificity and inhibition by beta-lactam antibiotics. 1. The membranes from Bacillus megaterium KM contained a DD-carboxypeptidase with optimum activity under the following conditions: pH 7; ionic strength, 1.3 M; temperature, 40 degrees C and below 20 degrees C. It did not require any divalent cation, but was inactivated by Cu2+ and Hg2+. It was stimulated by 2-mercaptoethanol and low concentrations of p-chloromercuribenzoate. 2. The membrane preparation also catalyzed a simple transpeptidation reaction using as carboxyl acceptors D-alanine or glycine. 3. The conditions for optimum activity, temperature-inactivation, temperature-dependence of the activity, carboxyl donor specificity, sensitivity to beta-lactam antibiotics, and insensitivity to potential peptide inhibitors of both enzyme activities, was identical. The DD-carboxypeptidase showed inhibition by D-alanine and Ac2-L-Lys-D-Ala. 4. The inhibition by beta-lactam antibiotic was reversible for both enzymic activities and the time-dependence for their recovery was identical. 5. The DD-carboxypeptidase was very sensitive to changes in the configuration and size of the side-chains of the C-terminal dipeptide of the substrate. Amino acid residues at the C-terminus that precluded the peptide from being a DD-carboxypeptidase substrate were not acceptors in the transpeptidation reaction. Dipeptides were not acceptors for the 'model transpeptidase'. 6. It is suggested that both activities are catalysed by the same enzyme molecule, whose physiological role is not the formation of peptide crosslinks during peptidoglycan biosynthesis. | [
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PMID:10162 | Kinetic investigation of the staphylococcal protease-catalyzed hydrolysis of synthetic substrates. | In investigating the staphylococcal protease-catalyzed hydrolysis of N-tert-butoxycarbonyl-L-glutamate alpha-phenyl ester, N-benzyloxycarbonyl-L-glutamate alpha-phenyl ester and N-benzyloxycarbonyl-L-glutamate alpha-p-nitroanilide, we obtained kinetic evidence consistent with the formation of an acyl-enzyme intermediate. We found that addition of a nucleophile, such as methanol, led to the partition of the common acyl-enzyme intermediate between water and the alcohol. With N-benzyl-oxycarbonyl-L-glutamate alpha-phenyl ester, a specific ester substrate, deacylation was shown to be the rate-limiting step. By studying the kcat/Km ratio of these hydrolyses as a function of pH, we have shown that two ionizable groups on the enzyme are essential to the catalytic process. One of these groups has a pK of 6.58 and the other, a pK of 8.25. The assignment of these pK values is discussed in connection with the known features of the serine proteinase reaction mechanism. In addition, monovalent anions were shown to inhibit staphylococcal protease hydrolyses. They seem to compete with the negative charge of the substrate, thus inhibiting its binding on the enzyme molecule. Finally we compared the kinetic parameters obtained with five proteases isolated from different strains of Staphylococcus aureus. | Kinetic investigation of the staphylococcal protease-catalyzed hydrolysis of synthetic substrates. In investigating the staphylococcal protease-catalyzed hydrolysis of N-tert-butoxycarbonyl-L-glutamate alpha-phenyl ester, N-benzyloxycarbonyl-L-glutamate alpha-phenyl ester and N-benzyloxycarbonyl-L-glutamate alpha-p-nitroanilide, we obtained kinetic evidence consistent with the formation of an acyl-enzyme intermediate. We found that addition of a nucleophile, such as methanol, led to the partition of the common acyl-enzyme intermediate between water and the alcohol. With N-benzyl-oxycarbonyl-L-glutamate alpha-phenyl ester, a specific ester substrate, deacylation was shown to be the rate-limiting step. By studying the kcat/Km ratio of these hydrolyses as a function of pH, we have shown that two ionizable groups on the enzyme are essential to the catalytic process. One of these groups has a pK of 6.58 and the other, a pK of 8.25. The assignment of these pK values is discussed in connection with the known features of the serine proteinase reaction mechanism. In addition, monovalent anions were shown to inhibit staphylococcal protease hydrolyses. They seem to compete with the negative charge of the substrate, thus inhibiting its binding on the enzyme molecule. Finally we compared the kinetic parameters obtained with five proteases isolated from different strains of Staphylococcus aureus. | [
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PMID:10164 | Plasma and brain amino acids in fulminant hepatic failure and their relationship to hepatic encephalopathy. | Amino acid concentrations were determined in plasma, whole blood, cerebrospinal fluid and brain tissue of 45 patients with grade 3 or 4 coma due to fulminant hepatic failure. The concentration of 15 of the 19 amino acids determined were significantly increased in blood and the increases were greatest for the amino acids concerned with neurotransmitter metabolism. There was no correlation, however, between the plasma concentration of these amino acids and changes in the grade of hepatic coma. The plasma concentrations of the branched chain amino acids were normal except in those patients who subsequently recovered in whom levels were slightly decreased. Phenylalanine- tyrosine and methionine were among the 15 out of 18 amino acids which were significantly increased in cerebrospinal fluid and among the 15 out of 21 amino acids which were significantly increased in the brain. The increase in tryptophan was associated with a significant elevation in brain 5-hydroxyindoleacetic acid concentration suggesting an increase in 5-hydroxytryptamine turnover in hepatic coma. Brain to plasma ratios of most amino acids in hepatic coma patients were similar to control subjects suggesting that plasma concentration is the main factor controlling the cerebral concentration. However, for the branched chain amino acids, cerebrospinal fluid and brain concentrations were increased when plasma concentrations were normal suggesting an increase in brain uptake. | Plasma and brain amino acids in fulminant hepatic failure and their relationship to hepatic encephalopathy. Amino acid concentrations were determined in plasma, whole blood, cerebrospinal fluid and brain tissue of 45 patients with grade 3 or 4 coma due to fulminant hepatic failure. The concentration of 15 of the 19 amino acids determined were significantly increased in blood and the increases were greatest for the amino acids concerned with neurotransmitter metabolism. There was no correlation, however, between the plasma concentration of these amino acids and changes in the grade of hepatic coma. The plasma concentrations of the branched chain amino acids were normal except in those patients who subsequently recovered in whom levels were slightly decreased. Phenylalanine- tyrosine and methionine were among the 15 out of 18 amino acids which were significantly increased in cerebrospinal fluid and among the 15 out of 21 amino acids which were significantly increased in the brain. The increase in tryptophan was associated with a significant elevation in brain 5-hydroxyindoleacetic acid concentration suggesting an increase in 5-hydroxytryptamine turnover in hepatic coma. Brain to plasma ratios of most amino acids in hepatic coma patients were similar to control subjects suggesting that plasma concentration is the main factor controlling the cerebral concentration. However, for the branched chain amino acids, cerebrospinal fluid and brain concentrations were increased when plasma concentrations were normal suggesting an increase in brain uptake. | [
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PMID:10165 | Serum concentrations of methaqualone after repeated oral doses of a combination formulation to human subjects. | Concentrations of methaqualone have been measured in the serum of five male human subjects receiving five consecutive evening doses of a combination formulation containing methaqualone (250 mg), carbromal (300 mg) and benactyzine (0.33 mg) in each tablet. After administration of the first dose, mean peak serum concentrations of methaqualone (1.2 mug/ml) occurred at 3 h. After obtaining peak levels, mean concentrations of methaqualone declined rapidly during the next 6 h and thereafter more slowly during the next 18 h. After administration of the last (fifth) dose, mean peak serum concentrations of methaqualone (1.9 mug/ml; 1.5 mug/ml above the predose level) occurred at 2 h. After attaining peak levels, mean concentrations of methaqualone declined rapidly during the next 6 h, and thereafter more slowly, with a half-life of approximately 10 h. Mean concentrations of methaqualone in serum samples 24 h after the second, third, fourth or fifth doses were not significantly different (0.3 mug/ml - 0.6 mug/ml) during this period of dosing. This suggests that significant accumulation of methaqualone in the serum did not occur during a period of five consecutive evening doses of the combination formulation. | Serum concentrations of methaqualone after repeated oral doses of a combination formulation to human subjects. Concentrations of methaqualone have been measured in the serum of five male human subjects receiving five consecutive evening doses of a combination formulation containing methaqualone (250 mg), carbromal (300 mg) and benactyzine (0.33 mg) in each tablet. After administration of the first dose, mean peak serum concentrations of methaqualone (1.2 mug/ml) occurred at 3 h. After obtaining peak levels, mean concentrations of methaqualone declined rapidly during the next 6 h and thereafter more slowly during the next 18 h. After administration of the last (fifth) dose, mean peak serum concentrations of methaqualone (1.9 mug/ml; 1.5 mug/ml above the predose level) occurred at 2 h. After attaining peak levels, mean concentrations of methaqualone declined rapidly during the next 6 h, and thereafter more slowly, with a half-life of approximately 10 h. Mean concentrations of methaqualone in serum samples 24 h after the second, third, fourth or fifth doses were not significantly different (0.3 mug/ml - 0.6 mug/ml) during this period of dosing. This suggests that significant accumulation of methaqualone in the serum did not occur during a period of five consecutive evening doses of the combination formulation. | [
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PMID:10166 | The immunological properties of haptens coupled to thymus-independent carrier molecules. III. The role of the immunogenicity and mitogenicity of the carrier in the induction of primary IgM anti-hapten responses. | Hapten (DNP-lys) conjugates of two putatively nonimmunogenic polymers, hyalutonic acid and poly-gamma-D-glutamic acid, induce significant primary IgM anti-DNP responses in C3H mice. Preparations of various immunogenic (Type 3 pneumococcal polysaccharide (SIII), levan, E. coli lipopolysaccharide) and nonimmunogenic (hyaluronic acid and poly-glutamic acid) polymers were tested for their ability to act as polyclonal mitogens in vitro. In serum-containing spleen cell cultures, only lipopolysaccharide stimulated substantial cell proliferation. In serum-free medium, and using high specific activity [3H]thymidine, lipopolysaccharide, levan, SIII and to a lesser degree hyaluronic acid induced significant thymidine incorporation. However, under the latter conditions cell survival and proliferation were much less impressive. There was no apparent correlation between the capacity of various polymers to induce lymphocyte proliferation and their "potency" as carriers for the generation of a primary IgM anti-DNP response. Furthermore while low doses of lipopolysaccharide elicited "polyclonal" antibody formation in vivo, high doses of SIII, levan and hyaluronic acid did not. These results indicate that T cell-independent B cell triggering is dependent on the polymeric nature of the antigen, and that polymers need not be immunogenic or mitogenic to act as carriers for the induction of primary IgM anti-hapten antibody responses. | The immunological properties of haptens coupled to thymus-independent carrier molecules. III. The role of the immunogenicity and mitogenicity of the carrier in the induction of primary IgM anti-hapten responses. Hapten (DNP-lys) conjugates of two putatively nonimmunogenic polymers, hyalutonic acid and poly-gamma-D-glutamic acid, induce significant primary IgM anti-DNP responses in C3H mice. Preparations of various immunogenic (Type 3 pneumococcal polysaccharide (SIII), levan, E. coli lipopolysaccharide) and nonimmunogenic (hyaluronic acid and poly-glutamic acid) polymers were tested for their ability to act as polyclonal mitogens in vitro. In serum-containing spleen cell cultures, only lipopolysaccharide stimulated substantial cell proliferation. In serum-free medium, and using high specific activity [3H]thymidine, lipopolysaccharide, levan, SIII and to a lesser degree hyaluronic acid induced significant thymidine incorporation. However, under the latter conditions cell survival and proliferation were much less impressive. There was no apparent correlation between the capacity of various polymers to induce lymphocyte proliferation and their "potency" as carriers for the generation of a primary IgM anti-DNP response. Furthermore while low doses of lipopolysaccharide elicited "polyclonal" antibody formation in vivo, high doses of SIII, levan and hyaluronic acid did not. These results indicate that T cell-independent B cell triggering is dependent on the polymeric nature of the antigen, and that polymers need not be immunogenic or mitogenic to act as carriers for the induction of primary IgM anti-hapten antibody responses. | [
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PMID:10167 | B cell tolerance induced by polymeric antigens. I. Comparison of the dose and epitope density requirements for inactivation of primed and unprimed B cells in vivo. | Hapten [2,4-dinitrophenyl (DNP)]-specific tolerance was induced in nonimmune or DNP-hemocyanin (DNP-KLH) primed mice by administering hapten-conjugated type 3 pneumococcal polysaccharide (DNP-lys-S3). The dose of DNP-lys2.5-S3 required to suppress the primary anti-DNP antibody responses was approximately ten times higher than that required to suppress the secondary response. Large doses of lightly substituted antigen (DNP-lys0.6-S3) had no effect on primary antibody responses, while small doses of this conjugate suppressed 90-95% of the secondary response. The conclusion from this (presumably B cell) tolerance model is that B lymphocytes "mature" in their susceptibility to tolerization following primary contact with immunogen, since primed cells are inactivated by lower doses of tolerogen, and by tolerogen with lower epitope density, than nonimmune B cells. These and other data suggest that the tolerance threshold of B lymphocytes is related to their state of differentiation, and especially to their antigen-binding characteristics. | B cell tolerance induced by polymeric antigens. I. Comparison of the dose and epitope density requirements for inactivation of primed and unprimed B cells in vivo. Hapten [2,4-dinitrophenyl (DNP)]-specific tolerance was induced in nonimmune or DNP-hemocyanin (DNP-KLH) primed mice by administering hapten-conjugated type 3 pneumococcal polysaccharide (DNP-lys-S3). The dose of DNP-lys2.5-S3 required to suppress the primary anti-DNP antibody responses was approximately ten times higher than that required to suppress the secondary response. Large doses of lightly substituted antigen (DNP-lys0.6-S3) had no effect on primary antibody responses, while small doses of this conjugate suppressed 90-95% of the secondary response. The conclusion from this (presumably B cell) tolerance model is that B lymphocytes "mature" in their susceptibility to tolerization following primary contact with immunogen, since primed cells are inactivated by lower doses of tolerogen, and by tolerogen with lower epitope density, than nonimmune B cells. These and other data suggest that the tolerance threshold of B lymphocytes is related to their state of differentiation, and especially to their antigen-binding characteristics. | [
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PMID:10168 | B cell tolerance induced by polymeric antigens. II. Effects of tolerance on hapten-binding lymphocyte levels in primary and secondary antibody responses. | Tolerogenic doses of hapten [2,4-dinitrophenyl (DNP)]-coupled type 3 pneumococcal polysaccharide (DNP-lys2.5-S3) totally abolished the anti-DNP rosette-forming cell (RFC) response to primary immunization with DNP-hemocyanin in mice, while lightly substituted antigen (DNP-lys0.6-S3) had little effect. Both antigens suppressed secondary anti-DNP RFC responses to DNP-KLH. Limiting doses of DNP-lys-S3 preferentially suppressed antibody-secreting cell levels, and had less effect on RFC. DNP-lys2.5-S3 was 500--1000-fold more potent in "blockading" primary RFC in vitro than DNP-lys0.6-S3, whereas both antigens were equally effective in blocking secondary RFC. These results suggest that the sensitivity of primed B lymphocytes to inactivation by DNP-lys-S3 is related to their high avidity for antigen. Furthermore, this appears to be largely due to a high density of immunoglobulin receptors on primed cells since the affinities of primary and secondary RFC for monovalent hapten were indistinguishable. Treatment of primarily immunized mice with DNP-lys2.5-S3 2 h before assay abolished 90% of RFC. Therefore, the reduction in RFC levels in tolerant mice may be due to cellular blockade by persisting tolerogen. However, it seems unlikely that simple blockade of antigen-reactive cells is the sole mechanism operative in this system. | B cell tolerance induced by polymeric antigens. II. Effects of tolerance on hapten-binding lymphocyte levels in primary and secondary antibody responses. Tolerogenic doses of hapten [2,4-dinitrophenyl (DNP)]-coupled type 3 pneumococcal polysaccharide (DNP-lys2.5-S3) totally abolished the anti-DNP rosette-forming cell (RFC) response to primary immunization with DNP-hemocyanin in mice, while lightly substituted antigen (DNP-lys0.6-S3) had little effect. Both antigens suppressed secondary anti-DNP RFC responses to DNP-KLH. Limiting doses of DNP-lys-S3 preferentially suppressed antibody-secreting cell levels, and had less effect on RFC. DNP-lys2.5-S3 was 500--1000-fold more potent in "blockading" primary RFC in vitro than DNP-lys0.6-S3, whereas both antigens were equally effective in blocking secondary RFC. These results suggest that the sensitivity of primed B lymphocytes to inactivation by DNP-lys-S3 is related to their high avidity for antigen. Furthermore, this appears to be largely due to a high density of immunoglobulin receptors on primed cells since the affinities of primary and secondary RFC for monovalent hapten were indistinguishable. Treatment of primarily immunized mice with DNP-lys2.5-S3 2 h before assay abolished 90% of RFC. Therefore, the reduction in RFC levels in tolerant mice may be due to cellular blockade by persisting tolerogen. However, it seems unlikely that simple blockade of antigen-reactive cells is the sole mechanism operative in this system. | [
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PMID:10169 | Recruitment of effector lymphocytes by initiator lymphocytes. In vivo migration of in vitro sensitized initiator T lymphocytes. | We previously found that mouse T lymphocytes sensitized in vitro against allo- or syngeneic fibroblasts, upon injection into syngeneic recipients, do not themselves differentiate into effector cells, but recruit effector T lymphocytes within the draining lymph nodes. As a result of sensitization, these initiator lymphocytes acquire a trypsin-sensitive membrane property which is necessary for recruitment. We now report studies on the in vivo migratory behavior of initiator lymphocytes following sensitization. We injected 51Cr-labeled initiator lymphocytes into recipient footpads and found significantly increased migration of sensitized cells to the draining popliteal lymph node (PLN) during the first day. By amputation of the foot at various times, we showed that migration during the first 12-24 hours was critical for subsequent recruitment. Trypsin treatment of initiator lymphocytes abolished this accelerated migration. Lymphocytes triggered nonspecifically by Con A migrated to the PLN like antigen-sensitized cells. We also compared the migration of injected lymphocytes from the footpad to the PLN in graft-versus-host and host-versus-graft reactions, and found these reactions to differ both from each other and from recruitment in terms of lymphocyte migration. These findings are discussed in terms of the physiology of the cell-mediated immune response and the notion of peripheral sensitization. | Recruitment of effector lymphocytes by initiator lymphocytes. In vivo migration of in vitro sensitized initiator T lymphocytes. We previously found that mouse T lymphocytes sensitized in vitro against allo- or syngeneic fibroblasts, upon injection into syngeneic recipients, do not themselves differentiate into effector cells, but recruit effector T lymphocytes within the draining lymph nodes. As a result of sensitization, these initiator lymphocytes acquire a trypsin-sensitive membrane property which is necessary for recruitment. We now report studies on the in vivo migratory behavior of initiator lymphocytes following sensitization. We injected 51Cr-labeled initiator lymphocytes into recipient footpads and found significantly increased migration of sensitized cells to the draining popliteal lymph node (PLN) during the first day. By amputation of the foot at various times, we showed that migration during the first 12-24 hours was critical for subsequent recruitment. Trypsin treatment of initiator lymphocytes abolished this accelerated migration. Lymphocytes triggered nonspecifically by Con A migrated to the PLN like antigen-sensitized cells. We also compared the migration of injected lymphocytes from the footpad to the PLN in graft-versus-host and host-versus-graft reactions, and found these reactions to differ both from each other and from recruitment in terms of lymphocyte migration. These findings are discussed in terms of the physiology of the cell-mediated immune response and the notion of peripheral sensitization. | [
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PMID:10170 | Immunologic induction of reticulum cell sarcoma: donor-type lymphomas in the graft-versus-host model. | The aim of this study was to investigate malignant lymphomas of donor origin induced in F1 mice undergoing a chronic graft-versus-host reaction (GVHR) after injection of parental strain spleen cells. A total of 3 X 10(8) or 4 X 10(8) C57BL/10 spleen cells were administered to 7-8-week-old H-2 incompatible (C57BL/10 X HTG)F1 hybrids either as single or fractionated i.p. injections. Recipients were killed at intervals ranging from 1 month to 1 year after the first injection and their lymphoid cells typed for host-derived and donor-derived histocompatibility antigens. In 49% of the 88 GVH F1 mice, cells failed to be killed by a hyperimmune serum against HTG (H-2g) but reacted normally with antisera to C57BL/10 (H-2b) and, in the 9 cases tested, to theta-C3H. The conclusion that the lymphoid cells of these mice were derived from donor cells was supported by the finding that these animals lacked immunoglobulins bearing host-derived Iga allotype in their serum. Forty-three percent of the GVH F1 mice developed reticulum cell sarcomas (RCS), 32% revealed hyperplastic lymphoreticular tissue, and 25% showed no grossly abnormal changes. Mice with donor-type lymphoid tissues were found in all three groups; 50% of the 38 RCS detected were donor-type neoplasms. The induction of donor-type RCS during the GVHR strengthens the concept of lymphomagenesis through persistent stimulation with antigen(s). | Immunologic induction of reticulum cell sarcoma: donor-type lymphomas in the graft-versus-host model. The aim of this study was to investigate malignant lymphomas of donor origin induced in F1 mice undergoing a chronic graft-versus-host reaction (GVHR) after injection of parental strain spleen cells. A total of 3 X 10(8) or 4 X 10(8) C57BL/10 spleen cells were administered to 7-8-week-old H-2 incompatible (C57BL/10 X HTG)F1 hybrids either as single or fractionated i.p. injections. Recipients were killed at intervals ranging from 1 month to 1 year after the first injection and their lymphoid cells typed for host-derived and donor-derived histocompatibility antigens. In 49% of the 88 GVH F1 mice, cells failed to be killed by a hyperimmune serum against HTG (H-2g) but reacted normally with antisera to C57BL/10 (H-2b) and, in the 9 cases tested, to theta-C3H. The conclusion that the lymphoid cells of these mice were derived from donor cells was supported by the finding that these animals lacked immunoglobulins bearing host-derived Iga allotype in their serum. Forty-three percent of the GVH F1 mice developed reticulum cell sarcomas (RCS), 32% revealed hyperplastic lymphoreticular tissue, and 25% showed no grossly abnormal changes. Mice with donor-type lymphoid tissues were found in all three groups; 50% of the 38 RCS detected were donor-type neoplasms. The induction of donor-type RCS during the GVHR strengthens the concept of lymphomagenesis through persistent stimulation with antigen(s). | [
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PMID:10171 | Graft-versus-host reaction induced by Peyer's patches. | Parental Peyer's patches were implanted into the anterior hepatic lobe of F1 hybrid rats. Using conventional histological techniques, phenomena typical of a graft-versus-host reaction were observed, such as paravascular lymphoid infiltration in the liver, cellular depletion of the thymus-dependent areas of spleen, and diffuse reticuloendothelial cell proliferation in various lymphoid tissues. Based on these findings and other observations it is concluded that a thymus-dependent cell population is present in the Peyer's patches. | Graft-versus-host reaction induced by Peyer's patches. Parental Peyer's patches were implanted into the anterior hepatic lobe of F1 hybrid rats. Using conventional histological techniques, phenomena typical of a graft-versus-host reaction were observed, such as paravascular lymphoid infiltration in the liver, cellular depletion of the thymus-dependent areas of spleen, and diffuse reticuloendothelial cell proliferation in various lymphoid tissues. Based on these findings and other observations it is concluded that a thymus-dependent cell population is present in the Peyer's patches. | [
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PMID:10172 | Role of epitope density in the induction of immunity and tolerance with thymus-independent antigens. I. Studies with 2,4-dinitrophenyl conjugates in vitro. | The effect of different degrees of conjugation of levan, dextran, pneumococcal polysaccharide SIII and the copolymer of D-glutamic acid and D-lysine with the 2,4-dinitrophenyl determinant (DNP) on the immunogenic and tolerogenic capacity of its haptenic conjugates was investigated in vitro. A strikingly uniform effect of hapten conjugation was observed despite the marked difference in the mol. wt. and structure (branched or linear) of the carrier molecules. Regarding the anti-DNP response, low density conjugates were immunogenic but not tolerogenic (even at high doses), higher conjugates were both, depending on concentration, while very high density conjugates were only tolerogenic. These results confirm and extend earlier findings made with DNP conjugates of polymeric flagellin and indicate the probable generality of this principle. Together with parallel in vivo studies (Eur. J. Immunol. 1975. 5:541), they reaffirm the importance of epitope density in the discrimination between immunity and tolerance. | Role of epitope density in the induction of immunity and tolerance with thymus-independent antigens. I. Studies with 2,4-dinitrophenyl conjugates in vitro. The effect of different degrees of conjugation of levan, dextran, pneumococcal polysaccharide SIII and the copolymer of D-glutamic acid and D-lysine with the 2,4-dinitrophenyl determinant (DNP) on the immunogenic and tolerogenic capacity of its haptenic conjugates was investigated in vitro. A strikingly uniform effect of hapten conjugation was observed despite the marked difference in the mol. wt. and structure (branched or linear) of the carrier molecules. Regarding the anti-DNP response, low density conjugates were immunogenic but not tolerogenic (even at high doses), higher conjugates were both, depending on concentration, while very high density conjugates were only tolerogenic. These results confirm and extend earlier findings made with DNP conjugates of polymeric flagellin and indicate the probable generality of this principle. Together with parallel in vivo studies (Eur. J. Immunol. 1975. 5:541), they reaffirm the importance of epitope density in the discrimination between immunity and tolerance. | [
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PMID:10173 | Effect of antipsychotic drugs on the firing of dorsal raphe cells. I. Role of adrenergic system. | The activity of serotonergic (5HT) neurons in the dorsal raphe nucleus was inhibited by the i.v. administration of certain antipsychotic drugs (methiothepin, clozapine and thioridazine). However, other antipsychotic agents (chlorpromazine, haloperidol and pimozide) did not inhibit raphe cell firing. The inhibitory potency of these drugs on raphe activity correlates with reported central noradrenergic blocking efficacy. An alpha-adrenergic blocking agent, piperoxane, but not the beta-blocking agents, propranolol and MJ 1999, inhibited raphe activity when administered systemically. All of these drugs appear to act indirectly since they (and NE) have relatively weak or variable effects when applied microiontophoretically to raphe neurons. The depressant effects of certain antipsychotic drugs and piperoxane on 5HT neurons appears to be mediated by a cnetral adrenergic system since (1) the depression could be reversed by the catecholamine releasing agents 1- and d-amphetamine; (2) the depression could be abolished by destruction of adrenergic pathways in the CNS by chemical, mechanical, or electrothermic lesions. While a precise localization has not yet been obtained, the data suggest that these drug effects may be mediated by an adrenergic pathway ascending from the lower brainstem. | Effect of antipsychotic drugs on the firing of dorsal raphe cells. I. Role of adrenergic system. The activity of serotonergic (5HT) neurons in the dorsal raphe nucleus was inhibited by the i.v. administration of certain antipsychotic drugs (methiothepin, clozapine and thioridazine). However, other antipsychotic agents (chlorpromazine, haloperidol and pimozide) did not inhibit raphe cell firing. The inhibitory potency of these drugs on raphe activity correlates with reported central noradrenergic blocking efficacy. An alpha-adrenergic blocking agent, piperoxane, but not the beta-blocking agents, propranolol and MJ 1999, inhibited raphe activity when administered systemically. All of these drugs appear to act indirectly since they (and NE) have relatively weak or variable effects when applied microiontophoretically to raphe neurons. The depressant effects of certain antipsychotic drugs and piperoxane on 5HT neurons appears to be mediated by a cnetral adrenergic system since (1) the depression could be reversed by the catecholamine releasing agents 1- and d-amphetamine; (2) the depression could be abolished by destruction of adrenergic pathways in the CNS by chemical, mechanical, or electrothermic lesions. While a precise localization has not yet been obtained, the data suggest that these drug effects may be mediated by an adrenergic pathway ascending from the lower brainstem. | [
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PMID:10174 | Effect of antipsychotic drugs on the firing of dorsal raphe cells. II. Reversal by picrotoxin. | As reported in the preceding study, the ability of certain antipsychotic and adrenolytic agents to inhibit the spontaneous firing of serotonergic 5HT neurons in the dorsal raphe nucleus appeared to be related to adrenergic blocking efficacy. However, the interaction between adrenergic and serotonergic systems was apparently indirect. In this phase of the study we investigated the hypothesis that another transmitter system could mediate this interaction. We examined the effects of two inhibitory amino acid transmitters (GABA and glycine) for possible effects on dorsal raphe cell firing using single cell recording and microiontophoretic techniques. In addition, the ability of the GABA antagonist, picrotoxin and the glycine antagonist, strychnine to reverse the effects of the antipsychotic and alpha-blocking drugs on dorsal raphe firing was tested. Both GABA and glycine were found to inhibit raphe cell firing selectively, allowing for a possible neurotransmitter function for these amino acids within the dorsal raphe nucleus. However, picrotoxin but not strychnine was found to reverse the effects of the antipsychotic and alpha-blocking drugs on raphe firing. Based on these results, we propose that the adrenergic input may influence 5HT neurons indirectly via a GABAergic interneuron or interposed GABA neuron. | Effect of antipsychotic drugs on the firing of dorsal raphe cells. II. Reversal by picrotoxin. As reported in the preceding study, the ability of certain antipsychotic and adrenolytic agents to inhibit the spontaneous firing of serotonergic 5HT neurons in the dorsal raphe nucleus appeared to be related to adrenergic blocking efficacy. However, the interaction between adrenergic and serotonergic systems was apparently indirect. In this phase of the study we investigated the hypothesis that another transmitter system could mediate this interaction. We examined the effects of two inhibitory amino acid transmitters (GABA and glycine) for possible effects on dorsal raphe cell firing using single cell recording and microiontophoretic techniques. In addition, the ability of the GABA antagonist, picrotoxin and the glycine antagonist, strychnine to reverse the effects of the antipsychotic and alpha-blocking drugs on dorsal raphe firing was tested. Both GABA and glycine were found to inhibit raphe cell firing selectively, allowing for a possible neurotransmitter function for these amino acids within the dorsal raphe nucleus. However, picrotoxin but not strychnine was found to reverse the effects of the antipsychotic and alpha-blocking drugs on raphe firing. Based on these results, we propose that the adrenergic input may influence 5HT neurons indirectly via a GABAergic interneuron or interposed GABA neuron. | [
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PMID:10176 | Regulation of enzymes of ethanol metabolism in yeast (Rhodotorula gracilis). | The three enzymes of ethanol metabolism alcohol dehydrogenase, aldehyde dehydrogenase and acetyl-CoA synthetase in the obligate aerobic yeast Rhodotorula gracilis are repressed by glucose and induced by C2 metabolic fuels with a regulatory pattern indicating a correlation in the control mechanisms. To try an identification of the molecular signals involved in the transmission of the inducing stimulus, experiments were carried out by blocking with 2 mM pyrazole the ethanol acetaldehyde metabolic step. Results indicate that ethanol is not specifically required as a molecular signal for induction. | Regulation of enzymes of ethanol metabolism in yeast (Rhodotorula gracilis). The three enzymes of ethanol metabolism alcohol dehydrogenase, aldehyde dehydrogenase and acetyl-CoA synthetase in the obligate aerobic yeast Rhodotorula gracilis are repressed by glucose and induced by C2 metabolic fuels with a regulatory pattern indicating a correlation in the control mechanisms. To try an identification of the molecular signals involved in the transmission of the inducing stimulus, experiments were carried out by blocking with 2 mM pyrazole the ethanol acetaldehyde metabolic step. Results indicate that ethanol is not specifically required as a molecular signal for induction. | [
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PMID:10178 | Lipoperoxidation rates and drug-oxidizing enzyme activities in the liver and placenta of some mammal species during the perinatal period. | Lipoperoxidation and drug-metabolizing enzymes were measured in livers and placentas of different mammal species during the perinatal perios. In placentas and fetal livers of rat, rabbit and guinea-pig, cofactor-supported lipoperoxidation was negligible, as were the activities of drug-oxidizing enzymes. Human fetal liver contained an intact drug-oxidizing electron transport chain, and lipoperoxidation activity was accordingly abserved. It is suggested that lesions mediated by lipoperoxidation may be possible in human fetus, but they are less probable in animal fetuses. | Lipoperoxidation rates and drug-oxidizing enzyme activities in the liver and placenta of some mammal species during the perinatal period. Lipoperoxidation and drug-metabolizing enzymes were measured in livers and placentas of different mammal species during the perinatal perios. In placentas and fetal livers of rat, rabbit and guinea-pig, cofactor-supported lipoperoxidation was negligible, as were the activities of drug-oxidizing enzymes. Human fetal liver contained an intact drug-oxidizing electron transport chain, and lipoperoxidation activity was accordingly abserved. It is suggested that lesions mediated by lipoperoxidation may be possible in human fetus, but they are less probable in animal fetuses. | [
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PMID:10179 | Similarities between sodium channels in excitable membranes and in epithelia. | The inhibitory effects of the pyrazine derivative, amiloride, on sodium transport in an amphibian epithelium has been studied as a function of pH. It is concluded that the charged (guanidinium) group interacts with a negatively charged acid grouping in the membrane. Similarities between sodium channels in excitable membranes and epithelia are highlighted. | Similarities between sodium channels in excitable membranes and in epithelia. The inhibitory effects of the pyrazine derivative, amiloride, on sodium transport in an amphibian epithelium has been studied as a function of pH. It is concluded that the charged (guanidinium) group interacts with a negatively charged acid grouping in the membrane. Similarities between sodium channels in excitable membranes and epithelia are highlighted. | [
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PMID:10180 | Different types of synaptic vesicles in axons of the retractor penis muscle of the bull. | Three types of axon profiles were observed in the smooth muscle of the retractor penis and the penile artery of the bull: 1. profiles containing small granular vesicles, presumably representing adrenergic axons; 2. profiles containing small agranular vesicles, presumably representing cholinergic axons; 3. profiles containing numerous large and small granular vesicles. The third type of profile was not found in the vas deferens or the metatarsal artery. It is therefore possible that this type of profile represents the non-adrenergic, non-cholinergic inhibitory nerves, the presence of which has previously been pharmacologically indicated in these tissues. | Different types of synaptic vesicles in axons of the retractor penis muscle of the bull. Three types of axon profiles were observed in the smooth muscle of the retractor penis and the penile artery of the bull: 1. profiles containing small granular vesicles, presumably representing adrenergic axons; 2. profiles containing small agranular vesicles, presumably representing cholinergic axons; 3. profiles containing numerous large and small granular vesicles. The third type of profile was not found in the vas deferens or the metatarsal artery. It is therefore possible that this type of profile represents the non-adrenergic, non-cholinergic inhibitory nerves, the presence of which has previously been pharmacologically indicated in these tissues. | [
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PMID:10215 | Properties of renin granules isolated from rat kidney. | Renin granules from rat kidney prepared at 25 degrees C show greater stability at 25 degrees C than at 0 degrees C when incubated in ionic medium. The sum of the renin in the supernatant fluid plus that in the pellet was the same at 25 degrees C as at 0 degrees C, thus ruling out the possibility that the extra release at 0 degrees C merely represented greater stability of free renin at 0 degrees C. In common with other secretory granules, renin granules were most stable at pH 6.0 and were osmotically sensitive. In contrast to neurosecretory and chromaffin granules, renin granules were stabilized by Mg-ATP in ionic medium. This result is similar to studies by others on lysosomes. It is concluded that the renin granules membrane shares many of the properties of other granule membranes. Some of these properties (temperature and pH lability) will have to be considered in the design of future experiments on renin storage and release. | Properties of renin granules isolated from rat kidney. Renin granules from rat kidney prepared at 25 degrees C show greater stability at 25 degrees C than at 0 degrees C when incubated in ionic medium. The sum of the renin in the supernatant fluid plus that in the pellet was the same at 25 degrees C as at 0 degrees C, thus ruling out the possibility that the extra release at 0 degrees C merely represented greater stability of free renin at 0 degrees C. In common with other secretory granules, renin granules were most stable at pH 6.0 and were osmotically sensitive. In contrast to neurosecretory and chromaffin granules, renin granules were stabilized by Mg-ATP in ionic medium. This result is similar to studies by others on lysosomes. It is concluded that the renin granules membrane shares many of the properties of other granule membranes. Some of these properties (temperature and pH lability) will have to be considered in the design of future experiments on renin storage and release. | [
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PMID:10217 | A sensitive method for measuring haemoglobin in gastric contents. | A sensitive method for measuring haemoglobin in gastric contents using the rate of change of optical density of a mixture of orthotolidine and hydrogen peroxide in the presence of haemoglobin, is described. The method can measure the blood normally lost in 10 min from the gastric mucosa. | A sensitive method for measuring haemoglobin in gastric contents. A sensitive method for measuring haemoglobin in gastric contents using the rate of change of optical density of a mixture of orthotolidine and hydrogen peroxide in the presence of haemoglobin, is described. The method can measure the blood normally lost in 10 min from the gastric mucosa. | [
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PMID:10218 | [Blood gas analysis and acid-base balance in acute myocardial infarction. Personal observations (author's transl)]. | Arterial pH, pO2 and pCO2 were analysed with Astup's micromethod on one hundred and three acute myocardial infarctions (A.M.I.) without metabolic, pulmonary and renal diseases. Following the clinical picute, the patients were divided into five groups and results were clinically and statistically evaluated (mean, standard deviation, Student's test "t", correlation coefficient "r" between pO2 and pulmonary arterial diastolic pressure): --Ist group (A.M.I. without complications): only mild hypoxemia; --IInd group (A.M.I. with slight left ventricular failure): more remarkable hypoxemia and hypocapnia, often with respiratory alkalosis; --IIIrd group (A.M.I. complicated by acute pulmonary oedema): mixed acidosis and severe hypoxemia; --IVth group (A.M.I. complicated by shock): prevailing metabolic acidosis and severe hypoxemia. Acidosis shows good correlations with the clinical picture; --Vth group (A.M.I. with serious arrhythmias): mixed and profound acidosis and important hypoxemia during ventricular fibrillation and cardiac arrest. In twenty patients hypoxemia and arterial pulmonary diastolic pressure showed a significant correlation. | [Blood gas analysis and acid-base balance in acute myocardial infarction. Personal observations (author's transl)]. Arterial pH, pO2 and pCO2 were analysed with Astup's micromethod on one hundred and three acute myocardial infarctions (A.M.I.) without metabolic, pulmonary and renal diseases. Following the clinical picute, the patients were divided into five groups and results were clinically and statistically evaluated (mean, standard deviation, Student's test "t", correlation coefficient "r" between pO2 and pulmonary arterial diastolic pressure): --Ist group (A.M.I. without complications): only mild hypoxemia; --IInd group (A.M.I. with slight left ventricular failure): more remarkable hypoxemia and hypocapnia, often with respiratory alkalosis; --IIIrd group (A.M.I. complicated by acute pulmonary oedema): mixed acidosis and severe hypoxemia; --IVth group (A.M.I. complicated by shock): prevailing metabolic acidosis and severe hypoxemia. Acidosis shows good correlations with the clinical picture; --Vth group (A.M.I. with serious arrhythmias): mixed and profound acidosis and important hypoxemia during ventricular fibrillation and cardiac arrest. In twenty patients hypoxemia and arterial pulmonary diastolic pressure showed a significant correlation. | [
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PMID:10219 | Salycylamide glucuronide formation in liver disease and its change by drugs. | Salicylamide glucuronide (SAMG) in 0-6 and 6-12 hours-urine specimens was determined after oral administration of salicylamide in 7 normal volunteers (NV), in 51 cases of various liver diseases and hyperbilirubinemias, and in 19 cases after drug administration, to predict the in vivo drug metabolism in man and its change by drugs. Maximal glucuronide formation was obtained by 1.0 g of salicylamide administered to NV; thus, this dosage was used in the present study. SAMG as percent of total salicylamide, the percent of SAMG, from 0-6 hours-urine specimens was high and constant in NV (71.3 +/- 8.3 (Mean +/- S.D.)). 0-0.08% of the total salicylamide was confirmed as free salicylamide in 0-12 hours-urine specimens of NV. The percent of SAMG of 0-6 hours-urine specimens was 57.2 +/- 8.6 in acute hepatitis, 66.6 +/- 10.9 in chronic hepatitis, and 48.6 +/- 10.7 in liver cirrhosis (mean +/- S.D.). Free salicylamide increased slightly in liver diseases. Serum bilirubin levels tended to be inversely correlated with the percent of SAMG. In most cases of Gilbert's syndrome, the percent of SAMG remained at a normal level. The percent of SAMG in cases with unconjugated hyperbilirubinemias of other geneses were almost within normal limits. Bucolome and phenobarbital increased the percent of SAMG in patients with various liver diseases. After rifampicin or phenytoin administration, the percent of SAMG of the patients with lung tuberculosis or epilepsy did not surpass that of NV. | Salycylamide glucuronide formation in liver disease and its change by drugs. Salicylamide glucuronide (SAMG) in 0-6 and 6-12 hours-urine specimens was determined after oral administration of salicylamide in 7 normal volunteers (NV), in 51 cases of various liver diseases and hyperbilirubinemias, and in 19 cases after drug administration, to predict the in vivo drug metabolism in man and its change by drugs. Maximal glucuronide formation was obtained by 1.0 g of salicylamide administered to NV; thus, this dosage was used in the present study. SAMG as percent of total salicylamide, the percent of SAMG, from 0-6 hours-urine specimens was high and constant in NV (71.3 +/- 8.3 (Mean +/- S.D.)). 0-0.08% of the total salicylamide was confirmed as free salicylamide in 0-12 hours-urine specimens of NV. The percent of SAMG of 0-6 hours-urine specimens was 57.2 +/- 8.6 in acute hepatitis, 66.6 +/- 10.9 in chronic hepatitis, and 48.6 +/- 10.7 in liver cirrhosis (mean +/- S.D.). Free salicylamide increased slightly in liver diseases. Serum bilirubin levels tended to be inversely correlated with the percent of SAMG. In most cases of Gilbert's syndrome, the percent of SAMG remained at a normal level. The percent of SAMG in cases with unconjugated hyperbilirubinemias of other geneses were almost within normal limits. Bucolome and phenobarbital increased the percent of SAMG in patients with various liver diseases. After rifampicin or phenytoin administration, the percent of SAMG of the patients with lung tuberculosis or epilepsy did not surpass that of NV. | [
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PMID:10220 | Influence of drugs and chemicals upon hepatic enzymes and proteins. II. The effects of various barbiturates on the induction and reduction of hepatic cytoplasmic organic anion-binding proteins. | The effects of seven barbiturates (phenobarbital, three N-phenylbarbiturates and three N-cyclohexylbarbiturates) on the hepatic cytoplasmic organic anion-binding proteins, Y and Z, were investigated in an attempt to observe the structure-activity relationship of baributrates to induction and reduction of these two proteins. Sulfobromophthalein (BSP) was fully bound by the Y and Z proteins at ten minutes of mixing with the 10,5000 X g supernate. In low concentrations of BSP, saturation of binding of BSP by the Z protein was very low, and with increasing concentration, BSP-binding by the Z protein increased rapidly. The Y protein bound BSP sufficiently even in low concentrations of the dye. BSP-binding capacity of the Yprotein was increased by phenobarbital, phetharbital and bucolome, and decreased by one of the N-phenylbarbiturates. BSP-binding capacity of the Zprotein tended to be decreased by phenobarbital and phetharbital, but to be increased by bucolome. The other N-phenyl- and N-cyclohexylbarbiturates had no effect on the binding capacities of the two proteins. From these results it was concluded that the regulation by the barbiturates of cytoplasmic proteins is different from that of the microsomal enzymes, and that both type and structural relation are important in the induction and reduction of the Y and Z proteins. | Influence of drugs and chemicals upon hepatic enzymes and proteins. II. The effects of various barbiturates on the induction and reduction of hepatic cytoplasmic organic anion-binding proteins. The effects of seven barbiturates (phenobarbital, three N-phenylbarbiturates and three N-cyclohexylbarbiturates) on the hepatic cytoplasmic organic anion-binding proteins, Y and Z, were investigated in an attempt to observe the structure-activity relationship of baributrates to induction and reduction of these two proteins. Sulfobromophthalein (BSP) was fully bound by the Y and Z proteins at ten minutes of mixing with the 10,5000 X g supernate. In low concentrations of BSP, saturation of binding of BSP by the Z protein was very low, and with increasing concentration, BSP-binding by the Z protein increased rapidly. The Y protein bound BSP sufficiently even in low concentrations of the dye. BSP-binding capacity of the Yprotein was increased by phenobarbital, phetharbital and bucolome, and decreased by one of the N-phenylbarbiturates. BSP-binding capacity of the Zprotein tended to be decreased by phenobarbital and phetharbital, but to be increased by bucolome. The other N-phenyl- and N-cyclohexylbarbiturates had no effect on the binding capacities of the two proteins. From these results it was concluded that the regulation by the barbiturates of cytoplasmic proteins is different from that of the microsomal enzymes, and that both type and structural relation are important in the induction and reduction of the Y and Z proteins. | [
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PMID:10221 | The estimation of gastric secretory capacity by the telemetering method of pH-sensitive radiocapsule. | Using a pH-sensitive radiotelemetering capsule with antimony electrode or glass electrode, the capacity of the gastric acid secretion was determined according to a modified Satveny's method on four healthy controls and 19 patients with peptic ulcer, and it was compared with that from the usual aspiration method. As to the basal condition, acid output couldn't be measured by the telemetering method in most cases because of high pH value in the stomach. There was no definite relation between two methods. On the other hand, acid output from the telemetering after stimulation of AOC-tetragastrin (4 gamma/kg, s.c.) showed a good reproducibility and good correlation with that from the aspiration method (r = +0.86). And the difference between two methods could be explained from the fact that some volume of secreted acid escaped into the duodenum before being neutralized by potassium bicarbonate, and that some of gastric juice remained not aspirated in the stomach. | The estimation of gastric secretory capacity by the telemetering method of pH-sensitive radiocapsule. Using a pH-sensitive radiotelemetering capsule with antimony electrode or glass electrode, the capacity of the gastric acid secretion was determined according to a modified Satveny's method on four healthy controls and 19 patients with peptic ulcer, and it was compared with that from the usual aspiration method. As to the basal condition, acid output couldn't be measured by the telemetering method in most cases because of high pH value in the stomach. There was no definite relation between two methods. On the other hand, acid output from the telemetering after stimulation of AOC-tetragastrin (4 gamma/kg, s.c.) showed a good reproducibility and good correlation with that from the aspiration method (r = +0.86). And the difference between two methods could be explained from the fact that some volume of secreted acid escaped into the duodenum before being neutralized by potassium bicarbonate, and that some of gastric juice remained not aspirated in the stomach. | [
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PMID:10222 | Effect of salts on the kinetic parameters and thermal stability of bovine brain acid phosphatase. | Bovine brain acid phosphatase is inhibited, at any pH, by an increase in ionic strength. The rate decrease is associated at pH 5, with a marked decrease in Km and, at pH 8, with a noticeable decrease in Vm. The rate of thermal inactivation of the enzyme is unaffected by increasing ionic strength up to 300 mM. These results are discussed in terms of interactions at the active site of the enzyme. | Effect of salts on the kinetic parameters and thermal stability of bovine brain acid phosphatase. Bovine brain acid phosphatase is inhibited, at any pH, by an increase in ionic strength. The rate decrease is associated at pH 5, with a marked decrease in Km and, at pH 8, with a noticeable decrease in Vm. The rate of thermal inactivation of the enzyme is unaffected by increasing ionic strength up to 300 mM. These results are discussed in terms of interactions at the active site of the enzyme. | [
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PMID:10223 | A pharmacological study of the angiotensin receptor and tachyphylaxis in smooth muscle. | The interaction of angiotensin with its receptor has been studied on the basis of the tachyphylaxis shown by the rat uterus towards angiotensin II when pH and Ca2+ concentration are below physiological levels. 14C-Angiotensin binding and 45Ca2+-uptake investigations suggest tachyphylaxis to be due to increased binding at low pH and Ca2+ concentration. Studies with alkylating (affinity labeled) angiotensin derivatives containing the N-mustard chlorambucil suggest a "Charnière type" inhibition at the Ca-binding site of receptor and an irreversible inhibition at an anionic site. Angiotensin inhibitors containing chlorambucil do not alkylate tissue but are competitive inhibitors suggesting that the aromatic side chain in angiotensin may induce conformational changes in the receptor. The results obtained lead to a logical model for the angiotensin receptor allowing for normal activation by the hormone as well as for production of tachyphylaxis. | A pharmacological study of the angiotensin receptor and tachyphylaxis in smooth muscle. The interaction of angiotensin with its receptor has been studied on the basis of the tachyphylaxis shown by the rat uterus towards angiotensin II when pH and Ca2+ concentration are below physiological levels. 14C-Angiotensin binding and 45Ca2+-uptake investigations suggest tachyphylaxis to be due to increased binding at low pH and Ca2+ concentration. Studies with alkylating (affinity labeled) angiotensin derivatives containing the N-mustard chlorambucil suggest a "Charnière type" inhibition at the Ca-binding site of receptor and an irreversible inhibition at an anionic site. Angiotensin inhibitors containing chlorambucil do not alkylate tissue but are competitive inhibitors suggesting that the aromatic side chain in angiotensin may induce conformational changes in the receptor. The results obtained lead to a logical model for the angiotensin receptor allowing for normal activation by the hormone as well as for production of tachyphylaxis. | [
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PMID:10224 | 5-Hydroxytryptamine and narcotic-analgesics interactions in the intestine. | The effects of 5-hydroxytriptamine (5-HT), 5-HT blocking agents, morphine, narcotic-antagonists and ganglionic blocking agents were tested in the dog intestine by close intra-arterial injection. Morphine, as 5-HT, induced an immediate increase in the intestinal tonus followed by phasic contractions. When 5-HT was injected immediately after cessation of morphine induced phasic contractions, a significant potentiation of the 5-HT induced intestinal contraction could be observed. The potentiation could be demonstrated for other narcotic-analgesics like dextromoramide and it is specific for 5-HT. 5-HT blocking agents like LSD, BOL, and cyproheptadine did not block either 5-HT or morphine. However, bufotenidine, a neural tryptaminergic blocking agent, blocked the effects of both 5-HT and morphine. The effects of 5-HT and morphine upon intestinal motility were also diminished by ganglionic depolarizing agents such as nicotine and DMPP. This effect could, however, be prevented by the pretreatment with hexamethonium. These results seem to confirm the hypothesis of a 5-HT mediator role in the intestinal contractile effect induced by the narcotic-analgesics. On the other hand, narcotic-antagonists such as nalorphine and cyclazocine, not only lacked the 5-HT potentiation effect but also prevented the 5-HT potentiation induced by morphine. Cyclazocine also showed a long lasting 5-HT blocking effect. These results seem to show that the 5-HT potentiating effect of morphine in vivo is very specific and characteristic of the narcotics and thus could be implicated in some of their central effects. | 5-Hydroxytryptamine and narcotic-analgesics interactions in the intestine. The effects of 5-hydroxytriptamine (5-HT), 5-HT blocking agents, morphine, narcotic-antagonists and ganglionic blocking agents were tested in the dog intestine by close intra-arterial injection. Morphine, as 5-HT, induced an immediate increase in the intestinal tonus followed by phasic contractions. When 5-HT was injected immediately after cessation of morphine induced phasic contractions, a significant potentiation of the 5-HT induced intestinal contraction could be observed. The potentiation could be demonstrated for other narcotic-analgesics like dextromoramide and it is specific for 5-HT. 5-HT blocking agents like LSD, BOL, and cyproheptadine did not block either 5-HT or morphine. However, bufotenidine, a neural tryptaminergic blocking agent, blocked the effects of both 5-HT and morphine. The effects of 5-HT and morphine upon intestinal motility were also diminished by ganglionic depolarizing agents such as nicotine and DMPP. This effect could, however, be prevented by the pretreatment with hexamethonium. These results seem to confirm the hypothesis of a 5-HT mediator role in the intestinal contractile effect induced by the narcotic-analgesics. On the other hand, narcotic-antagonists such as nalorphine and cyclazocine, not only lacked the 5-HT potentiation effect but also prevented the 5-HT potentiation induced by morphine. Cyclazocine also showed a long lasting 5-HT blocking effect. These results seem to show that the 5-HT potentiating effect of morphine in vivo is very specific and characteristic of the narcotics and thus could be implicated in some of their central effects. | [
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PMID:10225 | The effect of cold storage on the sensitivity to alpha and beta agonists in the isolated rabbit kidney. | Rabbit kidneys were perfused at 25 degrees C, and the effect of alpha and beta agonists was studied, before and after 24 hr of cold storage. Vascular and in vitro functional parameters were evaluated. We concluded that cold storage impairs partially the vascular alpha receptor. We could not find beta receptors under these conditions. An electron microscope analysis of these kidneys has shown some degree of autolysis of the tubular cells and good preservation of the vascular smooth muscle cells and membranes. These results are very important for kidney preservation before transplantation. | The effect of cold storage on the sensitivity to alpha and beta agonists in the isolated rabbit kidney. Rabbit kidneys were perfused at 25 degrees C, and the effect of alpha and beta agonists was studied, before and after 24 hr of cold storage. Vascular and in vitro functional parameters were evaluated. We concluded that cold storage impairs partially the vascular alpha receptor. We could not find beta receptors under these conditions. An electron microscope analysis of these kidneys has shown some degree of autolysis of the tubular cells and good preservation of the vascular smooth muscle cells and membranes. These results are very important for kidney preservation before transplantation. | [
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