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PMID:14767 | The effect of a single dose of reserpine administered prior to incubation on the development of tyrosine hydroxylase activity in chick sympathetic ganglia. | A single dose of reserpine administered into the yolk sac of chicken eggs prior to incubation produces two distinct periods of significant increase in tyrosine hydroxylase (TH) activity over controls. The first period is 21 days of incubation (55%) and the second is between day 14 and 30 after hatching (a.h.) (69%). Cholineacetyltransferase (ChAc) and dopadecarboxylase (DDC) are not modified in the two periods of increased TH activity. Reserpine had no effect on cholinergic parasympathetic synapses and neurons in the ciliary ganglion, as judged by ChAc activity. When reserpine was acutely administered in three different posthatching periods only the injection at the latest period (days 26 and 27) caused a significant (38%) increase in TH activity at day 30. Postsynaptic nicotinic receptors were blocked selectively by injecting chlorisondamine in the chick starting at hatching for one week. The administration of chlorisondamine almost completely abolished the reserpine induced increase of TH activity at day 15 a.h. The present results support the view that the development of enzyme activities specifically related to neurotransmitter biosynthesis in chick autonomic ganglia is regulated not only by transsynaptic influences but also by regulatory inputs originating in the periphery. | The effect of a single dose of reserpine administered prior to incubation on the development of tyrosine hydroxylase activity in chick sympathetic ganglia. A single dose of reserpine administered into the yolk sac of chicken eggs prior to incubation produces two distinct periods of significant increase in tyrosine hydroxylase (TH) activity over controls. The first period is 21 days of incubation (55%) and the second is between day 14 and 30 after hatching (a.h.) (69%). Cholineacetyltransferase (ChAc) and dopadecarboxylase (DDC) are not modified in the two periods of increased TH activity. Reserpine had no effect on cholinergic parasympathetic synapses and neurons in the ciliary ganglion, as judged by ChAc activity. When reserpine was acutely administered in three different posthatching periods only the injection at the latest period (days 26 and 27) caused a significant (38%) increase in TH activity at day 30. Postsynaptic nicotinic receptors were blocked selectively by injecting chlorisondamine in the chick starting at hatching for one week. The administration of chlorisondamine almost completely abolished the reserpine induced increase of TH activity at day 15 a.h. The present results support the view that the development of enzyme activities specifically related to neurotransmitter biosynthesis in chick autonomic ganglia is regulated not only by transsynaptic influences but also by regulatory inputs originating in the periphery. | [
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PMID:14768 | Responsiveness of neurones in the hippocampal region of anaesthetised and unanaesthetised cats to stimulation of sensory pathways. | The responses to sensory stimuli and to shocks to the optic pathways have been studied for 399 units in the hippocampal region of 19 cats. 172 units were recorded from anaesthetised cats and 229 from 7 unanaesthetised cats. In the unanaesthetised animals the proportions of units responding were high (60%) and did not differ significantly between regions for each type of stimulation investigated: visual, auditory and optic chiasma shocks. The proportion of units responsive to chiasmatic stimulation was related to the background firing rate. The latencies and duration of the responses were relatively long and variable. Both increases and decreases in responsiveness occurred. In a number of units response were erratic, sometimes being brisk and at other times totally absent. Responses during one series of stimuli were not always similar to those during a subsequent series. A higher proportion of units responded decrementally to a series of sensory stimuli than to a series of shocks to the optic chiasma. Movement by the animal was wihtout consistent effect on unit activity. Responses were markedly influenced according to whether or not the cat looked intently at an object. An important determinant of HR unit responses was the state of arousal, the effect being regionally distributed. In anaesthetised animals the proportions of responding units were substantially lower and the responses were weaker than those in the unanaesthetised cats. | Responsiveness of neurones in the hippocampal region of anaesthetised and unanaesthetised cats to stimulation of sensory pathways. The responses to sensory stimuli and to shocks to the optic pathways have been studied for 399 units in the hippocampal region of 19 cats. 172 units were recorded from anaesthetised cats and 229 from 7 unanaesthetised cats. In the unanaesthetised animals the proportions of units responding were high (60%) and did not differ significantly between regions for each type of stimulation investigated: visual, auditory and optic chiasma shocks. The proportion of units responsive to chiasmatic stimulation was related to the background firing rate. The latencies and duration of the responses were relatively long and variable. Both increases and decreases in responsiveness occurred. In a number of units response were erratic, sometimes being brisk and at other times totally absent. Responses during one series of stimuli were not always similar to those during a subsequent series. A higher proportion of units responded decrementally to a series of sensory stimuli than to a series of shocks to the optic chiasma. Movement by the animal was wihtout consistent effect on unit activity. Responses were markedly influenced according to whether or not the cat looked intently at an object. An important determinant of HR unit responses was the state of arousal, the effect being regionally distributed. In anaesthetised animals the proportions of responding units were substantially lower and the responses were weaker than those in the unanaesthetised cats. | [
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|
PMID:14769 | Target organ regulation of sympathetic neuron development. | The role of target organs in the morphological and biochemical development of sympathetic neurons was examined in the neonatal rat. The superior cervical ganglion (SCG) and its end organs, the salivary glands and iris were employed as a model system. Unilateral sialectomy and iridectomy prevented the normal developmental increase in ipsilateral ganglion tyrosine hydroxylase (T-OH) activity, a marker for adrenergic maturation. Enzyme activity remained depressed by approximately 30% for at least 6 months, the longest time tested. Ganglion morphometry was performed to investigate the basis of the abnormal biochemical ontogeny. Target organ removal significantly decreased the number of adrenergic neurons in the Scg by approximately 30%. Total ganglion volume was reduced in a parallel fashion. Thus, end organ extirpation may prevent the biochemical maturation of the SCG by decreasing adrenergic neuron survival. Sialectomy without iridectomy prevented the normal postnatal increase in ganglion T-OH activity, but did not alter iris activity. These observations suggest that target removal prevents the development of only those neurons destined to innervate that organ. In addition to preventing normal adrenergic neuron ontogeny, target extirpation also prevented the normal development of presynaptic choline acetyltransferase activity. Presynaptic ganglion terminal may have failed to mature normally secondary to adrenergic destruction, or may have responded in some other manner to target organ extirpation. | Target organ regulation of sympathetic neuron development. The role of target organs in the morphological and biochemical development of sympathetic neurons was examined in the neonatal rat. The superior cervical ganglion (SCG) and its end organs, the salivary glands and iris were employed as a model system. Unilateral sialectomy and iridectomy prevented the normal developmental increase in ipsilateral ganglion tyrosine hydroxylase (T-OH) activity, a marker for adrenergic maturation. Enzyme activity remained depressed by approximately 30% for at least 6 months, the longest time tested. Ganglion morphometry was performed to investigate the basis of the abnormal biochemical ontogeny. Target organ removal significantly decreased the number of adrenergic neurons in the Scg by approximately 30%. Total ganglion volume was reduced in a parallel fashion. Thus, end organ extirpation may prevent the biochemical maturation of the SCG by decreasing adrenergic neuron survival. Sialectomy without iridectomy prevented the normal postnatal increase in ganglion T-OH activity, but did not alter iris activity. These observations suggest that target removal prevents the development of only those neurons destined to innervate that organ. In addition to preventing normal adrenergic neuron ontogeny, target extirpation also prevented the normal development of presynaptic choline acetyltransferase activity. Presynaptic ganglion terminal may have failed to mature normally secondary to adrenergic destruction, or may have responded in some other manner to target organ extirpation. | [
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|
PMID:14771 | A comparative study of alkaline phosphatase in calcifying cartilage, odontoblasts and the enamel organ. | The enzyme alkaline phosphatase (AP) (EC 3.1.3.1) in three different calcification areas was studied by means of a spectrophotometric micro method using p-nitrophenylphosphate as a substrate. Rat maxillary incisor odontoblasts and enamel organ from the zones of matrix formation and maturation and tissue from rabbit metatarsal cartilage were allowed to react with the substrate in glycine-NaOH buffer at room temperature. The reaction was found to be linear for a minimum of 20 min. The pH optima for AP from these tissues were in the pH range of 10.0-10.3. In order to compare AP from the four calcification areas different parameters were studied. Heating at 56 degrees C or 60 degrees C for varying times revealed that the enzymes were almost completely inactivated after 10 min. Mg2+ ions activated the enzymes by about 25% at concentrations of 2.5 mM (enamel organ 1.25 mM); while only higher concentrations of Mg2+ had an inactivating effect, Ca2+ and PO3-4 ions were inactivating at varying concentrations. F- ions showed no effect on AP activity at concentrations below 250 mM (enamel organ 125 mM) but caused inactivation of the enzymes at about 50% at 1 M. EDTA was found to be a very effective AP inactivator at concentrations above 0.06 mM, whereas urea did not noticeably affect the enzyme reactions at concentrations below 1 M. At higher concentrations, inactivation was observed. In order to determine AP localization in the epiphyseal plate successive 40-mum-thick, freeze-sectioned slices were analyzed. The activity was highest nearest the zone of cartilage calcification and decreased towards the reserve cell zone. It was concluded that the same AP isoenzyme is present in these quite different calcification loci. | A comparative study of alkaline phosphatase in calcifying cartilage, odontoblasts and the enamel organ. The enzyme alkaline phosphatase (AP) (EC 3.1.3.1) in three different calcification areas was studied by means of a spectrophotometric micro method using p-nitrophenylphosphate as a substrate. Rat maxillary incisor odontoblasts and enamel organ from the zones of matrix formation and maturation and tissue from rabbit metatarsal cartilage were allowed to react with the substrate in glycine-NaOH buffer at room temperature. The reaction was found to be linear for a minimum of 20 min. The pH optima for AP from these tissues were in the pH range of 10.0-10.3. In order to compare AP from the four calcification areas different parameters were studied. Heating at 56 degrees C or 60 degrees C for varying times revealed that the enzymes were almost completely inactivated after 10 min. Mg2+ ions activated the enzymes by about 25% at concentrations of 2.5 mM (enamel organ 1.25 mM); while only higher concentrations of Mg2+ had an inactivating effect, Ca2+ and PO3-4 ions were inactivating at varying concentrations. F- ions showed no effect on AP activity at concentrations below 250 mM (enamel organ 125 mM) but caused inactivation of the enzymes at about 50% at 1 M. EDTA was found to be a very effective AP inactivator at concentrations above 0.06 mM, whereas urea did not noticeably affect the enzyme reactions at concentrations below 1 M. At higher concentrations, inactivation was observed. In order to determine AP localization in the epiphyseal plate successive 40-mum-thick, freeze-sectioned slices were analyzed. The activity was highest nearest the zone of cartilage calcification and decreased towards the reserve cell zone. It was concluded that the same AP isoenzyme is present in these quite different calcification loci. | [
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|
PMID:14772 | Buprenorphine hydrochloride: determination of analgesic potency. | An open evaluation of relief from severe pain following major abdominal operations was carried out on at least ten patients, who had given written consent, with 0.1 to 0.4 mg doses of buprenorphine hydrochloride administered intramuscularly. Statistical analysis of the data showed that 0.3 mg of this compound provided quite satisfactory relief from pain for up to six hours. Seven more consenting patients were given buprenorphine hydrochloride 0.5 to 0.6 mg, but they did not receive much greater or longer pain relief than those receiving 0.3 to 0.4 mg. However, the latter patients were younger and heavier. It was concluded that buprenorphine hydrochloride 0.2 to 0.4 mg provided relief of severe pain probably as well as is observed with morphine 10 mg for the average-size patient, but the duration of pain relief with the new compound is substantially longer than with other strong analgesics previously tested. The only common side effect noted was drowsiness, which was observed during the analgesic action of the compound. No appreciable alterations were seen in the respiration, pulse rate and blood pressure. On the basis of these tests, buprenorphine hydrochloride appears to be a satisfactory analgesic for severe postoperative pain, and it deserves extensive study. | Buprenorphine hydrochloride: determination of analgesic potency. An open evaluation of relief from severe pain following major abdominal operations was carried out on at least ten patients, who had given written consent, with 0.1 to 0.4 mg doses of buprenorphine hydrochloride administered intramuscularly. Statistical analysis of the data showed that 0.3 mg of this compound provided quite satisfactory relief from pain for up to six hours. Seven more consenting patients were given buprenorphine hydrochloride 0.5 to 0.6 mg, but they did not receive much greater or longer pain relief than those receiving 0.3 to 0.4 mg. However, the latter patients were younger and heavier. It was concluded that buprenorphine hydrochloride 0.2 to 0.4 mg provided relief of severe pain probably as well as is observed with morphine 10 mg for the average-size patient, but the duration of pain relief with the new compound is substantially longer than with other strong analgesics previously tested. The only common side effect noted was drowsiness, which was observed during the analgesic action of the compound. No appreciable alterations were seen in the respiration, pulse rate and blood pressure. On the basis of these tests, buprenorphine hydrochloride appears to be a satisfactory analgesic for severe postoperative pain, and it deserves extensive study. | [
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|
PMID:14773 | Rapid intubation with fazadinium and suxamethonium. | Fazadinium at two dose levels (1 mg/kg and 1.5 mg/kg) and suxamethonium at three dose levels (50 mg, 75 mg and 100 mg) were investigated in 106 adult patients to determine the time interval from injection to tracheal intubation. The intubating conditions were graded according to the scheme described by Lund and Stovner. Suxamethonium 100 mg gave the shortest time interval between the end of injection and intubation. There was no significant difference between the intubation time when smaller doses of suxamethonium (50 mg and 75 mg were used and those when AH8165 (1 mg and 1.5 mg/kg) were given. Suxamethonium 100 mg also produced a significantly higher incidence of excellent intubating conditions. The clinical implications of the findings are discussed. | Rapid intubation with fazadinium and suxamethonium. Fazadinium at two dose levels (1 mg/kg and 1.5 mg/kg) and suxamethonium at three dose levels (50 mg, 75 mg and 100 mg) were investigated in 106 adult patients to determine the time interval from injection to tracheal intubation. The intubating conditions were graded according to the scheme described by Lund and Stovner. Suxamethonium 100 mg gave the shortest time interval between the end of injection and intubation. There was no significant difference between the intubation time when smaller doses of suxamethonium (50 mg and 75 mg were used and those when AH8165 (1 mg and 1.5 mg/kg) were given. Suxamethonium 100 mg also produced a significantly higher incidence of excellent intubating conditions. The clinical implications of the findings are discussed. | [
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|
PMID:14774 | Characterization of the interaction between chymotrypsin and heparin. | Heparin forms a complex with chymotrypsin which is active towards glutaryl-L-phenylalanine-p-nitroanilide (GPANA) and glutaryl-L-phenylalanine-beta-naphthylamide (GPNA) at pH 7.6. The activity of chymotrypsin towards GPANA at pH 7.6 is enhanced in the presence of heparin. Heparin does not bind at the active site of the enzyme since proflavin is not displaced from the active site of chymotrypsin upon complex formation. The heparin-chymotrypsin complex migrates under basic polyacrylamide disc gel electrophoresis conditions to a position intermediate between heparin and free chymotrypsin. The complex is dissociable under acidic polyacrylamide gel electrophoresis conditions. It is estimated that one to three molecules of heparin can bind to each chymotrypsin molecule on the basis of electrophoretic and enzymic activity data. | Characterization of the interaction between chymotrypsin and heparin. Heparin forms a complex with chymotrypsin which is active towards glutaryl-L-phenylalanine-p-nitroanilide (GPANA) and glutaryl-L-phenylalanine-beta-naphthylamide (GPNA) at pH 7.6. The activity of chymotrypsin towards GPANA at pH 7.6 is enhanced in the presence of heparin. Heparin does not bind at the active site of the enzyme since proflavin is not displaced from the active site of chymotrypsin upon complex formation. The heparin-chymotrypsin complex migrates under basic polyacrylamide disc gel electrophoresis conditions to a position intermediate between heparin and free chymotrypsin. The complex is dissociable under acidic polyacrylamide gel electrophoresis conditions. It is estimated that one to three molecules of heparin can bind to each chymotrypsin molecule on the basis of electrophoretic and enzymic activity data. | [
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PMID:14775 | Isolation and properties of a ferredoxin from leaves of Sambucus racemosa L. | Ferredoxin was isolated in good yield from leaves of Sambucus racemosa L. by the following procedure: (1) homogenization in buffered acetone-water (1:1v/v); (2) ion-exchange chromatography on several columns of DEAE-cellulose; and (3) purification by gel filtration with Sephadex G-75. The ultraviolet and visible spectrum showed maxima at 277, 331, 423, and 466 nm. The electron paramagnetic resonance spectrum was centered around g = 1.957. The protein sustained an initial photoreduction rate of 86 mumol NADP per milligram chlorophyll per hour. The amino acid composition was found to be Lys 5, His 2, Arg1, Asx11, Thr5, Ser7, Glx17, Pro6, Gly7, Ala6-7, Cys4, Val8, Ile5, Leu7, Tyr3, Phe2, and Trp1. The molecule had a molecular weight of 10 700 and contained two atoms of iron. The amino-terminal residue was alanine. These properties are highly similar to those of other angiosperm ferredoxins. Sambucus ferredoxin was found to be most closely related to that of Leucaena. | Isolation and properties of a ferredoxin from leaves of Sambucus racemosa L. Ferredoxin was isolated in good yield from leaves of Sambucus racemosa L. by the following procedure: (1) homogenization in buffered acetone-water (1:1v/v); (2) ion-exchange chromatography on several columns of DEAE-cellulose; and (3) purification by gel filtration with Sephadex G-75. The ultraviolet and visible spectrum showed maxima at 277, 331, 423, and 466 nm. The electron paramagnetic resonance spectrum was centered around g = 1.957. The protein sustained an initial photoreduction rate of 86 mumol NADP per milligram chlorophyll per hour. The amino acid composition was found to be Lys 5, His 2, Arg1, Asx11, Thr5, Ser7, Glx17, Pro6, Gly7, Ala6-7, Cys4, Val8, Ile5, Leu7, Tyr3, Phe2, and Trp1. The molecule had a molecular weight of 10 700 and contained two atoms of iron. The amino-terminal residue was alanine. These properties are highly similar to those of other angiosperm ferredoxins. Sambucus ferredoxin was found to be most closely related to that of Leucaena. | [
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|
PMID:14776 | NADPH-dependent lipid peroxidation and its effects on aminopyrine N-demethylation in subcellular fractions of human neonatal liver. | NADPH-dependent lipid peroxidation was determined in humans, using subcellular fractions of livers obtained from newborn infants. As reported for other species, activity was concentrated in the microsomal fraction and was similar to that found in the rat. High activity of lipid peroxidation induced by iron decreased aminopyrine N-demethylation and slightly reduced linearity time for the reaction. Compared with the rat, however, human microsomes were more resistant to the effects of lipid peroxidation. If liped peroxidation occurs in vivo it is unlikely to affect drug oxidation to any great degree in human infants. | NADPH-dependent lipid peroxidation and its effects on aminopyrine N-demethylation in subcellular fractions of human neonatal liver. NADPH-dependent lipid peroxidation was determined in humans, using subcellular fractions of livers obtained from newborn infants. As reported for other species, activity was concentrated in the microsomal fraction and was similar to that found in the rat. High activity of lipid peroxidation induced by iron decreased aminopyrine N-demethylation and slightly reduced linearity time for the reaction. Compared with the rat, however, human microsomes were more resistant to the effects of lipid peroxidation. If liped peroxidation occurs in vivo it is unlikely to affect drug oxidation to any great degree in human infants. | [
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|
PMID:14777 | Flow rates of components in digesta of pigs prepared with re-entrant cannulas in the proximal duodenum and terminal ileum, and fed semipurified, hard wheat, and soft wheat diets. | Four pigs prepared with re-entrant cannulas in the proximal duodenum and terminal ileum were used to study flow rates of total digesta, insoluble dry matter, nitrogen, and amino acids entering and leaving the small intestine. The pigs received a semipurified diet, a hard wheat diet, or a soft wheat diet. These were approximately isonitrogenous. A higher rate of passage of digesta through the proximal duodenum and terminal ileum were measured in pigs receiving the hard wheat diet. Peak flow of digesta at the duodenum of all pigs occurred at 1 h post feeding. Peak flow of digesta at the ileum occurred at 9 h post feeding on the soft wheat diet, but somewhat earlier on the hard wheat and semipurified diet. More nitrogen and essential amino acids flowed in the solid fraction of duodenal digesta during the first 2 h post feeding for the wheat diets and 4 h post feeding for the semipurified diet. It was concluded that flow rate of most nutrients from the stomach and through the small intestine of pigs is modified by the composition and texture of the food ingested. It is postulated that efficiency of mixing of digesta with digestive secretions in the stomach is a major factor influencing rate of flow. | Flow rates of components in digesta of pigs prepared with re-entrant cannulas in the proximal duodenum and terminal ileum, and fed semipurified, hard wheat, and soft wheat diets. Four pigs prepared with re-entrant cannulas in the proximal duodenum and terminal ileum were used to study flow rates of total digesta, insoluble dry matter, nitrogen, and amino acids entering and leaving the small intestine. The pigs received a semipurified diet, a hard wheat diet, or a soft wheat diet. These were approximately isonitrogenous. A higher rate of passage of digesta through the proximal duodenum and terminal ileum were measured in pigs receiving the hard wheat diet. Peak flow of digesta at the duodenum of all pigs occurred at 1 h post feeding. Peak flow of digesta at the ileum occurred at 9 h post feeding on the soft wheat diet, but somewhat earlier on the hard wheat and semipurified diet. More nitrogen and essential amino acids flowed in the solid fraction of duodenal digesta during the first 2 h post feeding for the wheat diets and 4 h post feeding for the semipurified diet. It was concluded that flow rate of most nutrients from the stomach and through the small intestine of pigs is modified by the composition and texture of the food ingested. It is postulated that efficiency of mixing of digesta with digestive secretions in the stomach is a major factor influencing rate of flow. | [
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|
PMID:14778 | Evoked release from guinea pig cerebral cortex slices of endogenous 14C-labelled amino acids, labelled via D-[U-14C]glucose. | Release of endogenous amino acids labelled via D-[U-14C]glucose was compared with that of several exogenous labelled amino acids using slices of guinea pig cerebral cortex. Electrical field stimulation evoked a selective release of endogenous [14C]glutamate, [14C]aspartate, and gamma-amino[14C]butyrate (14C-labelled GABA). The selectivity of release correlated well with 14C incorporation into endogenous amino acids. Calculations of the fraction of the tissue radioactivity released indicated that the selectivity was not an artifact due to differential incorporation. Because glucose in mammalian brain is metabolized almost entirely by the so-called 'large compartment', it is tentatively concluded that the releasable 'transmitter pool' of glutamate, aspartate, and GABA is located in this 'large compartment'. | Evoked release from guinea pig cerebral cortex slices of endogenous 14C-labelled amino acids, labelled via D-[U-14C]glucose. Release of endogenous amino acids labelled via D-[U-14C]glucose was compared with that of several exogenous labelled amino acids using slices of guinea pig cerebral cortex. Electrical field stimulation evoked a selective release of endogenous [14C]glutamate, [14C]aspartate, and gamma-amino[14C]butyrate (14C-labelled GABA). The selectivity of release correlated well with 14C incorporation into endogenous amino acids. Calculations of the fraction of the tissue radioactivity released indicated that the selectivity was not an artifact due to differential incorporation. Because glucose in mammalian brain is metabolized almost entirely by the so-called 'large compartment', it is tentatively concluded that the releasable 'transmitter pool' of glutamate, aspartate, and GABA is located in this 'large compartment'. | [
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|
PMID:14779 | Ca2+-dependence of the evoked release from guinea pig cerebral cortex slices of endogenous 14C-labelled amino acids, labelled via D-[U-14C]glucose. | Spontaneous and electrically evoked release of exogenous labelled amino acids and endogenous amino acids labelled from D-[U-14C]glucose were compared in control and Ca2+-free medium using guinea pig cerebral cortex slices. Spontaneous release of all labelled amino acids, except that of endogenous 14C-labelled threonine-serine-glutamine (unseparated) and exogenous [14C]aspartate, was doubled in Ca2+-free medium. The major portion of the electrically evoked release of endogenous [14C]glutamate, [14C]aspartate, gamma-amino[14C]butyrate (14C-labelled GABA) and exogenous 3H-labelled GABA was Ca2+-inpendent. More than half of the evoked release of the other labelled amino acids was Ca2+-independent. As the pattern of Ca2+-dependence of the evoked release concurred with the selectivity of the evoked release for endogenous [14C]-glutamate, [14C]aspartate, and 14C-labelled GABA, it was concluded that these labelled amino acids were probably released from the amino acid 'transmitter pool'. | Ca2+-dependence of the evoked release from guinea pig cerebral cortex slices of endogenous 14C-labelled amino acids, labelled via D-[U-14C]glucose. Spontaneous and electrically evoked release of exogenous labelled amino acids and endogenous amino acids labelled from D-[U-14C]glucose were compared in control and Ca2+-free medium using guinea pig cerebral cortex slices. Spontaneous release of all labelled amino acids, except that of endogenous 14C-labelled threonine-serine-glutamine (unseparated) and exogenous [14C]aspartate, was doubled in Ca2+-free medium. The major portion of the electrically evoked release of endogenous [14C]glutamate, [14C]aspartate, gamma-amino[14C]butyrate (14C-labelled GABA) and exogenous 3H-labelled GABA was Ca2+-inpendent. More than half of the evoked release of the other labelled amino acids was Ca2+-independent. As the pattern of Ca2+-dependence of the evoked release concurred with the selectivity of the evoked release for endogenous [14C]-glutamate, [14C]aspartate, and 14C-labelled GABA, it was concluded that these labelled amino acids were probably released from the amino acid 'transmitter pool'. | [
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|
PMID:14780 | Effects of saline infusion and acute metabolic acidosis and alkalosis on water and electrolyte transport in the human colon. | Both the kidney and colon secrete bicarbonate and transport water and electrolytes. The respective contributions of these two organs to acid-base and electrolyte balance in normal man has thus been studied in eight healthy male volunteers who underwent simultaneous renal clearance studies, and colonic perfusion with a 0.9% saline or 7.2% mannitol solution, during metabolic alkalosis and acidosis, extracellular volume expansion, and control conditions. There was no influence of these acid-base conditions on electrolyte transport in the colon. In the urine, preferential loss of chloride over sodium averaged 81, 143 (P less than 0.001), and 141 (P less than 0.05) muequiv./min, during control, metabolic acidosis, and extracellular volume expansion conditions, respectively. During alkalosis more sodium than chloride was lost (146 muequiv./min) (P less than 0.001). Colonic pH averaged 7.41 during saline and 6.75 (P less than 0.005) during mannitol perfusion. Titratable acid was not produced in the colon during saline perfusion, and averaged 18 muequiv./min during mannitol perfusion. Urinary titratable acid increased from 19 to 25 muequiv./min (P less than 0.01) during volume expansion. With saline perfusion, bicarbonate secretion rate in the colon rose from 249 muequiv./min during control conditions to 289 muequiv./min during metabolic alkalosis (P less than 0.05). More bicarbonate was excreted in the urine during alkalosis when mannitol was introduced in the colon (243 muequiv./min) than when saline was perfused (152 muequiv./min) (P less than 0.05). This study indicates that the response of the human colon is trivial compared with that of the kidney during acute changes in acid-base balance. | Effects of saline infusion and acute metabolic acidosis and alkalosis on water and electrolyte transport in the human colon. Both the kidney and colon secrete bicarbonate and transport water and electrolytes. The respective contributions of these two organs to acid-base and electrolyte balance in normal man has thus been studied in eight healthy male volunteers who underwent simultaneous renal clearance studies, and colonic perfusion with a 0.9% saline or 7.2% mannitol solution, during metabolic alkalosis and acidosis, extracellular volume expansion, and control conditions. There was no influence of these acid-base conditions on electrolyte transport in the colon. In the urine, preferential loss of chloride over sodium averaged 81, 143 (P less than 0.001), and 141 (P less than 0.05) muequiv./min, during control, metabolic acidosis, and extracellular volume expansion conditions, respectively. During alkalosis more sodium than chloride was lost (146 muequiv./min) (P less than 0.001). Colonic pH averaged 7.41 during saline and 6.75 (P less than 0.005) during mannitol perfusion. Titratable acid was not produced in the colon during saline perfusion, and averaged 18 muequiv./min during mannitol perfusion. Urinary titratable acid increased from 19 to 25 muequiv./min (P less than 0.01) during volume expansion. With saline perfusion, bicarbonate secretion rate in the colon rose from 249 muequiv./min during control conditions to 289 muequiv./min during metabolic alkalosis (P less than 0.05). More bicarbonate was excreted in the urine during alkalosis when mannitol was introduced in the colon (243 muequiv./min) than when saline was perfused (152 muequiv./min) (P less than 0.05). This study indicates that the response of the human colon is trivial compared with that of the kidney during acute changes in acid-base balance. | [
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PMID:14781 | Induction of urogenital anomalies and some tumors in the progeny of mice receiving diethylstilbestrol during pregnancy. | Pregnant mice were given a single dose (10 mug/g body weight) of diethylstilbestrol (DES) on Days 7 to 19, which correspond to the first to fifth lunar months in humans, after the authors, using a 14C-labeled compound, confirmed easy placental penetration by DES. Treatment with DES on Days 15 to 19 resulted in the induction of persistent urogenital sinus (15.8 to 92.5%) and hypertrophy of the portio vaginalis (11.8 to 73.3%) in female offspring, and treatment on Days 17 and 19 resulted in the induction of undescended testes and their hypogenesis (70.4 to 73.3%) in male offspring, although treatment with DES at other stages of pregnancy and after birth did not cause these alterations. The incidence of various tumors (lung adenoma, granulosa cell tumor, etc.) increased significantly (31.0 to 37.9%) when DES was given on Days 15 and 17, which correspond to the stage sensitive to other carcinogens. However, adenosis and adenocarcinoma of the vagina were not observed in the offspring. | Induction of urogenital anomalies and some tumors in the progeny of mice receiving diethylstilbestrol during pregnancy. Pregnant mice were given a single dose (10 mug/g body weight) of diethylstilbestrol (DES) on Days 7 to 19, which correspond to the first to fifth lunar months in humans, after the authors, using a 14C-labeled compound, confirmed easy placental penetration by DES. Treatment with DES on Days 15 to 19 resulted in the induction of persistent urogenital sinus (15.8 to 92.5%) and hypertrophy of the portio vaginalis (11.8 to 73.3%) in female offspring, and treatment on Days 17 and 19 resulted in the induction of undescended testes and their hypogenesis (70.4 to 73.3%) in male offspring, although treatment with DES at other stages of pregnancy and after birth did not cause these alterations. The incidence of various tumors (lung adenoma, granulosa cell tumor, etc.) increased significantly (31.0 to 37.9%) when DES was given on Days 15 and 17, which correspond to the stage sensitive to other carcinogens. However, adenosis and adenocarcinoma of the vagina were not observed in the offspring. | [
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|
PMID:14782 | Production of formaldehyde from N5-methyltetrahydrofolate by normal and leukemic leukocytes. | Extracts of human normal and leukemic leukocytes contain an enzyme that catalyzes a transfer of labeled methyl carbon from N5-[14C]methyltetrahydrofolate to tryptamine. Evidence is presented that this reaction is not attributable to a methyltransferase but to the following reaction sequence: (a) an oxidation of N5-[14C]methyltetrahydrofolate to N5, N10-[14C]methylenetetrahydrofolate that is catalyzed by N5, N10-methylenetetrahydrofolate reductase (EC 1.1.1.68); (b) spontaneous release of [14C]formaldehyde from N5, N10-[14C]methylenetetrahydrofolate; and (c) nonenzymatic condensation of [14C]formaldehyde with tryptamine to form a radioactive carboline derivative. The occurrence of this sequence in leukocytes is suggested by data that show that the enzyme reaction is strongly stimulated by addition of flavin adenine dinucleotide and that the final product is chromatographically identical to the adduct formed in the reaction of [14C]formaldehyde with tryptamine. In the absence of tryptamine, a product accumulates that can react with other HCHO acceptors, i.e., beta-phenylethylamine and dimedone; another reaction product is tetrahydrofolate. Production of formaldehyde is relatively more active in normal lymphocytes than in normal granulocytes, but it is even higher in lymphocytes of chronic lymphocytic leukemia. Activity in granulocytes from a subject with chronic myelocytic leukemia is also elevated but to a lesser extent than activity in lymphocytes of chronic lymphocytic leukemia. Activity in granulocytes from a subject with chronic myelocytic leukemia is also elevated but to a lesser extent than activity in lymphocytes of chronic lymphocytic leukemia. Formaldehyde production in leukocytes is only slightly stimulated by addition of various cobalamins, and activity is normal in leukocytes from a vitamin B12-deficient patient. We conclude that the system is cobalamin independent. Thus, there exists an active pathway from N5-methyltetrahydrofolate to tetrahydrofolate other than the one catalyzed by cobalamin-dependent N5-methyltetrahydrofolate-homocysteine methyltransferase. | Production of formaldehyde from N5-methyltetrahydrofolate by normal and leukemic leukocytes. Extracts of human normal and leukemic leukocytes contain an enzyme that catalyzes a transfer of labeled methyl carbon from N5-[14C]methyltetrahydrofolate to tryptamine. Evidence is presented that this reaction is not attributable to a methyltransferase but to the following reaction sequence: (a) an oxidation of N5-[14C]methyltetrahydrofolate to N5, N10-[14C]methylenetetrahydrofolate that is catalyzed by N5, N10-methylenetetrahydrofolate reductase (EC 1.1.1.68); (b) spontaneous release of [14C]formaldehyde from N5, N10-[14C]methylenetetrahydrofolate; and (c) nonenzymatic condensation of [14C]formaldehyde with tryptamine to form a radioactive carboline derivative. The occurrence of this sequence in leukocytes is suggested by data that show that the enzyme reaction is strongly stimulated by addition of flavin adenine dinucleotide and that the final product is chromatographically identical to the adduct formed in the reaction of [14C]formaldehyde with tryptamine. In the absence of tryptamine, a product accumulates that can react with other HCHO acceptors, i.e., beta-phenylethylamine and dimedone; another reaction product is tetrahydrofolate. Production of formaldehyde is relatively more active in normal lymphocytes than in normal granulocytes, but it is even higher in lymphocytes of chronic lymphocytic leukemia. Activity in granulocytes from a subject with chronic myelocytic leukemia is also elevated but to a lesser extent than activity in lymphocytes of chronic lymphocytic leukemia. Activity in granulocytes from a subject with chronic myelocytic leukemia is also elevated but to a lesser extent than activity in lymphocytes of chronic lymphocytic leukemia. Formaldehyde production in leukocytes is only slightly stimulated by addition of various cobalamins, and activity is normal in leukocytes from a vitamin B12-deficient patient. We conclude that the system is cobalamin independent. Thus, there exists an active pathway from N5-methyltetrahydrofolate to tetrahydrofolate other than the one catalyzed by cobalamin-dependent N5-methyltetrahydrofolate-homocysteine methyltransferase. | [
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PMID:14783 | Suppression of antibody-mediated and cell-mediated murine immunity by the carcinogen N-[4-(5-nitro-2-furyl)-2-thiazolyl]acetamide. | N-[4-(5-Nitro-2-furyl)-2-thiazolyl]acetamide (NFTA) administered at 1000 ppm in diet to mice for 12 weeks induced a high incidence of lymphocytic leukemia. Effects of NFTA on antibody-mediated immunity and cell-mediated immunity of BALB/c mice were studied using the spleen plaque assay for detection of immunoglobulin M-producing cells and the graft-versus-host (GVH) reaction, respectively. NFTA suppressed both responses. With the spleen plaque assay, the number of antibody-forming cells (AFC) to sheep red blood cells was significantly less than in unmedicated, control mice after treated mice received NFTA at 1000 ppm for 6 days. The GVH reaction was not suppressed at 21 days, but was severely suppressed at 70 days, prior to the histological appearance of leukemia. Effect of dose was studied by administering NFTA at 100, 250, 500, and 1000 ppm of diet for 13 to 14 weeks and then determining the response in the spleen plaque assay and GVH reactions. The ratio of AFC/spleen of NFTA-treated groups to AFC/spleen of an unmedicated control group, at the above specified doses, was 0.86, 0.22, 0.33, and 0.54 in ascending dosage order beginning with 100 ppm. For the GVH reaction, the suppression of the cell-mediated immunity was directly proportional to the dose of NFTA. Suppression of the antibody-mediated immunity in relation to the induction of leukemia at 28 weeks was studied by feeding NFTA at 500 ppm for 14 weeks, followed by unmedicated diet for 14 weeks. During the 11th week, mice were immunized with SRBC; 5 days later the spleens were removed and the spleen plaque assay was performed. Eight of 18 mice fed NFTA developed leukemia. The number of AFC/spleen was 78 X 10(3) +/- 34 for those with leukemia and 68 X 10(3) +/- 24 (p greater than 0.5) for those without leukemia, compared with 170 X 10(3) +/- 74 for the control mice (p less than 0.01 for both groups, compared with controls). A closely related carcinogenic nitrofuran, N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide, did not suppress the antibody-mediated immunity response measured during the 11th week of administration. | Suppression of antibody-mediated and cell-mediated murine immunity by the carcinogen N-[4-(5-nitro-2-furyl)-2-thiazolyl]acetamide. N-[4-(5-Nitro-2-furyl)-2-thiazolyl]acetamide (NFTA) administered at 1000 ppm in diet to mice for 12 weeks induced a high incidence of lymphocytic leukemia. Effects of NFTA on antibody-mediated immunity and cell-mediated immunity of BALB/c mice were studied using the spleen plaque assay for detection of immunoglobulin M-producing cells and the graft-versus-host (GVH) reaction, respectively. NFTA suppressed both responses. With the spleen plaque assay, the number of antibody-forming cells (AFC) to sheep red blood cells was significantly less than in unmedicated, control mice after treated mice received NFTA at 1000 ppm for 6 days. The GVH reaction was not suppressed at 21 days, but was severely suppressed at 70 days, prior to the histological appearance of leukemia. Effect of dose was studied by administering NFTA at 100, 250, 500, and 1000 ppm of diet for 13 to 14 weeks and then determining the response in the spleen plaque assay and GVH reactions. The ratio of AFC/spleen of NFTA-treated groups to AFC/spleen of an unmedicated control group, at the above specified doses, was 0.86, 0.22, 0.33, and 0.54 in ascending dosage order beginning with 100 ppm. For the GVH reaction, the suppression of the cell-mediated immunity was directly proportional to the dose of NFTA. Suppression of the antibody-mediated immunity in relation to the induction of leukemia at 28 weeks was studied by feeding NFTA at 500 ppm for 14 weeks, followed by unmedicated diet for 14 weeks. During the 11th week, mice were immunized with SRBC; 5 days later the spleens were removed and the spleen plaque assay was performed. Eight of 18 mice fed NFTA developed leukemia. The number of AFC/spleen was 78 X 10(3) +/- 34 for those with leukemia and 68 X 10(3) +/- 24 (p greater than 0.5) for those without leukemia, compared with 170 X 10(3) +/- 74 for the control mice (p less than 0.01 for both groups, compared with controls). A closely related carcinogenic nitrofuran, N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide, did not suppress the antibody-mediated immunity response measured during the 11th week of administration. | [
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|
PMID:14784 | Inhibitors of L-asparagine synthetase, in vitro. | A systematic search has been made for inhibitors of L-asparagine synthetase (L-glutamine hydrolyzing, EC 6.3.5.4) from leukemia 5178Y/AR, a rodent neoplasm resistant to the oncolytic enzyme L-asparaginase (EC 3.5.1.1), The classes of chemicals examined in this search included substrate and product analogs, agents capable of reacting with sulfhydryl functions, and a variety of modifiers whose mechanism of interaction with proteins is known. In general, antagonists of L-glutamine and thiol reagents proved to be the most effective inhibitors of L-asparagine synthetase from this tumor source. Within these groups, certain structural prerequisites to inhibition are reported. Attempts to correlate oncolytic potency with enzyme-inhibitory potency were unsuccesful. | Inhibitors of L-asparagine synthetase, in vitro. A systematic search has been made for inhibitors of L-asparagine synthetase (L-glutamine hydrolyzing, EC 6.3.5.4) from leukemia 5178Y/AR, a rodent neoplasm resistant to the oncolytic enzyme L-asparaginase (EC 3.5.1.1), The classes of chemicals examined in this search included substrate and product analogs, agents capable of reacting with sulfhydryl functions, and a variety of modifiers whose mechanism of interaction with proteins is known. In general, antagonists of L-glutamine and thiol reagents proved to be the most effective inhibitors of L-asparagine synthetase from this tumor source. Within these groups, certain structural prerequisites to inhibition are reported. Attempts to correlate oncolytic potency with enzyme-inhibitory potency were unsuccesful. | [
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|
PMID:14785 | Coprecipitation of active uridine kinase and phosphomonoesterase from rat kidney by Zn2+-ions. III. Enzymes relevant to cancer chemotherapy. | Partially purified enzyme fraction from rat kidney possessing high uridine kinase and phosphomonoesterase activity was insolubilized by means of zinc precipitation without substantial loss of the activity. While uridine kinase in a soluble and Zn-precipitated form was inhibited by low concentrations (0.5-1.0 mM) of Zn2+-ions, phosphomonoesterase was fully active. In contrast to the soluble fraction, the two enzymes in zinc-precipitated and lyophilized preparations were stable on heating at 100 degrees C. Metal complexed proteins catalyze the dephosphorylation of 5'-UMP, 6-AzaUMP as well as of 2'(3')-UMP or 2,4-dinitrophenyl phosphate indicating thus the presence of several phosphomonoesterases in the complex. | Coprecipitation of active uridine kinase and phosphomonoesterase from rat kidney by Zn2+-ions. III. Enzymes relevant to cancer chemotherapy. Partially purified enzyme fraction from rat kidney possessing high uridine kinase and phosphomonoesterase activity was insolubilized by means of zinc precipitation without substantial loss of the activity. While uridine kinase in a soluble and Zn-precipitated form was inhibited by low concentrations (0.5-1.0 mM) of Zn2+-ions, phosphomonoesterase was fully active. In contrast to the soluble fraction, the two enzymes in zinc-precipitated and lyophilized preparations were stable on heating at 100 degrees C. Metal complexed proteins catalyze the dephosphorylation of 5'-UMP, 6-AzaUMP as well as of 2'(3')-UMP or 2,4-dinitrophenyl phosphate indicating thus the presence of several phosphomonoesterases in the complex. | [
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|
PMID:14790 | Effects of adrenergic amines on electrophysiological properties and automaticity of neonatal and adult canine Purkinje fibers: evidence for alpha- and beta-adrenergic actions. | We determined age-related differences in automaticity and responsiveness of cardiac Purkinje fibers from adult and neonatal dogs to graded concentrations of epinephrine, isoproterenol, and phenylephrine. Purkinje fibers were studied with standard microelectrode techniques during superfusion with Tyrode's solution at 37 degrees C. Control spontaneous rates for adults and neonates did not differ significantly. There was a biphasic response to all agonists such that rate decreased at low and increased at high concentrations. The decrease was greater with phenylephrine and epinephrine than with isoproterenol. The increase in rate was greater with isoproterenol and epinephrine than with phenylephrine. Propranolol shifted the dose-response curves downward and to the right for all agonists; phentolamine, shifter the curves upward and to the left. Epinephrine and isoproterenol dose-response curves for the neonates were upward and to the left of those for adults. Phenylephrine curves were identical for adults and neonates. Thus there are alpha- and beta-adrenergic effects on Purkinje fiber automaticity; the former decrease and the latter increase rate. Furthermore, the effects on automiticity of beta-adrenergic amines are greater in the neonates than in the adult. | Effects of adrenergic amines on electrophysiological properties and automaticity of neonatal and adult canine Purkinje fibers: evidence for alpha- and beta-adrenergic actions. We determined age-related differences in automaticity and responsiveness of cardiac Purkinje fibers from adult and neonatal dogs to graded concentrations of epinephrine, isoproterenol, and phenylephrine. Purkinje fibers were studied with standard microelectrode techniques during superfusion with Tyrode's solution at 37 degrees C. Control spontaneous rates for adults and neonates did not differ significantly. There was a biphasic response to all agonists such that rate decreased at low and increased at high concentrations. The decrease was greater with phenylephrine and epinephrine than with isoproterenol. The increase in rate was greater with isoproterenol and epinephrine than with phenylephrine. Propranolol shifted the dose-response curves downward and to the right for all agonists; phentolamine, shifter the curves upward and to the left. Epinephrine and isoproterenol dose-response curves for the neonates were upward and to the left of those for adults. Phenylephrine curves were identical for adults and neonates. Thus there are alpha- and beta-adrenergic effects on Purkinje fiber automaticity; the former decrease and the latter increase rate. Furthermore, the effects on automiticity of beta-adrenergic amines are greater in the neonates than in the adult. | [
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|
PMID:14791 | Autonomic control of cardiovascular functions during neonatal development and in adult sheep. | We studied the autonomic control of resting heart rate of systemic and pulmonary vascular blood pressures (BP) in chronically instrumented neonatal lambs 1-8 weeks of age. The maximum response to ganglionic blockade and sympathetic and parasympathetic antagonists was taken as an index of the magnitude of the total neural, adrenergic, and cholinergic tones. The reactivity of the circulatory parameters to adrenergic and cholinergic agonists also was investigated. All findings were compared with those in adult nonpregnant sheep studied concomitantly and with data previously obtained from term fetal lambs. The results of our studies show: (1) resting heart rate declines spontaneously throughout the 8 weeks of neonatal life approaching that of adult sheep; (2) the progressive bradycardia is not related to changes in the parasympathetic or sympathetic tone; (3) resting systemic BP is under strong neurohumoral control during the first two to three weeks of neonatal life; the control decreases progressively, becoming similar to that of adult sheep; (4) resting pulmonary artery pressure of neonatal and adult sheep has no neurohumoral control; (5) the systemic BP response of the neonate to autonomic agonists is greater than that of the term fetus and is similar to that of the adult; (6) in neonatal and adult sheep, compared to the term fetus, the pressor response to norepinephrine is accompanied by a baroreceptor-mediated bradycardia, and acetylcholine-induced systemci hypotension is accompanied by a "paradoxical" tachycardia mediated through beta-adrenergic stimulation; (7) in contrast to our finding for the fetus, the pulmonary vascular pressure of neonatal and adult sheep is unresponsive to autonomic agonists. | Autonomic control of cardiovascular functions during neonatal development and in adult sheep. We studied the autonomic control of resting heart rate of systemic and pulmonary vascular blood pressures (BP) in chronically instrumented neonatal lambs 1-8 weeks of age. The maximum response to ganglionic blockade and sympathetic and parasympathetic antagonists was taken as an index of the magnitude of the total neural, adrenergic, and cholinergic tones. The reactivity of the circulatory parameters to adrenergic and cholinergic agonists also was investigated. All findings were compared with those in adult nonpregnant sheep studied concomitantly and with data previously obtained from term fetal lambs. The results of our studies show: (1) resting heart rate declines spontaneously throughout the 8 weeks of neonatal life approaching that of adult sheep; (2) the progressive bradycardia is not related to changes in the parasympathetic or sympathetic tone; (3) resting systemic BP is under strong neurohumoral control during the first two to three weeks of neonatal life; the control decreases progressively, becoming similar to that of adult sheep; (4) resting pulmonary artery pressure of neonatal and adult sheep has no neurohumoral control; (5) the systemic BP response of the neonate to autonomic agonists is greater than that of the term fetus and is similar to that of the adult; (6) in neonatal and adult sheep, compared to the term fetus, the pressor response to norepinephrine is accompanied by a baroreceptor-mediated bradycardia, and acetylcholine-induced systemci hypotension is accompanied by a "paradoxical" tachycardia mediated through beta-adrenergic stimulation; (7) in contrast to our finding for the fetus, the pulmonary vascular pressure of neonatal and adult sheep is unresponsive to autonomic agonists. | [
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|
PMID:14792 | Baroreflex sensitivity in patients with Takayasu's aortitis. | Takayasu's aortitis is an arterial inflammatory disease of arteries of unknown etiology. Fainting is a common symptom and has been attributed to ypersensitivity of the baroreflex. We studied baroreflex sensitivity in 11 patients with Takayasu's aortitis and compared it with that of eight control subjects of comparable age. Baroreflex sensitivity was assessed by determining the slope of a regression line relating the rise of systolic arterial pressure to the prolongation of the R-R interval of the electrocardiogram during a transient rise of arterial pressure induced by an intravenous injection of phenylephrine. The average baroreflex slope of patients with Takayasu's arteritis (4.0 +/- 0.8 msec/mm Hg) was significantly less than that of control subjects (10.7 +/- 0.8 msec/mm Hg, P less than 0.001). Reduced baroreflex sensitivity in patients with Takayasu's aortitis may be due to the hardening of the arteries where baroreceptors lie, or to hypertension and/or cardiac disease which was present in most of the patients included in this study. Patients with Takayasu's aortitis who complained of fainting also showed the reduced baroreflex sensitivity. This indicates that fainting in this disease is not likely to be caused by the hyperreactivity of the baroreceptors as is commonly postulated. | Baroreflex sensitivity in patients with Takayasu's aortitis. Takayasu's aortitis is an arterial inflammatory disease of arteries of unknown etiology. Fainting is a common symptom and has been attributed to ypersensitivity of the baroreflex. We studied baroreflex sensitivity in 11 patients with Takayasu's aortitis and compared it with that of eight control subjects of comparable age. Baroreflex sensitivity was assessed by determining the slope of a regression line relating the rise of systolic arterial pressure to the prolongation of the R-R interval of the electrocardiogram during a transient rise of arterial pressure induced by an intravenous injection of phenylephrine. The average baroreflex slope of patients with Takayasu's arteritis (4.0 +/- 0.8 msec/mm Hg) was significantly less than that of control subjects (10.7 +/- 0.8 msec/mm Hg, P less than 0.001). Reduced baroreflex sensitivity in patients with Takayasu's aortitis may be due to the hardening of the arteries where baroreceptors lie, or to hypertension and/or cardiac disease which was present in most of the patients included in this study. Patients with Takayasu's aortitis who complained of fainting also showed the reduced baroreflex sensitivity. This indicates that fainting in this disease is not likely to be caused by the hyperreactivity of the baroreceptors as is commonly postulated. | [
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PMID:14793 | Amperometric determination of total cholesterol in serum, with use of immobilized cholesterol ester hydrolase and cholesterol oxidase. | We describe an electrochemical method for simple, rapid, and economical assay of total serum cholesterol with use of immobilized cholesterol esterase (EC 3.1.1.13) and cholesterol oxidase (EC 1.1.3.6). A rotating porous cell was specially designed to hold the immobilized enzymes firmly and to allow the reaction mixture to pass through the enzyme layer easily, thus catalyzing the enzymatic transformation quickly. Hydrogen peroxide resulting from a catalytic reactions was measured amperometrically at +0.60 V cs. a standard calomel electrode. The calibration curve for total serum cholesterol was linear from 0 to 5.00 g/liter. The method is specific, precise, and inexpensive. Our results correlate well with those obtained by the method of Abell et al. [Stand. Methods Clin. Chem. 2, 26 (1958)], the correlation coefficient being 0.992. Ascorbic acid or bilirubin in concentrations up to 700 mg/liter do not interfere. The immobilized enzymes are stable, and the same immobilized-enzyme stirrer can be used for at least 200 accurate, reproducible assays. | Amperometric determination of total cholesterol in serum, with use of immobilized cholesterol ester hydrolase and cholesterol oxidase. We describe an electrochemical method for simple, rapid, and economical assay of total serum cholesterol with use of immobilized cholesterol esterase (EC 3.1.1.13) and cholesterol oxidase (EC 1.1.3.6). A rotating porous cell was specially designed to hold the immobilized enzymes firmly and to allow the reaction mixture to pass through the enzyme layer easily, thus catalyzing the enzymatic transformation quickly. Hydrogen peroxide resulting from a catalytic reactions was measured amperometrically at +0.60 V cs. a standard calomel electrode. The calibration curve for total serum cholesterol was linear from 0 to 5.00 g/liter. The method is specific, precise, and inexpensive. Our results correlate well with those obtained by the method of Abell et al. [Stand. Methods Clin. Chem. 2, 26 (1958)], the correlation coefficient being 0.992. Ascorbic acid or bilirubin in concentrations up to 700 mg/liter do not interfere. The immobilized enzymes are stable, and the same immobilized-enzyme stirrer can be used for at least 200 accurate, reproducible assays. | [
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PMID:14794 | Evaluation of a new automatic calcium ion analyzer. | We evaluated a new automatic analyzer for ionized calcium (Ca2+), the Orion Model SS-20, based on a flowthrough ion-exchange electrode. Ca2+ was measured in heparinized whole blood and serum. Within-day variation was 1.2%, day-to-day variation1.6%, and analytical recovery 92.4%. Over the phsiological range interference by K+ and Mg2+ was negligible; major changes in ionic strength, induced by changes in N+ concentration, made correction for a sodium error necessary. Within the physiological range, Ca2+ was inversely correlated to variations in pH. Therefore, to compare Ca2+ values, correction to an apparent pH of 7.40 should be made. The calcium binding effect of heparin was negligible when minimal (4.4 int. units/ml) concentrations of heparin were used. Storage of serum at 4 degrees C for one week resulted in a 4 degrees decrease in apparent serum Ca2+, primarily owing to an increase in pH during storage. In normal material, mean values for blood-and serum-Ca2+(1.10 and 1.07 mmol/liter, respectively) were close to results obtained by previous systems. Errors caused by disturbances in the fluid flow and non-function of half the electrodes we received were the major inconveniences of the analyzer. We conclude that this new analyzer gives decisive advantages in measurement of Ca2+, making this importnant analysis possible as a routine laboratory test for the first time. | Evaluation of a new automatic calcium ion analyzer. We evaluated a new automatic analyzer for ionized calcium (Ca2+), the Orion Model SS-20, based on a flowthrough ion-exchange electrode. Ca2+ was measured in heparinized whole blood and serum. Within-day variation was 1.2%, day-to-day variation1.6%, and analytical recovery 92.4%. Over the phsiological range interference by K+ and Mg2+ was negligible; major changes in ionic strength, induced by changes in N+ concentration, made correction for a sodium error necessary. Within the physiological range, Ca2+ was inversely correlated to variations in pH. Therefore, to compare Ca2+ values, correction to an apparent pH of 7.40 should be made. The calcium binding effect of heparin was negligible when minimal (4.4 int. units/ml) concentrations of heparin were used. Storage of serum at 4 degrees C for one week resulted in a 4 degrees decrease in apparent serum Ca2+, primarily owing to an increase in pH during storage. In normal material, mean values for blood-and serum-Ca2+(1.10 and 1.07 mmol/liter, respectively) were close to results obtained by previous systems. Errors caused by disturbances in the fluid flow and non-function of half the electrodes we received were the major inconveniences of the analyzer. We conclude that this new analyzer gives decisive advantages in measurement of Ca2+, making this importnant analysis possible as a routine laboratory test for the first time. | [
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PMID:14796 | The diagnosis of Gaucher's disease in liver using 4-methylumbelliferyl-beta-D-glucopyranoside. | 1. Cases of Gaucher's disease could not be distinguished from controls by the assay of beta-glucosidase activity in water homogenates of liver using 4-methylumbelliferyl-beta-D glucopyranoside. 2. Two peaks of beta-glucosidase activity were separated by Sephadex G-150 gel filtration in control and Gaucher livers. In the presence of the elution buffer pH profiles of peak I showed a deficiency at pH 3.5-4.5 in Gaucher's disease. Gaucher and control peak II had similar pH profiles with little or no activity at pH 3.0-4.0. 3. A clear distinction between homogenates of Gaucher and control liver was obtained by assay at pH 4.0 in the presence of elution buffer, or of sodium chloride, a component of the elution buffer. | The diagnosis of Gaucher's disease in liver using 4-methylumbelliferyl-beta-D-glucopyranoside. 1. Cases of Gaucher's disease could not be distinguished from controls by the assay of beta-glucosidase activity in water homogenates of liver using 4-methylumbelliferyl-beta-D glucopyranoside. 2. Two peaks of beta-glucosidase activity were separated by Sephadex G-150 gel filtration in control and Gaucher livers. In the presence of the elution buffer pH profiles of peak I showed a deficiency at pH 3.5-4.5 in Gaucher's disease. Gaucher and control peak II had similar pH profiles with little or no activity at pH 3.0-4.0. 3. A clear distinction between homogenates of Gaucher and control liver was obtained by assay at pH 4.0 in the presence of elution buffer, or of sodium chloride, a component of the elution buffer. | [
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|
PMID:14797 | A new semiautomated fluorometric method for estimation of small amounts of L-dopa in human urine. | A semiautomated fluorometric method for the quantitative determination of urinary dopa, 3-(3,4-dihydroxyphenyl)-L-alanine, is described. It provides a simple, sensitive and reproducible analytical technique for routine use. Dopa is isolated from interfering substances, especially catecholamines, by adsorption onto aluminium oxide, elution with 0.1 M HCl, then passed through a cation-exchange column. By mild oxidation with potassium ferricyanide, dopa is cyclized to dopachrome. This is isomerised with strong alkali to 5,6-substituted indole, which is highly fluorescent but very unstable in the presence of oxygen. The addition of ascorbic acid and 3-mercaptopropionic acid to the alkaline solution stabilizes the fluorescence. Concentrations of chemicals, pH and reaction times are made optimal for maximal sensitivity. Recovery of dopa added to the urine samples averaged 79% (range, 75 to 83). Reproducibility of results from the same urine specimen gave a coefficient of variation of 2.4%. In healthy adults, we found daily excretion ranges from 17.3 to 54.9 (mean: 36.1) microng/24 h or from 0.015 to 0.048 (mean: 0.021) microng/mg cretinine. | A new semiautomated fluorometric method for estimation of small amounts of L-dopa in human urine. A semiautomated fluorometric method for the quantitative determination of urinary dopa, 3-(3,4-dihydroxyphenyl)-L-alanine, is described. It provides a simple, sensitive and reproducible analytical technique for routine use. Dopa is isolated from interfering substances, especially catecholamines, by adsorption onto aluminium oxide, elution with 0.1 M HCl, then passed through a cation-exchange column. By mild oxidation with potassium ferricyanide, dopa is cyclized to dopachrome. This is isomerised with strong alkali to 5,6-substituted indole, which is highly fluorescent but very unstable in the presence of oxygen. The addition of ascorbic acid and 3-mercaptopropionic acid to the alkaline solution stabilizes the fluorescence. Concentrations of chemicals, pH and reaction times are made optimal for maximal sensitivity. Recovery of dopa added to the urine samples averaged 79% (range, 75 to 83). Reproducibility of results from the same urine specimen gave a coefficient of variation of 2.4%. In healthy adults, we found daily excretion ranges from 17.3 to 54.9 (mean: 36.1) microng/24 h or from 0.015 to 0.048 (mean: 0.021) microng/mg cretinine. | [
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|
PMID:14798 | Phenylethanolamine N-methyltransferase and other enzymes of catecholamine metabolism in human brain. | The activities of tyrosine hydroxylase (TH), DOPA decarboxylase (DDC), dopamine beta-hydroxylase (DBH), phenylethanolamine N-methyltransferase (PNMT), and monoamine oxidase (MAO) with serotonin and phenylethylamine as substrates were measured in catecholaminergic regions of human brain from 10 controls and 3 patients with Parkinsonism. PNMT activity was detected in hypothalamus, thalamus and cerebellar nucleus of the control human brain, and was reduced in hypothalamus of Parkinsonian cases. Type A (with serotonin as substrate) and type B (with phenylethylamine as substrate) MAO activities were high in all brain regions with little individual variations in controls and Parkinsonian cases. TH activity was high in the controls and was markedly decreased, in substantia nigra, caudate nucleus, putamen and in pallidum, in all three cases of Parkinsonism. DDC activity in these regions was also decreased in 2 patients. However, one Parkinsonian case had only decreased TH and normal DDC activities. DBH activity in hypothalamus was also reduced in the Parkinsonian cases. | Phenylethanolamine N-methyltransferase and other enzymes of catecholamine metabolism in human brain. The activities of tyrosine hydroxylase (TH), DOPA decarboxylase (DDC), dopamine beta-hydroxylase (DBH), phenylethanolamine N-methyltransferase (PNMT), and monoamine oxidase (MAO) with serotonin and phenylethylamine as substrates were measured in catecholaminergic regions of human brain from 10 controls and 3 patients with Parkinsonism. PNMT activity was detected in hypothalamus, thalamus and cerebellar nucleus of the control human brain, and was reduced in hypothalamus of Parkinsonian cases. Type A (with serotonin as substrate) and type B (with phenylethylamine as substrate) MAO activities were high in all brain regions with little individual variations in controls and Parkinsonian cases. TH activity was high in the controls and was markedly decreased, in substantia nigra, caudate nucleus, putamen and in pallidum, in all three cases of Parkinsonism. DDC activity in these regions was also decreased in 2 patients. However, one Parkinsonian case had only decreased TH and normal DDC activities. DBH activity in hypothalamus was also reduced in the Parkinsonian cases. | [
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|
PMID:14799 | Influence of the pH on the antigen-antibody coupling in angiotensin I radioimmunoassay. Its consequences for the determination of the plasma renin activity. | The antigen-antibody complex formation in the angiotensin I radioimmunoassay appears to be influenced by the pH of the radioimmunoassay incubation mixture. This may lead to erroneous results in the determination of the plasma renin activity, when an aliquot of a plasma sample, buffered at pH 6 for optimum renin activity is brought into a radioimmunoassay mixture of another pH, while the radioimmunoassay standards are not corrected for this pH shift. In our experiments we studied this general pH effect, and evaluated the effect on the New England Nuclear Angiotensin I radioimmunoassay procedure. We propose a slight modification of this radioimmunoassay kit. | Influence of the pH on the antigen-antibody coupling in angiotensin I radioimmunoassay. Its consequences for the determination of the plasma renin activity. The antigen-antibody complex formation in the angiotensin I radioimmunoassay appears to be influenced by the pH of the radioimmunoassay incubation mixture. This may lead to erroneous results in the determination of the plasma renin activity, when an aliquot of a plasma sample, buffered at pH 6 for optimum renin activity is brought into a radioimmunoassay mixture of another pH, while the radioimmunoassay standards are not corrected for this pH shift. In our experiments we studied this general pH effect, and evaluated the effect on the New England Nuclear Angiotensin I radioimmunoassay procedure. We propose a slight modification of this radioimmunoassay kit. | [
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|
PMID:14802 | Acute overdosage with benzodiazepine derivatives. | A total of 773 admissions to Massachusetts General Hospital between 1962 and 1975 were due to acute overdosage with one or more psychotropic drugs. Benzodiazepine overdosage, particularly with diazepam, increased relative to other psychotropic drugs over the years. Only 12 admissions were due to benzodiazepine overdosage alone, and none of these patients were seriously ill or had significant complications. Multiple drugs were ingested in the other 87 cases, and the frequency and severity of complications among these individuals depended upon the type and quantity of other nonbenzodiazepines taken. For example, 21 of 31 patients who ingested benzodiazepines together with barbiturates experienced severe central nervous system (CNS) depression, and 14 of 31 required assisted ventilation. However, the frequency of such complications was nearly identical in a group of patients who ingested barbiturates alone. This report and a review of the literature suggest that serious intoxication following overdosage with a benzodiazepine derivative alone is unusual. Ingestion of benzodiazepines together with other drugs appears to be considerably more common than benzodiazepine overdosage alone as a cause of intoxication. The severity of intoxication in such cases of multiple drug ingestion probably depends largely on the type and quantity of nonbenzodiazepines. | Acute overdosage with benzodiazepine derivatives. A total of 773 admissions to Massachusetts General Hospital between 1962 and 1975 were due to acute overdosage with one or more psychotropic drugs. Benzodiazepine overdosage, particularly with diazepam, increased relative to other psychotropic drugs over the years. Only 12 admissions were due to benzodiazepine overdosage alone, and none of these patients were seriously ill or had significant complications. Multiple drugs were ingested in the other 87 cases, and the frequency and severity of complications among these individuals depended upon the type and quantity of other nonbenzodiazepines taken. For example, 21 of 31 patients who ingested benzodiazepines together with barbiturates experienced severe central nervous system (CNS) depression, and 14 of 31 required assisted ventilation. However, the frequency of such complications was nearly identical in a group of patients who ingested barbiturates alone. This report and a review of the literature suggest that serious intoxication following overdosage with a benzodiazepine derivative alone is unusual. Ingestion of benzodiazepines together with other drugs appears to be considerably more common than benzodiazepine overdosage alone as a cause of intoxication. The severity of intoxication in such cases of multiple drug ingestion probably depends largely on the type and quantity of nonbenzodiazepines. | [
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|
PMID:14803 | Effects of metabolic alkalosis, metabolic acidosis and uraemia on whole-body intracellular pH in man. | 1. Whole-body intracellular pH (pHi) was measured by the 14C-labelled DMO method in twenty-four control subjects, eighteen normal subjects with induced acute metabolic alkalosis, ten normal subjects with induced acute metabolic acidosis, twelve normal subjects with chronic acidosis and in fifteen patients with chronic renal insufficiency and acidosis. 2. The change in pHi per unit change in extracellular pH is significantly larger in acute metabolic alkalosis than in acute metabolic acidosis. In chronic metabolic acidosis, pHi decreased in proportion to the total amount of ammonium chloride administered; pHi was normal in patients with uraemic acidosis. 3. These observations confirm the role that tissue buffers play in the protection of the cellular environment in some forms of acidosis. When the acid load overwhelms tissue buffer capacity, pHi becomes a function of extracellular pH. 4. Cells seem more protected from acute acidosis than from acute alkalosis. | Effects of metabolic alkalosis, metabolic acidosis and uraemia on whole-body intracellular pH in man. 1. Whole-body intracellular pH (pHi) was measured by the 14C-labelled DMO method in twenty-four control subjects, eighteen normal subjects with induced acute metabolic alkalosis, ten normal subjects with induced acute metabolic acidosis, twelve normal subjects with chronic acidosis and in fifteen patients with chronic renal insufficiency and acidosis. 2. The change in pHi per unit change in extracellular pH is significantly larger in acute metabolic alkalosis than in acute metabolic acidosis. In chronic metabolic acidosis, pHi decreased in proportion to the total amount of ammonium chloride administered; pHi was normal in patients with uraemic acidosis. 3. These observations confirm the role that tissue buffers play in the protection of the cellular environment in some forms of acidosis. When the acid load overwhelms tissue buffer capacity, pHi becomes a function of extracellular pH. 4. Cells seem more protected from acute acidosis than from acute alkalosis. | [
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|
PMID:14804 | Enzyme activities in human liver biopsies: assay methods and activities of some lysosomal and membrane-bound enzymes in control tissue and serum. | 1. Highly sensitive technique are described for the assay of plasma membrane (5'-nucleotidase, alkaline phosphatase), microsomal (neutral alpha-glucosidase, leucyl-2-naphthylamidase) and biliary canalicular (gamma-glutamyltransferase) enzymes and for nine acid hydrolases (acid phosphatase, phosphodiesterase, beta-glucosidase, alpha-glucosidase, alpha-galactosidase, beta-galactosidase, alpha-mannosidase, N-acetyl-beta-glucosaminidase, beta-glucuronidase) in human liver. 2. Optimum and specific assay systems have been developed which give linear kinetics for all enzymes. 3. The range of enzyme activities in samples of human liver, obtained by closed needle biopsy, and sera have been determined. | Enzyme activities in human liver biopsies: assay methods and activities of some lysosomal and membrane-bound enzymes in control tissue and serum. 1. Highly sensitive technique are described for the assay of plasma membrane (5'-nucleotidase, alkaline phosphatase), microsomal (neutral alpha-glucosidase, leucyl-2-naphthylamidase) and biliary canalicular (gamma-glutamyltransferase) enzymes and for nine acid hydrolases (acid phosphatase, phosphodiesterase, beta-glucosidase, alpha-glucosidase, alpha-galactosidase, beta-galactosidase, alpha-mannosidase, N-acetyl-beta-glucosaminidase, beta-glucuronidase) in human liver. 2. Optimum and specific assay systems have been developed which give linear kinetics for all enzymes. 3. The range of enzyme activities in samples of human liver, obtained by closed needle biopsy, and sera have been determined. | [
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PMID:14805 | Lysosomal changes in liver tissue from patients with the Dubin-Johnson-Sprinz syndrome. | 1. Clinical, morphological and biochemical data, including data obtained from the application of subcellular fractionation techniques to liver biopsy specimens, are presented for two patients with the Dubin-Johnson-Sprinz (DJS) syndrome. 2. Subcellular fractionation experiments demonstrate that the lysosomes, which have strikingly reduced equilibrium densities, accumulate melanin. Morphological studies confirm the presence of pigments within lysosomes. 3. Although there are increased activities of lysosomal acid hydrolases in the liver tissue from patients with the DJS syndrome, the integrity of these organelles is essentially normal and therefore the accumulation of pigment would not be expected to initiate liver damage. The DJS syndrome is thus a benign type of secondary lysosomal storage disease. | Lysosomal changes in liver tissue from patients with the Dubin-Johnson-Sprinz syndrome. 1. Clinical, morphological and biochemical data, including data obtained from the application of subcellular fractionation techniques to liver biopsy specimens, are presented for two patients with the Dubin-Johnson-Sprinz (DJS) syndrome. 2. Subcellular fractionation experiments demonstrate that the lysosomes, which have strikingly reduced equilibrium densities, accumulate melanin. Morphological studies confirm the presence of pigments within lysosomes. 3. Although there are increased activities of lysosomal acid hydrolases in the liver tissue from patients with the DJS syndrome, the integrity of these organelles is essentially normal and therefore the accumulation of pigment would not be expected to initiate liver damage. The DJS syndrome is thus a benign type of secondary lysosomal storage disease. | [
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|
PMID:14806 | Leucocyte intracellular pH in patients with the metabolic acidosis of renal failure. | 1. Human leucocytes were isolated from venous blood and resuspended in the subjects plasma. 2. Intracellular pH measurements were made in vitro by the dimethyloxazolidinedione technique in 13 healthy subjects and in 11 subjects with renal failure and metabolic acidosis. 3. The intracellular pH of the healthy subjects was found to be 7-07 (SD 0-04), significantly lower than that of the patients, which was 7-11 (SD 0-03). For all estimations plasma PCO2 was maintained at approximately 5-5 kPa. The methodological and possible metabolic reasons for this difference in intracellular pH are discussed. | Leucocyte intracellular pH in patients with the metabolic acidosis of renal failure. 1. Human leucocytes were isolated from venous blood and resuspended in the subjects plasma. 2. Intracellular pH measurements were made in vitro by the dimethyloxazolidinedione technique in 13 healthy subjects and in 11 subjects with renal failure and metabolic acidosis. 3. The intracellular pH of the healthy subjects was found to be 7-07 (SD 0-04), significantly lower than that of the patients, which was 7-11 (SD 0-03). For all estimations plasma PCO2 was maintained at approximately 5-5 kPa. The methodological and possible metabolic reasons for this difference in intracellular pH are discussed. | [
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|
PMID:14808 | The influence of instructional set on schizophrenic vs. organic concreteness. | 36 male patients (12 schizophrenic, 12 organic, and 12 neurotic), age 19-57, each took two forms of the Gorham Proverbs test under two different instructional sets. It was hypothesized that for schizophrenics, a higher level of abstraction would be obtained on the proverbs task when the instructions required less personal involvement of the patient than when a more personalized set was used; whereas the performance of organic and neurotic patients would not be affected by the variation in instructional set. The results indicate that instructional set does significantly alter the concreteness of the schizophrenic's response to proverbs, but has little effect on the responses of the other two groups. | The influence of instructional set on schizophrenic vs. organic concreteness. 36 male patients (12 schizophrenic, 12 organic, and 12 neurotic), age 19-57, each took two forms of the Gorham Proverbs test under two different instructional sets. It was hypothesized that for schizophrenics, a higher level of abstraction would be obtained on the proverbs task when the instructions required less personal involvement of the patient than when a more personalized set was used; whereas the performance of organic and neurotic patients would not be affected by the variation in instructional set. The results indicate that instructional set does significantly alter the concreteness of the schizophrenic's response to proverbs, but has little effect on the responses of the other two groups. | [
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|
PMID:14812 | Pulmonary artery involvement in Takayasu's arteritis. | Although pulmonary artery disease in Takayasu's arteritis has been described since 1940, it has received little attention. The clinical, hemodynamic, and angiographic features of the pulmonary involvement were studied in 22 patients with systemic arterial disease. Pulmonary involvement was found in 50 percent of the cases. Moderate pulmonary hypertension was a common finding (73 percent). Lesions were generally localized to the large and medium pulmonary vessels. None of the patients had pulmonary symptoms, but in 63 percent there were clinical, radiologic and electrocardiographic findings suggesting pulmonary hypertension or right heart strain. We believe that the pulmonary circulation should be routinely studied in patients with Takayasu's arteritis and that pulmonary involvement should be included in the classification of the disease. | Pulmonary artery involvement in Takayasu's arteritis. Although pulmonary artery disease in Takayasu's arteritis has been described since 1940, it has received little attention. The clinical, hemodynamic, and angiographic features of the pulmonary involvement were studied in 22 patients with systemic arterial disease. Pulmonary involvement was found in 50 percent of the cases. Moderate pulmonary hypertension was a common finding (73 percent). Lesions were generally localized to the large and medium pulmonary vessels. None of the patients had pulmonary symptoms, but in 63 percent there were clinical, radiologic and electrocardiographic findings suggesting pulmonary hypertension or right heart strain. We believe that the pulmonary circulation should be routinely studied in patients with Takayasu's arteritis and that pulmonary involvement should be included in the classification of the disease. | [
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|
PMID:14813 | Three types of paranoid processes. | In classical descriptive psychiatry the term 'paranoid' is often used ambiguously -- referring to a variety of clinical processes which should be more clearly differentiated. Specifically, in this paper, we have differentiated three distinct sets of clinical phenomena all usually lumped together as 'paranoid': 1. Paranoid from a Sense of Guilt, 2. Paranoid from a sense of Low Self-Esteem, and 3. Paranoid from a Sense of Persecution. These three processes are distinct descriptively, dynamically and genetically. Further, this differentiation is most significant pragmatically as the treatment is different for each type. | Three types of paranoid processes. In classical descriptive psychiatry the term 'paranoid' is often used ambiguously -- referring to a variety of clinical processes which should be more clearly differentiated. Specifically, in this paper, we have differentiated three distinct sets of clinical phenomena all usually lumped together as 'paranoid': 1. Paranoid from a Sense of Guilt, 2. Paranoid from a sense of Low Self-Esteem, and 3. Paranoid from a Sense of Persecution. These three processes are distinct descriptively, dynamically and genetically. Further, this differentiation is most significant pragmatically as the treatment is different for each type. | [
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|
PMID:14809 | Application of the Starling resistor concept to the lungs during CPPV. | The application of the Starling resistor concept of the lungs during continuous positive pressure ventilation (CPPV) was evaluated. Ventilation and hemodynamics were studied in eight anesthetized and paralyzed dogs before and during the use of CPPV. CPPV resulted in an increase in transpulmonary pressure and functional residual capacity (FRC), and a decrease in arterial pH and mixed venous oxygen tension (PvO2). Cardica index decreased 32% (p less than 0.01) and stroke volume 51% (p less than 0.01). Neither right nor left transmural pressures changed but pulmonary vascular resistance increased 189% (p less than 0.01). This study supports the concept that the pulmonary vasculature behaves as a Starling resistor during the use of CPPV. The increase in pulmonary vascular resistance must be considered when transpulmonary pressure is raised by CPPV. | Application of the Starling resistor concept to the lungs during CPPV. The application of the Starling resistor concept of the lungs during continuous positive pressure ventilation (CPPV) was evaluated. Ventilation and hemodynamics were studied in eight anesthetized and paralyzed dogs before and during the use of CPPV. CPPV resulted in an increase in transpulmonary pressure and functional residual capacity (FRC), and a decrease in arterial pH and mixed venous oxygen tension (PvO2). Cardica index decreased 32% (p less than 0.01) and stroke volume 51% (p less than 0.01). Neither right nor left transmural pressures changed but pulmonary vascular resistance increased 189% (p less than 0.01). This study supports the concept that the pulmonary vasculature behaves as a Starling resistor during the use of CPPV. The increase in pulmonary vascular resistance must be considered when transpulmonary pressure is raised by CPPV. | [
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|
PMID:14816 | [Sympathetic responsiveness and antihypertensive effect of beta-receptor blockade in essential hypertension: the effect of atenolol (author's transl)]. | Plasmin noradrenaline concentration after bicycle exercise (200 W for 2 min), compared with base line concentration, was used as an index of sympathetic responsiveness in patients with essential hypertension. Atenolol (JCI 66082, a "cardioselective" beta-blocker) was given in a daily dose of 200 mg to 16 patients for five weeks. This caused a decrease in supine blood pressure of 37/23 and, on standing, of 36/25 mm Hg compared with the placebo period. There was a significant correlation between the ratio of the increase in plasma noradrenaline concentration on exercise over its base line concentration and the subsequent fall in mean arterial pressure (r=0.840; P less than 0.001). There was a less significant correlation between plasma renin concentration and subsequent decrease in mean arterial pressure (r=0.542; P less than 0.05). Administrations of atenolol caused a rise in plasma noradrenaline both on lying and after exercise (P less than 0.0125), and a fall in plasma renin concentration (P less than 0.01). The results suggest that the antihypertensive effect of atenolol is related to the responsiveness of the sympathetic nervous sytem. Adrenergic activity is apparently an important determinant of blood pressure response to beta-blockade. | [Sympathetic responsiveness and antihypertensive effect of beta-receptor blockade in essential hypertension: the effect of atenolol (author's transl)]. Plasmin noradrenaline concentration after bicycle exercise (200 W for 2 min), compared with base line concentration, was used as an index of sympathetic responsiveness in patients with essential hypertension. Atenolol (JCI 66082, a "cardioselective" beta-blocker) was given in a daily dose of 200 mg to 16 patients for five weeks. This caused a decrease in supine blood pressure of 37/23 and, on standing, of 36/25 mm Hg compared with the placebo period. There was a significant correlation between the ratio of the increase in plasma noradrenaline concentration on exercise over its base line concentration and the subsequent fall in mean arterial pressure (r=0.840; P less than 0.001). There was a less significant correlation between plasma renin concentration and subsequent decrease in mean arterial pressure (r=0.542; P less than 0.05). Administrations of atenolol caused a rise in plasma noradrenaline both on lying and after exercise (P less than 0.0125), and a fall in plasma renin concentration (P less than 0.01). The results suggest that the antihypertensive effect of atenolol is related to the responsiveness of the sympathetic nervous sytem. Adrenergic activity is apparently an important determinant of blood pressure response to beta-blockade. | [
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|
PMID:14818 | [Effect of beta-adrenergic blockers on complement]. | The authors, examined the effect of beta-adrenergic blockers Propranolol, Pindolol, Practonol and P11 on complement in the serum of guinea pigs after continuous administration. The follow-up was made dynamicaly: on the 10th, 20th and 30th day. There was a considerable lowering of the complement level on the 20th and 30th day in all animals treated with beta-adrenergic blockers. The experiments in vitro showed that the effect of Propranolol and and P11 on the serum of persons, guinea pigs and rabbits presented data on the complement inhibition, having the direct effect on the first component of the complementary systems. | [Effect of beta-adrenergic blockers on complement]. The authors, examined the effect of beta-adrenergic blockers Propranolol, Pindolol, Practonol and P11 on complement in the serum of guinea pigs after continuous administration. The follow-up was made dynamicaly: on the 10th, 20th and 30th day. There was a considerable lowering of the complement level on the 20th and 30th day in all animals treated with beta-adrenergic blockers. The experiments in vitro showed that the effect of Propranolol and and P11 on the serum of persons, guinea pigs and rabbits presented data on the complement inhibition, having the direct effect on the first component of the complementary systems. | [
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|
PMID:14819 | [Participation of the adrenals in the pathogenesis of metabolic acid-base disorders]. | The authors examined in dynamics the changes in the functional state of the adrenals on 240 rabbits, which served as models for acute metabolic deviations in the acid-base balance. The obtained results showed that the acute metabolic acidosis increased moderately the values of ACTH and 17-hydroxycorticosteroids in blood without changing their concentration on the adrenal tissue. It lowered strongly the content of catecholamines (adrenaline and noradranaline) in the adrenal medular part. The metabolic alkalosis raised the concentration of ACTH in blood plasma and increased the amount of corticosteroids in blood and adrenals. There was no well formed parallelism in normalizing acid-base and hormonal indices. As a consequence of this a stage of postaciodotic catecholamine adrenal deficit was formed as well as metabasic hypercorticism in the experimental animals. | [Participation of the adrenals in the pathogenesis of metabolic acid-base disorders]. The authors examined in dynamics the changes in the functional state of the adrenals on 240 rabbits, which served as models for acute metabolic deviations in the acid-base balance. The obtained results showed that the acute metabolic acidosis increased moderately the values of ACTH and 17-hydroxycorticosteroids in blood without changing their concentration on the adrenal tissue. It lowered strongly the content of catecholamines (adrenaline and noradranaline) in the adrenal medular part. The metabolic alkalosis raised the concentration of ACTH in blood plasma and increased the amount of corticosteroids in blood and adrenals. There was no well formed parallelism in normalizing acid-base and hormonal indices. As a consequence of this a stage of postaciodotic catecholamine adrenal deficit was formed as well as metabasic hypercorticism in the experimental animals. | [
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|
PMID:14820 | [Comparative study of the effect of haloperidol and maptil on the karyotype and immunoproliferative response of lymphocytes]. | The author examined the action of haloperidol and magetyl on the mitotic activity, blast transformation and karyotype of the lymphocytic cultures, obtained from the peripheral blood in vitro. The experiments were carried out on flymphycytes, obtained from ten healthy individuals. Nontreated cultures, obtained from the same cultures, were used as controls. The following concentrations were used: haloperidol--0.1 mkg/ml, 10 mkg/ml and 10.0 mkg/ml; mageptyl--2.0 mkg/ml, 10,0 mkg/ml and 50 mkg/ml. There was a significant inhibition of the mitotic and blast transformation index after using concentrations of 10.0 mkg/ml both of haloperidol and magentyl. Concentration of 50.0 mkg/ml was toxic for the lymphocytes, obtaine from all donors. Furthermore the two preparations did not induce genomic mutations. Polyploidia, established in the treated cultures, varied slightly from that of the nontreated cultures. There was a weak mutagenic activity of the two preparations at chromosomal level in a concentration of 10.0 mkg/ml. | [Comparative study of the effect of haloperidol and maptil on the karyotype and immunoproliferative response of lymphocytes]. The author examined the action of haloperidol and magetyl on the mitotic activity, blast transformation and karyotype of the lymphocytic cultures, obtained from the peripheral blood in vitro. The experiments were carried out on flymphycytes, obtained from ten healthy individuals. Nontreated cultures, obtained from the same cultures, were used as controls. The following concentrations were used: haloperidol--0.1 mkg/ml, 10 mkg/ml and 10.0 mkg/ml; mageptyl--2.0 mkg/ml, 10,0 mkg/ml and 50 mkg/ml. There was a significant inhibition of the mitotic and blast transformation index after using concentrations of 10.0 mkg/ml both of haloperidol and magentyl. Concentration of 50.0 mkg/ml was toxic for the lymphocytes, obtaine from all donors. Furthermore the two preparations did not induce genomic mutations. Polyploidia, established in the treated cultures, varied slightly from that of the nontreated cultures. There was a weak mutagenic activity of the two preparations at chromosomal level in a concentration of 10.0 mkg/ml. | [
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|
PMID:14821 | Uptake of NADPH by islet secretion granule membranes. | Reduced nicotinamide adenine dinucleotide phosphate (NADPH), which stimulated the release of insulin from toadfish islet cells and from isolated secretion granules, was taken up by the membranes prepare from these secretion granules. Oxidized nicotinamide adenine dinucleotide phosphate, which did not release insulin from the granules, was taken up to a much lesser extent. The uptake of NADPH by the granule membranes was time dependent, reaching equilibrium in about 30 min. The maximum amount of NADPH taken up was 0.6 nmol/mg membrane protein and the concentration of NADPH needed to obtain maximum uptake was 6.5 x 10(-4)m, which was approximately the same concentration of NADPH needed to produce maximum release of insulin from the secretion granules. | Uptake of NADPH by islet secretion granule membranes. Reduced nicotinamide adenine dinucleotide phosphate (NADPH), which stimulated the release of insulin from toadfish islet cells and from isolated secretion granules, was taken up by the membranes prepare from these secretion granules. Oxidized nicotinamide adenine dinucleotide phosphate, which did not release insulin from the granules, was taken up to a much lesser extent. The uptake of NADPH by the granule membranes was time dependent, reaching equilibrium in about 30 min. The maximum amount of NADPH taken up was 0.6 nmol/mg membrane protein and the concentration of NADPH needed to obtain maximum uptake was 6.5 x 10(-4)m, which was approximately the same concentration of NADPH needed to produce maximum release of insulin from the secretion granules. | [
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|
PMID:14822 | Failure of neurotransmitter blockers to alter PGE2-induced LH release. | The purpose of this study is to ascertain whether or not prostaglandin (PG) E2 induces LH release by modifying or mudulating the release or action of neural transmitters. PGE2 injected inv into spayed rats primed two days earlier with 10 mug estradiol benzoate increased the plasma levels of LH 10 min later as measured by radio-immunoassay. The peak of plasma LH was not changed by prior treatment with beta- or alpha-adrenergic receptor blockers, propranolol or phenoxybenzamine. The peak level of plasma LH did not alter in rats treated with DL-alpha-methyl-ptyrosine methyl ester HC1 (alpha-MPT) or sodium diethyldithiocarbamate (DDC). Similarly, the peak of plasma LH was not changed by prior treatment with imipramine. Adminisration of PGE2 produced an increase in anterior pituitary and plasma, but not hypothalamic cyclic AMP concomitantly with the elevation in plasma LH. Although it is possible that the effect of PGE2 could be mediated by another transmitter system, as yet unknown, or that the effect of PGE2 on LH release could be mediated via the adenylate cyclase-cyclic AMP system, the results indicate that PGE2 does not act trans-synaptically, but probably acts directly on LH-RH neurons. | Failure of neurotransmitter blockers to alter PGE2-induced LH release. The purpose of this study is to ascertain whether or not prostaglandin (PG) E2 induces LH release by modifying or mudulating the release or action of neural transmitters. PGE2 injected inv into spayed rats primed two days earlier with 10 mug estradiol benzoate increased the plasma levels of LH 10 min later as measured by radio-immunoassay. The peak of plasma LH was not changed by prior treatment with beta- or alpha-adrenergic receptor blockers, propranolol or phenoxybenzamine. The peak level of plasma LH did not alter in rats treated with DL-alpha-methyl-ptyrosine methyl ester HC1 (alpha-MPT) or sodium diethyldithiocarbamate (DDC). Similarly, the peak of plasma LH was not changed by prior treatment with imipramine. Adminisration of PGE2 produced an increase in anterior pituitary and plasma, but not hypothalamic cyclic AMP concomitantly with the elevation in plasma LH. Although it is possible that the effect of PGE2 could be mediated by another transmitter system, as yet unknown, or that the effect of PGE2 on LH release could be mediated via the adenylate cyclase-cyclic AMP system, the results indicate that PGE2 does not act trans-synaptically, but probably acts directly on LH-RH neurons. | [
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|
PMID:14825 | Purification of an endonuclease from adenovirus-infected KB cells. | This report describes the purification of an endonuclease from extracts of adenovirus-type-2-infected KB cells. Endonuclease activity can also be detected in extracts of uninfected KB cells and the enzyme activities from extracts of uninfected and adenovirus-infected cells are very similar, if not identical. The enzyme has its maximal activity at pH 4.0. The enzyme found in uninfected and adenovirus-infectedcells is, however, strikingly different from an endonuclease isolated from calf serum. Hence, the endonuclease described is probably not a contaminant derived from the medium in which the KB cells were propagated. The endonuclease in crude extracts from uninfected or adenovirus-infected KB cells can be activated or its activity enhanced by treatment of the extracts with proteolytic enzymes, like pronase or trypsin. Evidence has been presented suggesting that this activation is due to proteolytic cleavage of an inhibitor present in crude extracts of uninfected and adenovirus-type-2-infected KB cells. A second endonuclease has been found in extracts of infected and uninfected cells with optimal activity at pH 7.2 and this endonuclease can be separated from the one with a pH optimum at 4.0. | Purification of an endonuclease from adenovirus-infected KB cells. This report describes the purification of an endonuclease from extracts of adenovirus-type-2-infected KB cells. Endonuclease activity can also be detected in extracts of uninfected KB cells and the enzyme activities from extracts of uninfected and adenovirus-infected cells are very similar, if not identical. The enzyme has its maximal activity at pH 4.0. The enzyme found in uninfected and adenovirus-infectedcells is, however, strikingly different from an endonuclease isolated from calf serum. Hence, the endonuclease described is probably not a contaminant derived from the medium in which the KB cells were propagated. The endonuclease in crude extracts from uninfected or adenovirus-infected KB cells can be activated or its activity enhanced by treatment of the extracts with proteolytic enzymes, like pronase or trypsin. Evidence has been presented suggesting that this activation is due to proteolytic cleavage of an inhibitor present in crude extracts of uninfected and adenovirus-type-2-infected KB cells. A second endonuclease has been found in extracts of infected and uninfected cells with optimal activity at pH 7.2 and this endonuclease can be separated from the one with a pH optimum at 4.0. | [
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|
PMID:14826 | Characterization of the pH 4.0 endonuclease from adenovirus-type-2-infected KB cells. | The properties of the pH 4.0 endonuclease from adenovirus-type-2-infected KB cells were determined. The enzyme has a molecular weight of approximately 40000. Its pH optimum is at pH 4.0, it is not inhibited by ethylenediaminetetraacetate (EDTA), and it is active at temperatures up to 60 degree C. The enzyme cleaves adenovirus DNA in a stepwise manner. The limit digestion product has a molecular weight of 120000-200000. There is evidence that the cleavage reaction proceeds via an initial single-strand nick. Under the conditions tested the endonuclease did not seem to reveal a high degree of specificity as to the recognition of cleavage sites, or else the sites recognized occurred very frequently. | Characterization of the pH 4.0 endonuclease from adenovirus-type-2-infected KB cells. The properties of the pH 4.0 endonuclease from adenovirus-type-2-infected KB cells were determined. The enzyme has a molecular weight of approximately 40000. Its pH optimum is at pH 4.0, it is not inhibited by ethylenediaminetetraacetate (EDTA), and it is active at temperatures up to 60 degree C. The enzyme cleaves adenovirus DNA in a stepwise manner. The limit digestion product has a molecular weight of 120000-200000. There is evidence that the cleavage reaction proceeds via an initial single-strand nick. Under the conditions tested the endonuclease did not seem to reveal a high degree of specificity as to the recognition of cleavage sites, or else the sites recognized occurred very frequently. | [
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|
PMID:14827 | Affinity labeling of rat-kidney gamma-glutamyl transpeptidase. | The reaction of gamma-glutamyl transpeptidase from rat kidney with a glutamine analog, 6-diazo-5-oxo-L-norleucine, resulted in irreversible inactivation of the enzyme. The concentration of this reagent giving a half-maximum rate of inactivation was 6 mMat pH 7.5. The inactivation was prevented by the presence of reduced glutathione in a competitive fashion, which indicates the active-site-directed nature of this reagent. The rate of inactivation was greatly accelerated in the presence of maleate, which is known to enhance the glutaminase activity of this enzyme. The presence of maleate increased the maximum velocity of the inactivation, but did not affect the affinity of the enzyme for 6-diazo-5-oxo-L-norleucine. Inactivation of the enzyme with 6-diazo-5-oxo-L-[6=14C]norleucine as well as with 6-diazo-5-oxo-L[1,2,3,4,5-14C]norleucine resulted in a stoichiometric incorporation of radioactivity into the enzyme protein via covalent linkage. The amount of radioactivity incorporated was 1 mol 14C label/248000 g enzyme protein. A native enzyme preparation showing a single protein band on polyacrylamide gel electrophoresis gave four distinct bands upon sodium dodecylsulfate/polyacrylamide gel electrophoresis. Upon sodium dodecylsulfate/polyacrylamide gel electrophoresis of the 14C-labeled enzyme, only the band moving the fastest towards the anode was found to contain radioactivity. This finding indicates that this protein band represents the catalytic component of the enzyme. | Affinity labeling of rat-kidney gamma-glutamyl transpeptidase. The reaction of gamma-glutamyl transpeptidase from rat kidney with a glutamine analog, 6-diazo-5-oxo-L-norleucine, resulted in irreversible inactivation of the enzyme. The concentration of this reagent giving a half-maximum rate of inactivation was 6 mMat pH 7.5. The inactivation was prevented by the presence of reduced glutathione in a competitive fashion, which indicates the active-site-directed nature of this reagent. The rate of inactivation was greatly accelerated in the presence of maleate, which is known to enhance the glutaminase activity of this enzyme. The presence of maleate increased the maximum velocity of the inactivation, but did not affect the affinity of the enzyme for 6-diazo-5-oxo-L-norleucine. Inactivation of the enzyme with 6-diazo-5-oxo-L-[6=14C]norleucine as well as with 6-diazo-5-oxo-L[1,2,3,4,5-14C]norleucine resulted in a stoichiometric incorporation of radioactivity into the enzyme protein via covalent linkage. The amount of radioactivity incorporated was 1 mol 14C label/248000 g enzyme protein. A native enzyme preparation showing a single protein band on polyacrylamide gel electrophoresis gave four distinct bands upon sodium dodecylsulfate/polyacrylamide gel electrophoresis. Upon sodium dodecylsulfate/polyacrylamide gel electrophoresis of the 14C-labeled enzyme, only the band moving the fastest towards the anode was found to contain radioactivity. This finding indicates that this protein band represents the catalytic component of the enzyme. | [
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|
PMID:14828 | Superoxide dismutase from Thermus aquaticus. Isolation and characterisation of manganese and apo enzymes. | Superoxide dismutase has been isolated and characterised from the extreme thermophile Thermus aquaticus. The pure enzyme is a reddish-purple manganese-containing protein with a molecular weight of approximately 80000 +/- 5000. Combination of gel electrophoresis in dodecylsulphate and amino acid analysis shows that it is composed of four identical subunit polypeptide chains consisting of approximately 186 amino acids. The tetrameric protein contains two atoms of manganese. A stable manganese-free apoprotein has been prepared by treatment with EDTA in 8 M urea at acidic pH. The apoprotein regains the tetrameric structure in the absence of manganese but is inactive. Reconstitution of active Mn-enzyme was achieved byaddition of Mm2+ apoprotein in 8 M urea at acid pH. Reconstitution was monitored by absorption spectroscopy, manganese analysis and regain of activity and by these criteria the reconstituted enyzme with two atoms Mn per mole is indistinguishable from the native enzyme. The enhanced stability of the thermophile apoenzyme and Mn-enzyme is of advantage for studies of the structure and mechanism of action of superoxide dismutase. The N-terminal amino acid sequence to the 40th residue of the submit was determined by automated Edman degradation. The sequence has a close resemblance to that of the dimeric Mn-enzyme from another thermophile, Bacillus stearothermophilus. | Superoxide dismutase from Thermus aquaticus. Isolation and characterisation of manganese and apo enzymes. Superoxide dismutase has been isolated and characterised from the extreme thermophile Thermus aquaticus. The pure enzyme is a reddish-purple manganese-containing protein with a molecular weight of approximately 80000 +/- 5000. Combination of gel electrophoresis in dodecylsulphate and amino acid analysis shows that it is composed of four identical subunit polypeptide chains consisting of approximately 186 amino acids. The tetrameric protein contains two atoms of manganese. A stable manganese-free apoprotein has been prepared by treatment with EDTA in 8 M urea at acidic pH. The apoprotein regains the tetrameric structure in the absence of manganese but is inactive. Reconstitution of active Mn-enzyme was achieved byaddition of Mm2+ apoprotein in 8 M urea at acid pH. Reconstitution was monitored by absorption spectroscopy, manganese analysis and regain of activity and by these criteria the reconstituted enyzme with two atoms Mn per mole is indistinguishable from the native enzyme. The enhanced stability of the thermophile apoenzyme and Mn-enzyme is of advantage for studies of the structure and mechanism of action of superoxide dismutase. The N-terminal amino acid sequence to the 40th residue of the submit was determined by automated Edman degradation. The sequence has a close resemblance to that of the dimeric Mn-enzyme from another thermophile, Bacillus stearothermophilus. | [
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|
PMID:14829 | PH-dependent changes of 2,3-bisphosphoglycerate in human red cells during transitional and steady states in vitro. | A systematic study of the pH-dependent changes in the range 6.6--7.4 of 2,3-bisphosphoglycerate and the adenine nucleotides was performed in the presence and absence of glucose during transitional and steady states. 1. The results indicatethat 2,3-gisphosphoglycerate phosphatase breaks down 2,3-bisphosphoglycerate nearly independent of pH at a rate of 480 mumol 2,3-bisphosphoglycerate x1 cells-1xh-1.2,3-Bisphosphoglycerate mutase is practically completely inhibited below pH value increases in long-term experiments with lower 2,3-bisphosphoglycerate levels. The formation of pyruvate corresponds to the breakdown of 2,3-bisphosphoglycerate afterconsumption of an unknown reducing substance. | PH-dependent changes of 2,3-bisphosphoglycerate in human red cells during transitional and steady states in vitro. A systematic study of the pH-dependent changes in the range 6.6--7.4 of 2,3-bisphosphoglycerate and the adenine nucleotides was performed in the presence and absence of glucose during transitional and steady states. 1. The results indicatethat 2,3-gisphosphoglycerate phosphatase breaks down 2,3-bisphosphoglycerate nearly independent of pH at a rate of 480 mumol 2,3-bisphosphoglycerate x1 cells-1xh-1.2,3-Bisphosphoglycerate mutase is practically completely inhibited below pH value increases in long-term experiments with lower 2,3-bisphosphoglycerate levels. The formation of pyruvate corresponds to the breakdown of 2,3-bisphosphoglycerate afterconsumption of an unknown reducing substance. | [
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|
PMID:14830 | Carboxyl groups at the two active centers of sucrase-isomaltase from rabbit small intestine. | 1. Seveal selective reagents were employed to identify the amino acid residues essential for the catalytic activity of sucrase-isomaltase. 2. Modification of histidine, lysine and carboxyl residues resulted in a partial inactivation of the enzyme. Substrates or competitive inhibitors provided protection against inactivation only in the reaction of carboxyl groups with carbodiimide (+lycine ethyl ester) or with diazoacetic ethyl ester. This indicated the occurrence of carboxyl groups at the two active centers of the enzyme complex. 3. Protection against inactivation of the enzyme by carbodiimide was provided also by the presence of alkali and alkaline earth metal ions, which are non-essential activators of sucrase-isomaltase. The presence of Na+ and Ba2+ protected approximately one carboxyl group per active center from reacting with carbodiimide plus glycine ethyl ester. 4. The carbodiimide-reactive groups were not identical with the two carboxylate groups recently found to react with conduritol-B-epoxide, an active-site-directed inhibitor of sucrase-isomaltase (Quaroni, A. and Semenza, G., 1976, J. Biol. Chem 251,3250--3253). A possible role for the carbodiimide-reactive carboxyl groups at the active centers of sucrase-isomaltase is discussed. | Carboxyl groups at the two active centers of sucrase-isomaltase from rabbit small intestine. 1. Seveal selective reagents were employed to identify the amino acid residues essential for the catalytic activity of sucrase-isomaltase. 2. Modification of histidine, lysine and carboxyl residues resulted in a partial inactivation of the enzyme. Substrates or competitive inhibitors provided protection against inactivation only in the reaction of carboxyl groups with carbodiimide (+lycine ethyl ester) or with diazoacetic ethyl ester. This indicated the occurrence of carboxyl groups at the two active centers of the enzyme complex. 3. Protection against inactivation of the enzyme by carbodiimide was provided also by the presence of alkali and alkaline earth metal ions, which are non-essential activators of sucrase-isomaltase. The presence of Na+ and Ba2+ protected approximately one carboxyl group per active center from reacting with carbodiimide plus glycine ethyl ester. 4. The carbodiimide-reactive groups were not identical with the two carboxylate groups recently found to react with conduritol-B-epoxide, an active-site-directed inhibitor of sucrase-isomaltase (Quaroni, A. and Semenza, G., 1976, J. Biol. Chem 251,3250--3253). A possible role for the carbodiimide-reactive carboxyl groups at the active centers of sucrase-isomaltase is discussed. | [
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|
PMID:14831 | A mutant ATP synthetase of Escherichia coli with an altered sensitivity to N,N' -dicyclohexylcarbodiimide: characterization in native membranes and reconstituted proteoliposomes. | Dicyclohexylcarbodiimide-resistant mutants of Escherichia coli were isolated and characterized In one mutant the unc genes and affects the membrane-integrated part of the ATP synthetase. The sensitivity of ATP synthetase functions to N,N' -dicyclohexylcarbodiimide was compared in wild-type and mutant membranes. The membrane-integrated part of the wild-type ATP synthetase is highly sensitive to ATP-dependent membrane energization and restoration of lactate-dependent energization of ATPase-depleted membranes. In mutant membranes this concentration has only a slight effect on these activities whereas a severe inhibition is obtained at 200 muM. Using the highly water-soluble 1-ethyl-3(3-dimethylaminopropyl)-carbodiimide theactivities of wild-type and mutant membranes are inhibited to the same extent. TheATP synthetase of wild-type and mutant was partially purified and incorporated muM. Uinto liposomes. These showed an uncoupler-sensitive ATP-32Pi exchange and ATP-dependent quenching of acridine-dye fluorescence. The activities of mutant and wild-type proteoliposomes exhibit the same pattern of sensitivity to dicyclohexylcarbodiimide as the corresponding membranes. | A mutant ATP synthetase of Escherichia coli with an altered sensitivity to N,N' -dicyclohexylcarbodiimide: characterization in native membranes and reconstituted proteoliposomes. Dicyclohexylcarbodiimide-resistant mutants of Escherichia coli were isolated and characterized In one mutant the unc genes and affects the membrane-integrated part of the ATP synthetase. The sensitivity of ATP synthetase functions to N,N' -dicyclohexylcarbodiimide was compared in wild-type and mutant membranes. The membrane-integrated part of the wild-type ATP synthetase is highly sensitive to ATP-dependent membrane energization and restoration of lactate-dependent energization of ATPase-depleted membranes. In mutant membranes this concentration has only a slight effect on these activities whereas a severe inhibition is obtained at 200 muM. Using the highly water-soluble 1-ethyl-3(3-dimethylaminopropyl)-carbodiimide theactivities of wild-type and mutant membranes are inhibited to the same extent. TheATP synthetase of wild-type and mutant was partially purified and incorporated muM. Uinto liposomes. These showed an uncoupler-sensitive ATP-32Pi exchange and ATP-dependent quenching of acridine-dye fluorescence. The activities of mutant and wild-type proteoliposomes exhibit the same pattern of sensitivity to dicyclohexylcarbodiimide as the corresponding membranes. | [
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|
PMID:14832 | Proton movements coupled to sugar transport via the galactose transport system in Salmonella typhimurium. | We have studied proton movements associated with substrate transport via the galactose transport system in Salmonella typhimurium. The addition of galactose to lightly buffered suspensions of anaerobic, non-metabolizing cells of Salmonella typhimurium, specifically induced for the galactose transport system, causes an increase in extracellularpH as galactose and protons enter the cell together. Other substrates for this transport system, D-fucose, 2-deoxygalactose, glucose and 2-deoxyglucose similarly cause an influx of protons when transported. In contrast, transport via the other major transport system for galactose, the methylgalactoside transport system, is not coupled to H+ influx. Comparison of kinetic data obtained from pH measurements with data obtained from measurement of active transport of galactose via the galactose transport system suggests that the apparent Km of the galactose transport system for this sugar differs under energized and non-energized conditions. At pH 7.2 the permeant anion SCN- increases both the rate and extent of galactose-induced proton influx; at pH 6 the rate, but not the extent is increased by SCN-. | Proton movements coupled to sugar transport via the galactose transport system in Salmonella typhimurium. We have studied proton movements associated with substrate transport via the galactose transport system in Salmonella typhimurium. The addition of galactose to lightly buffered suspensions of anaerobic, non-metabolizing cells of Salmonella typhimurium, specifically induced for the galactose transport system, causes an increase in extracellularpH as galactose and protons enter the cell together. Other substrates for this transport system, D-fucose, 2-deoxygalactose, glucose and 2-deoxyglucose similarly cause an influx of protons when transported. In contrast, transport via the other major transport system for galactose, the methylgalactoside transport system, is not coupled to H+ influx. Comparison of kinetic data obtained from pH measurements with data obtained from measurement of active transport of galactose via the galactose transport system suggests that the apparent Km of the galactose transport system for this sugar differs under energized and non-energized conditions. At pH 7.2 the permeant anion SCN- increases both the rate and extent of galactose-induced proton influx; at pH 6 the rate, but not the extent is increased by SCN-. | [
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|
PMID:14833 | Intramolecularly-quenched fluorescent peptides as fluorogenic substrates ofleucine aminopeptidase and inhibitors of clostridial aminopeptidase. | Fluorogenic oligopeptide derivatives of the type Lys(ABz)-ONBzl, where ABz iso-aminobenzoyl (anthraniloyl), X stands for Ala Phe, or Ala-Ala, and ONBzlis p-nitrobenzyloxy, were synthesized and shown to be hydrolyzed by leucine aminopeptidase. The hydrolysis is accompanied by an increase in fluorescence due to disruptionof the intramolecular quenching of the fluorescent anthraniloyl moiety by the nitrobenzyester group. The spectral characteristics of the compounds are not consistent withan energy transfer mechanism according to Förster, therefore the quenching isassumed to be caused by a direct encouter between the quenching and the fluorecentgroups. The change in fluorescence that accompanies the enzymic hydrolysis ofthe first peptide bound was used for quantitative measurement of the activity ofthe activity of leucine aminopeptidase and for the determination of some of itskinetic parameters. A bacterial aminopeptidase from Clostrdium histolyticumthat is very similar to leucine aminopeptidase in its substrate specificity inits substrate specificity did not hydrolyze the above peptidederivatives. Thehydrolysis of leucine p-nitroanilide by this enzyme was found to be inhibitedby the three peptides and the corresponding inhibition constants were determined. | Intramolecularly-quenched fluorescent peptides as fluorogenic substrates ofleucine aminopeptidase and inhibitors of clostridial aminopeptidase. Fluorogenic oligopeptide derivatives of the type Lys(ABz)-ONBzl, where ABz iso-aminobenzoyl (anthraniloyl), X stands for Ala Phe, or Ala-Ala, and ONBzlis p-nitrobenzyloxy, were synthesized and shown to be hydrolyzed by leucine aminopeptidase. The hydrolysis is accompanied by an increase in fluorescence due to disruptionof the intramolecular quenching of the fluorescent anthraniloyl moiety by the nitrobenzyester group. The spectral characteristics of the compounds are not consistent withan energy transfer mechanism according to Förster, therefore the quenching isassumed to be caused by a direct encouter between the quenching and the fluorecentgroups. The change in fluorescence that accompanies the enzymic hydrolysis ofthe first peptide bound was used for quantitative measurement of the activity ofthe activity of leucine aminopeptidase and for the determination of some of itskinetic parameters. A bacterial aminopeptidase from Clostrdium histolyticumthat is very similar to leucine aminopeptidase in its substrate specificity inits substrate specificity did not hydrolyze the above peptidederivatives. Thehydrolysis of leucine p-nitroanilide by this enzyme was found to be inhibitedby the three peptides and the corresponding inhibition constants were determined. | [
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|
PMID:14834 | Hypoxanthine levels of plasma during hypoxemia in dogs. | Tissue hypoxia was induced in one group of dogs by clamping of the endotracheal tube and in another group by artificial ventilation with a mixture of nitrogen and air. The hypoxanthine concentration of venous and arterial plasma increased significantly during severe hypoxemia. When the hypoxemia was relieved, an increased venous-arterial hypoxanthine difference appeared indicating that the lung metabolism of hypoxanthine was slowed down during alveolar hypoxia. It is concluded that the level of plasma hypoxanthine in dogs during hypoxemia is dependent on the degree of tissue hypoxia, peripheral vasoregulation, and lung metabolism. | Hypoxanthine levels of plasma during hypoxemia in dogs. Tissue hypoxia was induced in one group of dogs by clamping of the endotracheal tube and in another group by artificial ventilation with a mixture of nitrogen and air. The hypoxanthine concentration of venous and arterial plasma increased significantly during severe hypoxemia. When the hypoxemia was relieved, an increased venous-arterial hypoxanthine difference appeared indicating that the lung metabolism of hypoxanthine was slowed down during alveolar hypoxia. It is concluded that the level of plasma hypoxanthine in dogs during hypoxemia is dependent on the degree of tissue hypoxia, peripheral vasoregulation, and lung metabolism. | [
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|
PMID:14835 | Protective action of timolol in acute myocardial ischemia. | Timolol, a beta-adrenoceptor blocking agent with little or no cardiodepressant activity, was studied in acute myocardial ischemia in cats. Timolol, at a dose of 25 mug/kg, blocked 75 to 80% of the cardiac response to isoproterenol. This dose also significantly reduced heart rate in cats subjected to acute myocardial ischemia by ligation of the left coronary artery. Timolol significantly prevented the spread of ischemic damage in the myocardium as assessed by (a) curtailing the increase in plasma creatine phosphokinase (CPK) activity, (b) preventing the loss of CPK from the ischemic portion of the myocardium, and (c) restoring the elevated S-T segment of the electrocardiogram toward normal. Timolol did not significantly retard the increase in fragility of lysosomes in ischemic myocardial tissue. The mechanism of the protective effect to timolol on the ischemic myocardium appears to be via reducing myocardial oxygen demand by decreasing heart rate. | Protective action of timolol in acute myocardial ischemia. Timolol, a beta-adrenoceptor blocking agent with little or no cardiodepressant activity, was studied in acute myocardial ischemia in cats. Timolol, at a dose of 25 mug/kg, blocked 75 to 80% of the cardiac response to isoproterenol. This dose also significantly reduced heart rate in cats subjected to acute myocardial ischemia by ligation of the left coronary artery. Timolol significantly prevented the spread of ischemic damage in the myocardium as assessed by (a) curtailing the increase in plasma creatine phosphokinase (CPK) activity, (b) preventing the loss of CPK from the ischemic portion of the myocardium, and (c) restoring the elevated S-T segment of the electrocardiogram toward normal. Timolol did not significantly retard the increase in fragility of lysosomes in ischemic myocardial tissue. The mechanism of the protective effect to timolol on the ischemic myocardium appears to be via reducing myocardial oxygen demand by decreasing heart rate. | [
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|
PMID:14836 | Studies on the antagonism by raphe lesions of the antinociceptive action of systemic morphine. | In rats, lesions were placed in the dorsal/median raphe (DMR), in the ventral raphe (VR: raphe magnus), in both the dorsal/median and ventral raphe (DMVR) or in the reticular formation (RF). The effect of the lesions on the antinociception and catalepsy produced by 3 doses of morphine (3, 10 and 30 mg/kg) was examined. The lesions had no significant effect on the catalepsy produced by any of the doses of morphine tested. DMR lesions produced a partial attentuation of the antinociceptive action of both the 3 and 10 mg doses. VR lesions produced a complete blockade of the 3 mg and only a partial attenuation of the 10 mg dose. In contrast, the combined (DMVR) lesions yielded virtually a total blockade of the 3 and 10 mg. Yet, as with the DMR and VR groups, the DMVR lesions failed to produce a significant antagonism on either of the nociceptive tests at the 30 mg dose. These findings suggest that the ascending and descending fiber systems emanating from the dorsal/median and ventral raphe, respectively, facilitate the expression of morphine-induced analgesia but that neither system alone can be regarded as essential for the manifestation of the antinociceptive effects of systematically administered morphine. | Studies on the antagonism by raphe lesions of the antinociceptive action of systemic morphine. In rats, lesions were placed in the dorsal/median raphe (DMR), in the ventral raphe (VR: raphe magnus), in both the dorsal/median and ventral raphe (DMVR) or in the reticular formation (RF). The effect of the lesions on the antinociception and catalepsy produced by 3 doses of morphine (3, 10 and 30 mg/kg) was examined. The lesions had no significant effect on the catalepsy produced by any of the doses of morphine tested. DMR lesions produced a partial attentuation of the antinociceptive action of both the 3 and 10 mg doses. VR lesions produced a complete blockade of the 3 mg and only a partial attenuation of the 10 mg dose. In contrast, the combined (DMVR) lesions yielded virtually a total blockade of the 3 and 10 mg. Yet, as with the DMR and VR groups, the DMVR lesions failed to produce a significant antagonism on either of the nociceptive tests at the 30 mg dose. These findings suggest that the ascending and descending fiber systems emanating from the dorsal/median and ventral raphe, respectively, facilitate the expression of morphine-induced analgesia but that neither system alone can be regarded as essential for the manifestation of the antinociceptive effects of systematically administered morphine. | [
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|
PMID:14837 | Attenuation of morphine analgesia in rats with lesions of the locus coeruleus and dorsal raphe nucleus. | The nociceptive reflex activity and analgesic effect of morphine were studied in rats using the hind paw stimulation test. The stimulation threshold was significantly increased in animals with bilateral destruction of the locus coeruleus (LC), and was reduced after lesion of the dorsal raphe nucleus (DR). LC lesions produced a selective lowering of noradrenaline (NA) content in the forebrain, while DR lesions resulted in a reduction in serotonin levels. Lesioning both LC and DR significantly reduced both NA and serotonin contents even when the stimulation threshold was not altered. Morphine produced a significant and dose-dependent elevation of the stimulation threshold in sham-operated animals, while morphine analgesia was almost completely inhibited by destruction of LC, DR and both the nuclei. These results imply that a depression of LC-mediated noradrenergic tone results in a decreased sensitivity to painful stimuli, whereas a reduction of raphe-derived serotonergic tone produces the opposite effect against LC. It is suggested, however, that both of these monoamines from the LC and DR are necessary for the analgesic effect of morphine. | Attenuation of morphine analgesia in rats with lesions of the locus coeruleus and dorsal raphe nucleus. The nociceptive reflex activity and analgesic effect of morphine were studied in rats using the hind paw stimulation test. The stimulation threshold was significantly increased in animals with bilateral destruction of the locus coeruleus (LC), and was reduced after lesion of the dorsal raphe nucleus (DR). LC lesions produced a selective lowering of noradrenaline (NA) content in the forebrain, while DR lesions resulted in a reduction in serotonin levels. Lesioning both LC and DR significantly reduced both NA and serotonin contents even when the stimulation threshold was not altered. Morphine produced a significant and dose-dependent elevation of the stimulation threshold in sham-operated animals, while morphine analgesia was almost completely inhibited by destruction of LC, DR and both the nuclei. These results imply that a depression of LC-mediated noradrenergic tone results in a decreased sensitivity to painful stimuli, whereas a reduction of raphe-derived serotonergic tone produces the opposite effect against LC. It is suggested, however, that both of these monoamines from the LC and DR are necessary for the analgesic effect of morphine. | [
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|
PMID:14838 | Pharmacological characterisation of the presynaptic alpha-adrenoceptor in the rat vas deferens. | The interactions between alpha-adrenoceptor agonists and antagonists on the twitch response of the rat isolated vas deferens to low frequency motor nerve stimulatioh have been examined. Oxymetazoline, clonidine, naphazoline and BAY-1470 caused a concentration-dependent inhibition of the twitch response at concentrations lower than those causing smooth muscle stimulation. The twitch-inhibitory effect of these compounds is thought to result from stimulation of presynaptically located alpha-adrenoceptors. In contrast, phenylephrine and methoxamine exerted little inhibitory effect at concentrations less than those which produced postsynaptic stimulation. The inhibitory effect of clonidine was antagonised by phentolamine, piperoxan, yohimbine and tolazoline, but not by thymoxamine. These results characterise the presynaptic alpha-adrenoceptors in the rat vas deferens as being of the same type as those present in the rabbit pulmonary artery and rat heart, but different from those located post-synaptically in sympathetically innervated tissues. | Pharmacological characterisation of the presynaptic alpha-adrenoceptor in the rat vas deferens. The interactions between alpha-adrenoceptor agonists and antagonists on the twitch response of the rat isolated vas deferens to low frequency motor nerve stimulatioh have been examined. Oxymetazoline, clonidine, naphazoline and BAY-1470 caused a concentration-dependent inhibition of the twitch response at concentrations lower than those causing smooth muscle stimulation. The twitch-inhibitory effect of these compounds is thought to result from stimulation of presynaptically located alpha-adrenoceptors. In contrast, phenylephrine and methoxamine exerted little inhibitory effect at concentrations less than those which produced postsynaptic stimulation. The inhibitory effect of clonidine was antagonised by phentolamine, piperoxan, yohimbine and tolazoline, but not by thymoxamine. These results characterise the presynaptic alpha-adrenoceptors in the rat vas deferens as being of the same type as those present in the rabbit pulmonary artery and rat heart, but different from those located post-synaptically in sympathetically innervated tissues. | [
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|
PMID:14839 | The effect of guanethidine and hydrochlorothiazide on blood pressure and vascular tyrosine hydroxylase activity in the spontaneously hypertensive rat. | The present study examined the effects of antihypertensive drugs (hydrochlorothiazide and guanethidine) on blood pressure and tyrosine hydroxylase (TH) activity in the spontaneously hypertensive rat (SHR). Hydrochlorothiazide (50 mg/kg X 4 days) lowered blood pressure in the SHR to a degree equivalent to that produced by reserpine (0.3 mg/kg X 3 days). However, while reserpine increased vascular and adrenal TH activity, hydrochlorothiazide had no effect. Guanethidine (30 mg/kg X 2 days) reduced blood pressure in the SHR and also depleted cardiac, vascular and adrenal gland catecholamines; However, guanethidine administration did not increase TH activity in the mesenteric vasculature or adrenal glands. These studies indicate that at equieffective blood pressure lowering doses, different antihypertensive drugs have different effects on TH activity in the SHR. Neither blood pressure reduction nor catecholamine depletion in peripheral tissues are sufficient prerequisties for increasing TH activity. The data support the suggestion, however, that amine depletion in the central nervous system or ganglia may be an important factor in the regulation of TH. | The effect of guanethidine and hydrochlorothiazide on blood pressure and vascular tyrosine hydroxylase activity in the spontaneously hypertensive rat. The present study examined the effects of antihypertensive drugs (hydrochlorothiazide and guanethidine) on blood pressure and tyrosine hydroxylase (TH) activity in the spontaneously hypertensive rat (SHR). Hydrochlorothiazide (50 mg/kg X 4 days) lowered blood pressure in the SHR to a degree equivalent to that produced by reserpine (0.3 mg/kg X 3 days). However, while reserpine increased vascular and adrenal TH activity, hydrochlorothiazide had no effect. Guanethidine (30 mg/kg X 2 days) reduced blood pressure in the SHR and also depleted cardiac, vascular and adrenal gland catecholamines; However, guanethidine administration did not increase TH activity in the mesenteric vasculature or adrenal glands. These studies indicate that at equieffective blood pressure lowering doses, different antihypertensive drugs have different effects on TH activity in the SHR. Neither blood pressure reduction nor catecholamine depletion in peripheral tissues are sufficient prerequisties for increasing TH activity. The data support the suggestion, however, that amine depletion in the central nervous system or ganglia may be an important factor in the regulation of TH. | [
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|
PMID:14843 | [Endogenous formation of carcinogenic-N-nitroso compounds in rats after application of drugs and nitrite (author's transl)]. | During the last years in vitro investigations have demonstrated the possibility of nitrosation of different amines and amides including a number of drugs. The aim of the presented investigations was to test whether an endogenous nitrosation of piperazine, Obesin (propyhexedrine), and ephedrine is possible in long term experiments with rats. With the exception of group No. 3, the experiments were carried out with hooded rats (table 1). Group No. 1: 15 rats were given 100 mg/kg b.w. piperazine (Merck, Darmstadt) and 80 mg/kg b.w. NaNO2 by gastric tube. After the fourth application because of intoxication a reduction to 80 mg/kg b.w. piperazine and 50 mg/kg b.w. NaNO2 was necessary. Duration of treatment (in two weeks intervals): 7 months. Group No. 2: Pregnant rats received 80 mg/kg b.w. piperazine and 50 mg/kg b.w. NaNO2 by gastric tube once in the last third of pregnancy. In 13 offsprings the carcinogenic action was tested. Group No. 3: Pregnant Wistar rats were given 100 mg/kg b.w. piperazine and 70 mg/kg b.w. NaN92 by gastric tube once in the last third of pregnancy. Group No. 4: 20 hooded rats received 40 mg/kg b.w. Obesin (propylhexedrine, Fahlberg-List Magdeburg) and 40 mg/kg b.w. NaM92 once in two weekly intervals by gastric tube. Duration of the treatment: 6 months. 5 controls received 40 mg/kg b.w. Obesin or 40 mg/kg b.w. NaNO2 resp. Group No. 5: Hooded rats were given 40 mg/kg b.w. Obesin and 40 mg/kg b.w. NaNO2 by gastric tube once in the last third of pregnancy. 32 offsprings were observed. Group No. 6: 26 animals received 80 mg/kg b.w. ephedrine (D,L-Ephedrin "Fahlberg" DAB 7) and 50 mg/kg b.w. NaNO2 once per two weeks by gastric tube. Duration of the experiment: 6 months. 5 controls were applied 80 mg/kg b.w. ephedrine. Group No. 7: 30 hooded rats received a standard diet containing 0.5 per cent ephedrine and 0.2 per cent NaNO2. Duration of the experiment: 50 days. Only ephedrine was added in the same concentration to the diet of 10 control animals. results (tables 2--7): All animals that survived for more than 80 days were included in the analysis. In group No. 1 after a latency of 254--432 days 5 out of 14 rats developed malignant tumours of the nasal cavities (1), esophagus (1), leukoses (2), or soft tissue sarcoma (1). The spectrum of tumours indicates the endogenous formation of N-nitrosopiperazines. In groups 2 and 3 no significant carcinogenic action can be verified. In 56 per cent of hooded rats of group 4 reticulum-cell sarcomas, that were mainly localized in the paracoecal region (7) as well as leukoses (3) were registered. These findings were regarded as a carcinogenic action of the N-nitroso derivative of Obesin. In group No. 5 there is an insignificant carcinogenic effect. 2 paracoecal reticulum-cell sarcomas, 2 leukoses, 1 soft tissue sarcoma, 1 palsma-cytosis and 1 spindle cell sarcoma of the heart were found. After application of ephedrine and nitrite in group No. 6 and group No... | [Endogenous formation of carcinogenic-N-nitroso compounds in rats after application of drugs and nitrite (author's transl)]. During the last years in vitro investigations have demonstrated the possibility of nitrosation of different amines and amides including a number of drugs. The aim of the presented investigations was to test whether an endogenous nitrosation of piperazine, Obesin (propyhexedrine), and ephedrine is possible in long term experiments with rats. With the exception of group No. 3, the experiments were carried out with hooded rats (table 1). Group No. 1: 15 rats were given 100 mg/kg b.w. piperazine (Merck, Darmstadt) and 80 mg/kg b.w. NaNO2 by gastric tube. After the fourth application because of intoxication a reduction to 80 mg/kg b.w. piperazine and 50 mg/kg b.w. NaNO2 was necessary. Duration of treatment (in two weeks intervals): 7 months. Group No. 2: Pregnant rats received 80 mg/kg b.w. piperazine and 50 mg/kg b.w. NaNO2 by gastric tube once in the last third of pregnancy. In 13 offsprings the carcinogenic action was tested. Group No. 3: Pregnant Wistar rats were given 100 mg/kg b.w. piperazine and 70 mg/kg b.w. NaN92 by gastric tube once in the last third of pregnancy. Group No. 4: 20 hooded rats received 40 mg/kg b.w. Obesin (propylhexedrine, Fahlberg-List Magdeburg) and 40 mg/kg b.w. NaM92 once in two weekly intervals by gastric tube. Duration of the treatment: 6 months. 5 controls received 40 mg/kg b.w. Obesin or 40 mg/kg b.w. NaNO2 resp. Group No. 5: Hooded rats were given 40 mg/kg b.w. Obesin and 40 mg/kg b.w. NaNO2 by gastric tube once in the last third of pregnancy. 32 offsprings were observed. Group No. 6: 26 animals received 80 mg/kg b.w. ephedrine (D,L-Ephedrin "Fahlberg" DAB 7) and 50 mg/kg b.w. NaNO2 once per two weeks by gastric tube. Duration of the experiment: 6 months. 5 controls were applied 80 mg/kg b.w. ephedrine. Group No. 7: 30 hooded rats received a standard diet containing 0.5 per cent ephedrine and 0.2 per cent NaNO2. Duration of the experiment: 50 days. Only ephedrine was added in the same concentration to the diet of 10 control animals. results (tables 2--7): All animals that survived for more than 80 days were included in the analysis. In group No. 1 after a latency of 254--432 days 5 out of 14 rats developed malignant tumours of the nasal cavities (1), esophagus (1), leukoses (2), or soft tissue sarcoma (1). The spectrum of tumours indicates the endogenous formation of N-nitrosopiperazines. In groups 2 and 3 no significant carcinogenic action can be verified. In 56 per cent of hooded rats of group 4 reticulum-cell sarcomas, that were mainly localized in the paracoecal region (7) as well as leukoses (3) were registered. These findings were regarded as a carcinogenic action of the N-nitroso derivative of Obesin. In group No. 5 there is an insignificant carcinogenic effect. 2 paracoecal reticulum-cell sarcomas, 2 leukoses, 1 soft tissue sarcoma, 1 palsma-cytosis and 1 spindle cell sarcoma of the heart were found. After application of ephedrine and nitrite in group No. 6 and group No... | [
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|
PMID:14844 | Activity of two components of serum ribonuclease under conditions of physical exercise. | Serum ribonuclease activity before and after physical exercise in healthy persons was estimated. It is found that a work load of 6000 kgm/5 min increased ribonuclease activity measured at pH 8.5 and decreased the activity of the same enzyme measured at pH 7.0 in the presence of Zn SO4. The observed changes were more pronounced in untrained than in trained persons. | Activity of two components of serum ribonuclease under conditions of physical exercise. Serum ribonuclease activity before and after physical exercise in healthy persons was estimated. It is found that a work load of 6000 kgm/5 min increased ribonuclease activity measured at pH 8.5 and decreased the activity of the same enzyme measured at pH 7.0 in the presence of Zn SO4. The observed changes were more pronounced in untrained than in trained persons. | [
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|
PMID:14845 | Effect of various factors on the activity of trehalase from the larvae of Sesamia inferens Walker (Insect). | Trehalase from the salivary glands and the midgut of Sesamia inferens showed optimum activity at pH 5.8, and at temperatures of 50 and 60 degrees C respectively. The increase in the incubation period, enzyme concentration, and substrate concentration respectively increased the end-product, the hydrolysis, and the rate of hydrolysis of the substrate. Dialysis did not affect, tryptophan accelerated, and other amino acids and end-product inhibited the enzyme activity. | Effect of various factors on the activity of trehalase from the larvae of Sesamia inferens Walker (Insect). Trehalase from the salivary glands and the midgut of Sesamia inferens showed optimum activity at pH 5.8, and at temperatures of 50 and 60 degrees C respectively. The increase in the incubation period, enzyme concentration, and substrate concentration respectively increased the end-product, the hydrolysis, and the rate of hydrolysis of the substrate. Dialysis did not affect, tryptophan accelerated, and other amino acids and end-product inhibited the enzyme activity. | [
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|
PMID:14849 | Oxygen and carbon dioxide in the regulation of respiration. | When a sea-level resident ascends to a high altitude, his breathing immediately increases because of hypoxic stimulation of the peripheral chemoreceptors. In many species the aortic bodies are relatively unimportant in this response compared to the carotid bodies. When the subject stays at that altitude, his breathing increases progressively in the next few hours and days in a process termed ventilatory acclimatization and does not immediately return to control levels when hypoxia is terminated. Evidence is summarized indicating that this chronic process does not depend on the peripheral chemoreceptors or an initial respiratory alkalosis. Historical review indicates that the process of ventilatory acclimatization was initially attributed to renal excretion of plasma bicarbonate with development of a metabolic acidosis; but subsequent measurements indicated this process did not lower the arterial pH sufficiently to account for the ventilatory stimulation. More recently, ventilatory acclimatization has been attributed to accelerated removal of bicarbonate from the cerebrospinal fluid (CSF), producing a metabolic acidosis in the region of the medullary chemoreceptors; but still more recent observations indicate that this process, contrary to earlier observations, does not lower the CSF pH sufficiently to account for the ventilatory stimulation, either. Some other mechanism should be sought. | Oxygen and carbon dioxide in the regulation of respiration. When a sea-level resident ascends to a high altitude, his breathing immediately increases because of hypoxic stimulation of the peripheral chemoreceptors. In many species the aortic bodies are relatively unimportant in this response compared to the carotid bodies. When the subject stays at that altitude, his breathing increases progressively in the next few hours and days in a process termed ventilatory acclimatization and does not immediately return to control levels when hypoxia is terminated. Evidence is summarized indicating that this chronic process does not depend on the peripheral chemoreceptors or an initial respiratory alkalosis. Historical review indicates that the process of ventilatory acclimatization was initially attributed to renal excretion of plasma bicarbonate with development of a metabolic acidosis; but subsequent measurements indicated this process did not lower the arterial pH sufficiently to account for the ventilatory stimulation. More recently, ventilatory acclimatization has been attributed to accelerated removal of bicarbonate from the cerebrospinal fluid (CSF), producing a metabolic acidosis in the region of the medullary chemoreceptors; but still more recent observations indicate that this process, contrary to earlier observations, does not lower the CSF pH sufficiently to account for the ventilatory stimulation, either. Some other mechanism should be sought. | [
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|
PMID:14850 | New developments in our knowledge of the chemistry of renin. | Although the role of renin in hypertension continues to be incompletely defined, recent progress in the chemistry of renin has been considerable. Extensive purifications of hog kidney renin and the renin-like mouse submaxillary gland enzyme have been achieved. Various inhibitory peptides based on tetradecapeptide renin substrate have been useful in renin kinetic studies and in renin affinity chromatography. Classification of renin as an acid protease results from its marked inhibition by pepstatin and from the discovery that free carboxyl at the active site is essential for activity in human and hog kidney and mouse submaxillary gland enzymes. The presence of pseudorenin in all tissues has limited the use of model peptides as renin substrates in plasma and crude tissue extracts, since the proteolytic properties of the two enzymes are nearly identical. The existence of renin in multiple, chromatographically separable forms has been known. More recently inactive forms have been found in plasma, amniotic fluid, and hog and rabbit kidneys. Prolonged storage or treatment with acid, trypsin, or pepsin causes activation; in some instances the conversion is from a higher than normal molecular weight. The implications of these findings with respect to the renin-angiotensin system need much further investigation. | New developments in our knowledge of the chemistry of renin. Although the role of renin in hypertension continues to be incompletely defined, recent progress in the chemistry of renin has been considerable. Extensive purifications of hog kidney renin and the renin-like mouse submaxillary gland enzyme have been achieved. Various inhibitory peptides based on tetradecapeptide renin substrate have been useful in renin kinetic studies and in renin affinity chromatography. Classification of renin as an acid protease results from its marked inhibition by pepstatin and from the discovery that free carboxyl at the active site is essential for activity in human and hog kidney and mouse submaxillary gland enzymes. The presence of pseudorenin in all tissues has limited the use of model peptides as renin substrates in plasma and crude tissue extracts, since the proteolytic properties of the two enzymes are nearly identical. The existence of renin in multiple, chromatographically separable forms has been known. More recently inactive forms have been found in plasma, amniotic fluid, and hog and rabbit kidneys. Prolonged storage or treatment with acid, trypsin, or pepsin causes activation; in some instances the conversion is from a higher than normal molecular weight. The implications of these findings with respect to the renin-angiotensin system need much further investigation. | [
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|
PMID:14851 | [Enzymatic processes and chemical composition of the cecal contents of swine fed carbohydrates from different plants]. | A considerable amount of low-molecular acids (14.5-18.2 meqv/100 ml) is formed in the coecum in consequence of fermentation of cellulose, starch, and other components of chyme with microflora. The total concentration and molecular per cent ratio of the acetic, propionic, butyric, and lactic acids depended on the composition of the carbohydrate ration. | [Enzymatic processes and chemical composition of the cecal contents of swine fed carbohydrates from different plants]. A considerable amount of low-molecular acids (14.5-18.2 meqv/100 ml) is formed in the coecum in consequence of fermentation of cellulose, starch, and other components of chyme with microflora. The total concentration and molecular per cent ratio of the acetic, propionic, butyric, and lactic acids depended on the composition of the carbohydrate ration. | [
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PMID:14852 | Time dependence of the effect of splenectomy on graft-versus-host reactivity of lymph node cells. | Changes in cell-mediated reactivity of lymph node cells at various intervals after splenectomy were investigated in three assays measuring the GVH reactivity of parental cells in F1 hybrids --splenomegaly test in very young recipients and popliteal lymph node enlargement assay in adults measuring the proliferative component of the reaction, and mortality assay in sublethally irradiated recipients measuring the killer activity of the cell inoculum. During the early postsplenectomy period the reactivity of the particular amounts of lymph node cells was lower than that of cells from normal donors, but at about 3 weeks after splenectomy it was higher. The increase was of short duration in the proliferation assay and at 5 weeks the reactivity declined markedly below the control values. The increase in activity persisted for 5 weeks after splenectomy in the "killer" assay. It is probable that the described changes in cell-mediated reactivity are involved in the total effect of splenectomy on the host's complex immune response, especially against normal and tumour allografts. | Time dependence of the effect of splenectomy on graft-versus-host reactivity of lymph node cells. Changes in cell-mediated reactivity of lymph node cells at various intervals after splenectomy were investigated in three assays measuring the GVH reactivity of parental cells in F1 hybrids --splenomegaly test in very young recipients and popliteal lymph node enlargement assay in adults measuring the proliferative component of the reaction, and mortality assay in sublethally irradiated recipients measuring the killer activity of the cell inoculum. During the early postsplenectomy period the reactivity of the particular amounts of lymph node cells was lower than that of cells from normal donors, but at about 3 weeks after splenectomy it was higher. The increase was of short duration in the proliferation assay and at 5 weeks the reactivity declined markedly below the control values. The increase in activity persisted for 5 weeks after splenectomy in the "killer" assay. It is probable that the described changes in cell-mediated reactivity are involved in the total effect of splenectomy on the host's complex immune response, especially against normal and tumour allografts. | [
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PMID:14858 | Aversive prenatal stimulation: effects on behavioral, biochemical, and somatic ontogeny in the rat. | Electric foot shock administered to pregnant rats altered the ontogeny of spontaneous motor activity in their pups. Prenatally stimulated (PMS) offspring were more active than controls on Days 1-10 but less active during the 3rd postpartum week. The age of peak activity, a major developmental landmark, occurred in PMS pups around 10 days of age; in controls maximum activity was not seen until the 3rd week. This effect was independent of the gender of the offspring and the timing of the gestational stimulation. Its appearance in both cross-fostered and fostered pups indicated the prenatal origin of the effect. The maturation of spontaneous alternation behavior and several reflexes and the appearance of physical features were not affected by prenatal stimulation. Moreover, both PMS and control groups exhibited an age-related increase in brain concentrations of norepinephrine, serotonin, and 5-hydroxyindoleacetic acid. These findings indicate that spontaneous motor activity is uniquely sensitive to PMS, and as far as can be determined here, PMS produces no generalized alteration in behavioral and physical ontogeny. | Aversive prenatal stimulation: effects on behavioral, biochemical, and somatic ontogeny in the rat. Electric foot shock administered to pregnant rats altered the ontogeny of spontaneous motor activity in their pups. Prenatally stimulated (PMS) offspring were more active than controls on Days 1-10 but less active during the 3rd postpartum week. The age of peak activity, a major developmental landmark, occurred in PMS pups around 10 days of age; in controls maximum activity was not seen until the 3rd week. This effect was independent of the gender of the offspring and the timing of the gestational stimulation. Its appearance in both cross-fostered and fostered pups indicated the prenatal origin of the effect. The maturation of spontaneous alternation behavior and several reflexes and the appearance of physical features were not affected by prenatal stimulation. Moreover, both PMS and control groups exhibited an age-related increase in brain concentrations of norepinephrine, serotonin, and 5-hydroxyindoleacetic acid. These findings indicate that spontaneous motor activity is uniquely sensitive to PMS, and as far as can be determined here, PMS produces no generalized alteration in behavioral and physical ontogeny. | [
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PMID:14859 | Decreased rat hepatic guanylate cyclase activity in streptozotocin-induced diabetes mellitus. | Guanylate cyclase is found in virtually all cells, but its physiologic role and the effect of hormones on its activity have not been clarified. Hepatic soluble guanylate cyclase activity (37,000 g supernatant) in rats with diabetes-mellitus-like syndrome induced by streptozotocin, 65 mg./kg. i.v., was 140 +/- 8 pmoles accumulated/mg. protein/10 min. (n = 13 rats) as against 279 +/- 16 pmoles accumulated/mg. protein/10 min. (n = 12 rats) in normal rats. The average blood sugar for the 12 normal rats was 100 +/- 4 mg./100 ml. and 546 +/- 32 mg./100 ml. for 13 diabetic rats. The decreased soluble hepatic guanylate cyclase activity in diabetic rats was completely restored to normal with 10 U. regular insulin, i.p. The maximum increase in guanylate cyclase activity was observed as early as five minutes and as late as two hours after insulin administration. Insulin restoration of guanylate cyclase was dose-related over a range of 1 U. to 10 U., i.p. Hepatic cyclic GMP levels in vivo paralleled in-vitro guanylate cyclase activity, being 29 +/- 0.4 pmoles/gm. wet weight in normals, 17 +/- 0.4 pmoles/gm. wet weight in streptozotocin-diabetic rats, and 38 +/- 0.4 pmoles/gm. wet weight two hours after the injection of 10 U. regular insulin. We conclude that rat hepatic guanylate cyclase is decreased in streptozotocin-induced diabetes and that insulin modulates this enzyme. The administration of exogenous insulin in normal animals did not further augment hepatic guanylate cyclase activity. | Decreased rat hepatic guanylate cyclase activity in streptozotocin-induced diabetes mellitus. Guanylate cyclase is found in virtually all cells, but its physiologic role and the effect of hormones on its activity have not been clarified. Hepatic soluble guanylate cyclase activity (37,000 g supernatant) in rats with diabetes-mellitus-like syndrome induced by streptozotocin, 65 mg./kg. i.v., was 140 +/- 8 pmoles accumulated/mg. protein/10 min. (n = 13 rats) as against 279 +/- 16 pmoles accumulated/mg. protein/10 min. (n = 12 rats) in normal rats. The average blood sugar for the 12 normal rats was 100 +/- 4 mg./100 ml. and 546 +/- 32 mg./100 ml. for 13 diabetic rats. The decreased soluble hepatic guanylate cyclase activity in diabetic rats was completely restored to normal with 10 U. regular insulin, i.p. The maximum increase in guanylate cyclase activity was observed as early as five minutes and as late as two hours after insulin administration. Insulin restoration of guanylate cyclase was dose-related over a range of 1 U. to 10 U., i.p. Hepatic cyclic GMP levels in vivo paralleled in-vitro guanylate cyclase activity, being 29 +/- 0.4 pmoles/gm. wet weight in normals, 17 +/- 0.4 pmoles/gm. wet weight in streptozotocin-diabetic rats, and 38 +/- 0.4 pmoles/gm. wet weight two hours after the injection of 10 U. regular insulin. We conclude that rat hepatic guanylate cyclase is decreased in streptozotocin-induced diabetes and that insulin modulates this enzyme. The administration of exogenous insulin in normal animals did not further augment hepatic guanylate cyclase activity. | [
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|
PMID:14860 | Aspects of carbohydrate metabolism in developing brain. | This review considers carbohydrate metabolism in the developing brain, in particular the proportion of glucose metabolized via the pentose phosphate pathway. Although small in amount, this fraction serves a vital rôle in some aspects of brain function. Evidence is presented that the pentose phosphate pathway subserves different functions as the developing brain progresses through the stages of growth and myelination to full neurological competence. The general aspects considered are the changing patterns of brain enzymes during development; the flux of glucose through the alternative pathways of glucose metabolism in the developing brain; the functional significance of the pentose phosphate pathway; and the regional and functional association of the pentose phosphate pathway activity and the detoxication of biogenic amínes. | Aspects of carbohydrate metabolism in developing brain. This review considers carbohydrate metabolism in the developing brain, in particular the proportion of glucose metabolized via the pentose phosphate pathway. Although small in amount, this fraction serves a vital rôle in some aspects of brain function. Evidence is presented that the pentose phosphate pathway subserves different functions as the developing brain progresses through the stages of growth and myelination to full neurological competence. The general aspects considered are the changing patterns of brain enzymes during development; the flux of glucose through the alternative pathways of glucose metabolism in the developing brain; the functional significance of the pentose phosphate pathway; and the regional and functional association of the pentose phosphate pathway activity and the detoxication of biogenic amínes. | [
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|
PMID:14861 | Serum gastrin levels before and after 6 weeks of cimetidine therapy in patients with duodenal ulcer. | Fasting serum gastrins were carried out at the beginning and end of a 6-week double-blind trial of cimetidine and placebo in 30 patients with duodenal ulcer. Cimetidine did not cause any significant change in fasting serum gastrin levels after 6 weeks of therapy. | Serum gastrin levels before and after 6 weeks of cimetidine therapy in patients with duodenal ulcer. Fasting serum gastrins were carried out at the beginning and end of a 6-week double-blind trial of cimetidine and placebo in 30 patients with duodenal ulcer. Cimetidine did not cause any significant change in fasting serum gastrin levels after 6 weeks of therapy. | [
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|
PMID:14862 | The effect of the alkaline tide on serum-ionized calcium concentration in man. | The effect of the gastric alkaline tide on serum-ionized calcium levels was determined in human subjects. Gastric acid seretion was stimulated by a standard steak meal, human synthetic gastrin, and betazole hydrochloride. Ionized calcium levels fell to a similar extent after each stimulus. The mean decrease in calcium ion concentration in all experiments was 5.4% of the basal concentration. The fall in serum calcium ion concentration was highly correlated with the rise in serum pH. We speculate that increased formation of calcium bicarbonate complex in the serum as well as increased binding of ionized calcium by serum protein accounts for the surprisingly large effect of the alkaline tide on serumionized calcium levels. | The effect of the alkaline tide on serum-ionized calcium concentration in man. The effect of the gastric alkaline tide on serum-ionized calcium levels was determined in human subjects. Gastric acid seretion was stimulated by a standard steak meal, human synthetic gastrin, and betazole hydrochloride. Ionized calcium levels fell to a similar extent after each stimulus. The mean decrease in calcium ion concentration in all experiments was 5.4% of the basal concentration. The fall in serum calcium ion concentration was highly correlated with the rise in serum pH. We speculate that increased formation of calcium bicarbonate complex in the serum as well as increased binding of ionized calcium by serum protein accounts for the surprisingly large effect of the alkaline tide on serumionized calcium levels. | [
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PMID:14865 | Lower esophageal sphincter pressure in cirrhotic men with ascites: before and after diuresis. | Lower esophageal sphincter pressure (LESP) was measured in 10 biopsy-proved cirrhotics with esophageal varices and tense ascites before and after diuresis to evaluate of ascites might play in the development of variceal bleeding. In the 10 cirrhotic men studied, basal LESP was 30.9 +/- 1.7 mm Hg before and 22.7 +/- 1.3 mm Hg (P less than 0.01) after a diuresis which resulted in a mean 12-kg weight loss. LESP responses to abdominal compression were also evaluated. The change in LESP in response to a standard degree of abdominal compression was greater in the presence of ascites (8.5 +/- 0.4) than in its absence (6.3 +/- 0.4) (P less than 0.01). Basal gastric pH and fasting plasma gastrin concentrations did not differ during the two testing periods. Based on these data and the rarity with which cirrhotic patients with ascites complain of heartburn, it is concluded that reflux esophagitis caused by failure of the lower esophageal sphincter to remain competent is unlikely to be a significant etiological factor in the development of variceal bleeding. | Lower esophageal sphincter pressure in cirrhotic men with ascites: before and after diuresis. Lower esophageal sphincter pressure (LESP) was measured in 10 biopsy-proved cirrhotics with esophageal varices and tense ascites before and after diuresis to evaluate of ascites might play in the development of variceal bleeding. In the 10 cirrhotic men studied, basal LESP was 30.9 +/- 1.7 mm Hg before and 22.7 +/- 1.3 mm Hg (P less than 0.01) after a diuresis which resulted in a mean 12-kg weight loss. LESP responses to abdominal compression were also evaluated. The change in LESP in response to a standard degree of abdominal compression was greater in the presence of ascites (8.5 +/- 0.4) than in its absence (6.3 +/- 0.4) (P less than 0.01). Basal gastric pH and fasting plasma gastrin concentrations did not differ during the two testing periods. Based on these data and the rarity with which cirrhotic patients with ascites complain of heartburn, it is concluded that reflux esophagitis caused by failure of the lower esophageal sphincter to remain competent is unlikely to be a significant etiological factor in the development of variceal bleeding. | [
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|
PMID:14866 | Small intestinal injury in the graft versus host reaction: an innocent bystander phenomenon. | Marked morphological and functional damage to the small intestine occurs during the graft versus host reaction (GVHR); the structural lesion is characterized by crypt hypertrophy and villus shortening. The purpose of this study was to determine whether the intestine is injured in the GVHR as an antigenic target of the grafted immunocompetent cells or as an innocent bystander to donor-host lymphoid interaction. Implants of fetal mouse small intestine from either C57BL/6 or (C57BL/L X DBA/2)F1 (BDF1) hybrid donors were placed under the kidney capsule of adult BDF1 male mice. After 4 weeks, both groups were injected with parental C57BL/6 spleen cells, syngeneic BDF1 spleen cells, or no cells. Two weeks after injection the mice were killed and the implants were removed for histological processing and measurement of villus height and crypt depth. The villus-crypt ratio in the BDF1 implants of mice receiving C57BL/6 cells was 1.32 +/- 0.3 compared to 2.51 +/- 0.4 in controls receiving either syngeneic BDF1 cells or no cells (P less than 0.001). The villus-crypt ratio in the C57BL/6 implants of mice receiving C57BL/6 cells was 1.79 +/- 0.4 compared to 2.48 +/- 0.4 in the controls receiving either syngeneic BDF1 cells or no cells (P less than 0.001). Because the latter implant is antigenically identical to the spleen cells eliciting the GVHR, we conclude that the small bowel is injured in the GVHR as an innocent bystander to cytotoxic donor-host lymphoid interaction. | Small intestinal injury in the graft versus host reaction: an innocent bystander phenomenon. Marked morphological and functional damage to the small intestine occurs during the graft versus host reaction (GVHR); the structural lesion is characterized by crypt hypertrophy and villus shortening. The purpose of this study was to determine whether the intestine is injured in the GVHR as an antigenic target of the grafted immunocompetent cells or as an innocent bystander to donor-host lymphoid interaction. Implants of fetal mouse small intestine from either C57BL/6 or (C57BL/L X DBA/2)F1 (BDF1) hybrid donors were placed under the kidney capsule of adult BDF1 male mice. After 4 weeks, both groups were injected with parental C57BL/6 spleen cells, syngeneic BDF1 spleen cells, or no cells. Two weeks after injection the mice were killed and the implants were removed for histological processing and measurement of villus height and crypt depth. The villus-crypt ratio in the BDF1 implants of mice receiving C57BL/6 cells was 1.32 +/- 0.3 compared to 2.51 +/- 0.4 in controls receiving either syngeneic BDF1 cells or no cells (P less than 0.001). The villus-crypt ratio in the C57BL/6 implants of mice receiving C57BL/6 cells was 1.79 +/- 0.4 compared to 2.48 +/- 0.4 in the controls receiving either syngeneic BDF1 cells or no cells (P less than 0.001). Because the latter implant is antigenically identical to the spleen cells eliciting the GVHR, we conclude that the small bowel is injured in the GVHR as an innocent bystander to cytotoxic donor-host lymphoid interaction. | [
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PMID:14867 | Stimulatory (H1) and inhibitory (H2) histamine receptors in gallbladder muscle. | The nature of histamine receptors in gallbladder muscle and examined using specific histamine-receptor agonists and antagonists. The H2-receptor antagonist, metiamide, augmented the contractile response to histamine indicating that gallbladder muscle possessed stimulatory H1 receptors and inhibitory H2 receptors. The independent inhibitory character of H2 receptors was confirmed by (1) induction of relaxation with histamine after H1-receptor blockade and the suppression of this relaxation with metiamide, and (2) induction of relaxation with a specific H2-receptor agonist, 4-methyl histamine and the suppression of this relaxation with metiamide. Further, blockade of H2 but not of H1 receptors augmented the response to the octapeptide of cholecystokinin. The nature of this effect was such that the apparent affinity of the octapeptide for its own receptor was increased. The finding raised the possibility that in their native unoccupied state, H2 receptors may modify the response to hormonal agents. | Stimulatory (H1) and inhibitory (H2) histamine receptors in gallbladder muscle. The nature of histamine receptors in gallbladder muscle and examined using specific histamine-receptor agonists and antagonists. The H2-receptor antagonist, metiamide, augmented the contractile response to histamine indicating that gallbladder muscle possessed stimulatory H1 receptors and inhibitory H2 receptors. The independent inhibitory character of H2 receptors was confirmed by (1) induction of relaxation with histamine after H1-receptor blockade and the suppression of this relaxation with metiamide, and (2) induction of relaxation with a specific H2-receptor agonist, 4-methyl histamine and the suppression of this relaxation with metiamide. Further, blockade of H2 but not of H1 receptors augmented the response to the octapeptide of cholecystokinin. The nature of this effect was such that the apparent affinity of the octapeptide for its own receptor was increased. The finding raised the possibility that in their native unoccupied state, H2 receptors may modify the response to hormonal agents. | [
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|
PMID:14870 | [Pharmacological analysis of effects of perimetazine on isolated smooth muscle]. | Effects of perimetazine on the motility of the isolated smooth muscle of guinea pig (ileum, taenia coli, uterus, vas deferens and trachea) and the ileum of rabbit were studied. The results obtained are as follows: Perimetazine showed a specific antihistamine, antiadrenaline and antiserotonin action. Moreover, antiacetylcholine and anti-BaC12 actions were observed with high doses of perimetazine as well as chlorpromazine and such actions were attributed to the nonspecific direct action on the smooth muscle. Both the spontaneous movement and the tonus of guinea pig taenia coli were also inhibited by a high concentration of perimetazine. The spontaneous membrane action potentials of the taenia coli recorded by the use of the sucrose-gap method were inhibited by a high concentration of perimetazine both in frequency and in amplitude with relaxation of the tonus. The action potentials were abolished however and change in the resting membrane level was not clearly observed. | [Pharmacological analysis of effects of perimetazine on isolated smooth muscle]. Effects of perimetazine on the motility of the isolated smooth muscle of guinea pig (ileum, taenia coli, uterus, vas deferens and trachea) and the ileum of rabbit were studied. The results obtained are as follows: Perimetazine showed a specific antihistamine, antiadrenaline and antiserotonin action. Moreover, antiacetylcholine and anti-BaC12 actions were observed with high doses of perimetazine as well as chlorpromazine and such actions were attributed to the nonspecific direct action on the smooth muscle. Both the spontaneous movement and the tonus of guinea pig taenia coli were also inhibited by a high concentration of perimetazine. The spontaneous membrane action potentials of the taenia coli recorded by the use of the sucrose-gap method were inhibited by a high concentration of perimetazine both in frequency and in amplitude with relaxation of the tonus. The action potentials were abolished however and change in the resting membrane level was not clearly observed. | [
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|
PMID:14871 | [Findings in the gastric mucosa and gastric secretion in rats treated with methyl O-(4-hydroxy-3-methoxycinnamoyl)reserpate (CD-3400) and reserpine derivatives]. | CD-3400 developed by Nippon Chemiphar Co. Ltd., is a new antihypertensive agent belonging to the class of rauwolfa alkaloids. Influence of the agent on gastric mucosa, healing process of acetic acid-induced gastric ulcer and gastric juice in rats was investigated and compared with effects of reserpine and rescinnamine. CD-3400-induced gastric lesions were fewer in number than those produced with reserpine and rescinnamine in fasted rats. After a three day treatment of CD-3400 to fed rats, however, there were few gastric lesions, while reserpine- and rescinnamine-induced gastric lesions were aggravated to a greater extent that when a single administration was given to fasted rats. Influence of CD-3400, reserpine and rescinnamine on the healing process of acetic acid-induced ulcer was insignificant, but treatment with high doses of reserpine and rescinnamine resulted in death. Pretreatment with CD-3400 and reserpine produced a decrease in gastric acid and K+, and an increase in Na+. Repeated administration of reserpine for 5 days resulted in a decrease of both gastric volume and acid, while such was not seen with CD-3400. Treatment with anticholinergic agents such as atropine sulfate and atropine methylbromide inhibited CD-3400- and reserpine-induced gastric lesions. From these results, it would appear that cholinergic factors play a role in the pathogenesis of CD-3400-induced gastric lesions, as in the case with reserpine, and that the responses of these lesions to reserpine and CD-3400 correlate with changes of ionic fluxes in gastric juice. | [Findings in the gastric mucosa and gastric secretion in rats treated with methyl O-(4-hydroxy-3-methoxycinnamoyl)reserpate (CD-3400) and reserpine derivatives]. CD-3400 developed by Nippon Chemiphar Co. Ltd., is a new antihypertensive agent belonging to the class of rauwolfa alkaloids. Influence of the agent on gastric mucosa, healing process of acetic acid-induced gastric ulcer and gastric juice in rats was investigated and compared with effects of reserpine and rescinnamine. CD-3400-induced gastric lesions were fewer in number than those produced with reserpine and rescinnamine in fasted rats. After a three day treatment of CD-3400 to fed rats, however, there were few gastric lesions, while reserpine- and rescinnamine-induced gastric lesions were aggravated to a greater extent that when a single administration was given to fasted rats. Influence of CD-3400, reserpine and rescinnamine on the healing process of acetic acid-induced ulcer was insignificant, but treatment with high doses of reserpine and rescinnamine resulted in death. Pretreatment with CD-3400 and reserpine produced a decrease in gastric acid and K+, and an increase in Na+. Repeated administration of reserpine for 5 days resulted in a decrease of both gastric volume and acid, while such was not seen with CD-3400. Treatment with anticholinergic agents such as atropine sulfate and atropine methylbromide inhibited CD-3400- and reserpine-induced gastric lesions. From these results, it would appear that cholinergic factors play a role in the pathogenesis of CD-3400-induced gastric lesions, as in the case with reserpine, and that the responses of these lesions to reserpine and CD-3400 correlate with changes of ionic fluxes in gastric juice. | [
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|
PMID:14872 | [Effects of methyl O-(4-hydroxy-3-methoxycinnamoly)reserpate (CD-3400) on the peripheral nervous system: especially on the smooth muscle organs]. | Effects of methyl O-(4-hydroxy-3-methoxycinnamoyl) reserpate (CD-3400), a new antihypertensive agent, on the peripheral nervous system in mice, rats and guinea pigs were investigated and compared with effects of reserpine and rescinnamine. Oral administration of CD-3400 in doses from 40 to 320 mg/kg revealed a miotic action and such was weaker than that seen with reserpine and rescinnamine. The intestinal propulsion in mice was accelerated by pretreatment with reserpine, but no so with CD-3400. In isolated guinea pig vas deferens, CD-3400 at a concentration of 1 X 10(-5) M inhibited on norepinephrine-induced contraction non-competitively. However, in insolated rat vas deferens pretreated with CD-3400 (1 mg/kg, i.p.) for 5 days, norepinephrine-induced contraction was potentiated and this effect was weaker than that seen with reserpine and rescinnamine. In isolated rat vas deferens pretreated with CD-3400 for 1, 2 and 5 days,, the contraction induced by tyramine (3 X 10(-5)M) was significantly inhibited. The effect was qualitively similar to those of reserpine and rescinnamine. The intravenous administration of these three rauwolfia alkaloids in doses of 1, 2 and 4 mg/kg had no effect on the spontaneous movement of rat uterus. In isolated rat uterus pretreated with CD-3400, no significant effect was observed on oxytocin- and isoproterenol-induced responses. The peripheral actions of CD-3400 are attributed to a deterioration in the function the sympathetic nerve resulting in depletion of catecholamine stores. Efect of CD-3400 were slightly weaker than those of reserpine and rescinnamine. | [Effects of methyl O-(4-hydroxy-3-methoxycinnamoly)reserpate (CD-3400) on the peripheral nervous system: especially on the smooth muscle organs]. Effects of methyl O-(4-hydroxy-3-methoxycinnamoyl) reserpate (CD-3400), a new antihypertensive agent, on the peripheral nervous system in mice, rats and guinea pigs were investigated and compared with effects of reserpine and rescinnamine. Oral administration of CD-3400 in doses from 40 to 320 mg/kg revealed a miotic action and such was weaker than that seen with reserpine and rescinnamine. The intestinal propulsion in mice was accelerated by pretreatment with reserpine, but no so with CD-3400. In isolated guinea pig vas deferens, CD-3400 at a concentration of 1 X 10(-5) M inhibited on norepinephrine-induced contraction non-competitively. However, in insolated rat vas deferens pretreated with CD-3400 (1 mg/kg, i.p.) for 5 days, norepinephrine-induced contraction was potentiated and this effect was weaker than that seen with reserpine and rescinnamine. In isolated rat vas deferens pretreated with CD-3400 for 1, 2 and 5 days,, the contraction induced by tyramine (3 X 10(-5)M) was significantly inhibited. The effect was qualitively similar to those of reserpine and rescinnamine. The intravenous administration of these three rauwolfia alkaloids in doses of 1, 2 and 4 mg/kg had no effect on the spontaneous movement of rat uterus. In isolated rat uterus pretreated with CD-3400, no significant effect was observed on oxytocin- and isoproterenol-induced responses. The peripheral actions of CD-3400 are attributed to a deterioration in the function the sympathetic nerve resulting in depletion of catecholamine stores. Efect of CD-3400 were slightly weaker than those of reserpine and rescinnamine. | [
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|
PMID:14873 | Separation and properties of alpha-mannosidase and mannanase from the basidiomycete Phellinus abietis. | Proteins of a crude enzyme preparation obtained from the cultivation medium of the basidiomycete Phellinus abietis were separated by gel filtration and ion-exchange chromatography. The preparation contained a minimum of three enzymes capable of splitting alpha-D-mannosidic bonds: alpha-mannosidase, exomannanase, and endomannanase, which were separated. Some properties of the mannanase complex of the crude enzyme preparation, and of a partially purified alpha-mannosidase were examined. The mannanase complex exhibited two pH optima, its temperature optimum being at 45 degrees C. The pH optimum of purified alpha-mannosidase was at pH 5.0, the temperature optimum being at 45 degrees C. The pH optimum of purifed alpha-mannosidase was at pH 5.0, the temperature optimum at at 60 degrees C; the enzyme had a relatively high heat stability. The Km of alpha-mannosidase for p-nitrophenyl alpha-D-mannopyranoside was 1.5 X 10(-5) M. Pure alpha-mannosidase did not split mannan. | Separation and properties of alpha-mannosidase and mannanase from the basidiomycete Phellinus abietis. Proteins of a crude enzyme preparation obtained from the cultivation medium of the basidiomycete Phellinus abietis were separated by gel filtration and ion-exchange chromatography. The preparation contained a minimum of three enzymes capable of splitting alpha-D-mannosidic bonds: alpha-mannosidase, exomannanase, and endomannanase, which were separated. Some properties of the mannanase complex of the crude enzyme preparation, and of a partially purified alpha-mannosidase were examined. The mannanase complex exhibited two pH optima, its temperature optimum being at 45 degrees C. The pH optimum of purified alpha-mannosidase was at pH 5.0, the temperature optimum being at 45 degrees C. The pH optimum of purifed alpha-mannosidase was at pH 5.0, the temperature optimum at at 60 degrees C; the enzyme had a relatively high heat stability. The Km of alpha-mannosidase for p-nitrophenyl alpha-D-mannopyranoside was 1.5 X 10(-5) M. Pure alpha-mannosidase did not split mannan. | [
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|
PMID:14874 | [A new method for the clinical determination of urinary porphobilinogen and delta-aminolevulinic acid]. | A new method has been developed for clinical determination of porphobilinogen and delta-aminolevulinic acid in urine. Compared with the method of Mauzerall and Granick, being the most commonly used method for clinical application, there is an essential reduction of work and needed time. During the flow of 0.5 ml urine on a 0.7 cm x 2 cm column of macroporous cation exchange resin in H+-form porphobilinogen and delta-aminolevulinic acid are bound to the resin. After rinsing the column with 3 ml N acetic acid and 1 ml H2O the bound hemprecursors were eluted with 5 ml buffer. Concentration of porphobilinogen and of the pyrrol, synthesized from delta-aminolevulinic acid by Knorr-reaction is measured photometrically at 546 nm after Ehrich-reaction. After a short-time regeneration of columns -- 5 ml buffer, 3 ml N acetic acid -- these columns can be reused at least 20 times. | [A new method for the clinical determination of urinary porphobilinogen and delta-aminolevulinic acid]. A new method has been developed for clinical determination of porphobilinogen and delta-aminolevulinic acid in urine. Compared with the method of Mauzerall and Granick, being the most commonly used method for clinical application, there is an essential reduction of work and needed time. During the flow of 0.5 ml urine on a 0.7 cm x 2 cm column of macroporous cation exchange resin in H+-form porphobilinogen and delta-aminolevulinic acid are bound to the resin. After rinsing the column with 3 ml N acetic acid and 1 ml H2O the bound hemprecursors were eluted with 5 ml buffer. Concentration of porphobilinogen and of the pyrrol, synthesized from delta-aminolevulinic acid by Knorr-reaction is measured photometrically at 546 nm after Ehrich-reaction. After a short-time regeneration of columns -- 5 ml buffer, 3 ml N acetic acid -- these columns can be reused at least 20 times. | [
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|
PMID:14875 | [The effect of anesthesia on the postoperative behavior of liver enzymes and protein fractions]. | Liver tests were performed postoperatively on 25 women with "healthy" livers. Patients were divided into 3 groups according to the type of anesthesia used: 1. halothane; 2. neuroleptic; 3. peridural. In all 3 groups the variations in serum proteins and liver enzymes were similar up to the 15th postoperative day. Variations of LDH, AP, LAP, and CHE were within normal range; slight to moderate increases above normal were observed for GPT, GOT, GLDH, and gamma-GT. All 3 groups barely differ with respect to enzyme levels during the postoperative course. We therefore interpret these changes to indicate a reaction of the liver to the stress of operation rather than to the mode of anesthesia. | [The effect of anesthesia on the postoperative behavior of liver enzymes and protein fractions]. Liver tests were performed postoperatively on 25 women with "healthy" livers. Patients were divided into 3 groups according to the type of anesthesia used: 1. halothane; 2. neuroleptic; 3. peridural. In all 3 groups the variations in serum proteins and liver enzymes were similar up to the 15th postoperative day. Variations of LDH, AP, LAP, and CHE were within normal range; slight to moderate increases above normal were observed for GPT, GOT, GLDH, and gamma-GT. All 3 groups barely differ with respect to enzyme levels during the postoperative course. We therefore interpret these changes to indicate a reaction of the liver to the stress of operation rather than to the mode of anesthesia. | [
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|
PMID:14890 | Bacteriostatic effect of human milk and bovine colostrum on Escherichia coli: importance of bicarbonate. | At pH 7.4 and in the presence of NaHCO3, human milk and bovine colostrum inhibited the growth of Escherichia coli O111. Adding sufficient iron to saturate the iron-binding capacity of the lactoferrin present in the milk or colostrum prevented bacteriostasis. At pH 6.8 neither molk nor colostrum inhibited E. coli 0111. Adjusting the pH to 7.4 with NaHCO3 resulted in the development of bacteriostatic activity. Adjusting the pH to 7.4 with NaOH was ineffective. Dialyzed colostrum and milk inhibited bacterial growth at pH 6.8 in the absence of added NaHCO3; addition of citrate or iron abolished bacteriostasis. The chromatographic elution profile of tyrosyl-transfer ribonucleic acid (tRNA) from iron-replete E. coli differs significantly from that of tyrosyl-tRNA from iron-deficient organisms. Examination of the elution profile tyrosyl-tRNA from E. coli 0111 growing in colostrum without added NaHCO3 showed that such bacteria were fully replete in iron. The nature of the elution profile of tyrosyl-tRNA also showed that iron was freely available to the bacteria when citrate was added to dialyzed colostrum but not available in its absence, even at pH 6.8. Results support the idea that the bacteriostatic action of milk and colostrum, due to the combined action of antibody and lactoferrin, depends on the addition of bicarbonate to counteract the iron-mobilizing effect of the citrate normally present in these secretions. | Bacteriostatic effect of human milk and bovine colostrum on Escherichia coli: importance of bicarbonate. At pH 7.4 and in the presence of NaHCO3, human milk and bovine colostrum inhibited the growth of Escherichia coli O111. Adding sufficient iron to saturate the iron-binding capacity of the lactoferrin present in the milk or colostrum prevented bacteriostasis. At pH 6.8 neither molk nor colostrum inhibited E. coli 0111. Adjusting the pH to 7.4 with NaHCO3 resulted in the development of bacteriostatic activity. Adjusting the pH to 7.4 with NaOH was ineffective. Dialyzed colostrum and milk inhibited bacterial growth at pH 6.8 in the absence of added NaHCO3; addition of citrate or iron abolished bacteriostasis. The chromatographic elution profile of tyrosyl-transfer ribonucleic acid (tRNA) from iron-replete E. coli differs significantly from that of tyrosyl-tRNA from iron-deficient organisms. Examination of the elution profile tyrosyl-tRNA from E. coli 0111 growing in colostrum without added NaHCO3 showed that such bacteria were fully replete in iron. The nature of the elution profile of tyrosyl-tRNA also showed that iron was freely available to the bacteria when citrate was added to dialyzed colostrum but not available in its absence, even at pH 6.8. Results support the idea that the bacteriostatic action of milk and colostrum, due to the combined action of antibody and lactoferrin, depends on the addition of bicarbonate to counteract the iron-mobilizing effect of the citrate normally present in these secretions. | [
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|
PMID:14891 | Reproduction of the eosinopenia of acute infection by passive transfer of a material obtained from inflammatory exudate. | Studies of the eosinopenic effect of acute inflammation were conducted in mice previously rendered eosinophilic with trichinosis. Exudate removed from a pneumococcal abscess contained material (eosinopenic factor [EF]) capable of causing eosinopenia of 4- to 24-h duration when injected intraperitoneally into eosinophilic mice. The material passed through a 0.45-micronm filter, but was retained by a dialysis membrane capable of retaining protein molecules of greater than approximately 30,000 molecular weight. EF was soluble in 7% perchloric acid, was not destroyed by pneumococcal proteolytic enzymes in the presence of Trasylol, but was inactivated by heating to 56 degrees C for 30 min. EF was detectable in the exudate after 10 h and had reached its highest concentration after 20 h. When the effect of EF was expressed as a percent suppression of control eosinophil levels, there was a geometric dose response. Eosinopenia could not be ascribed to steroids present in the preparation, and the EF was effective in adrenalectomized animals. Eosinopenia was not induced by transfer of similarly treated heat-killed pneumococci, pneumococcal culture filtrate, or normal serum. The eosinopenia of acute infection may be the direct effect of a substance present at the site of acute inflammation. | Reproduction of the eosinopenia of acute infection by passive transfer of a material obtained from inflammatory exudate. Studies of the eosinopenic effect of acute inflammation were conducted in mice previously rendered eosinophilic with trichinosis. Exudate removed from a pneumococcal abscess contained material (eosinopenic factor [EF]) capable of causing eosinopenia of 4- to 24-h duration when injected intraperitoneally into eosinophilic mice. The material passed through a 0.45-micronm filter, but was retained by a dialysis membrane capable of retaining protein molecules of greater than approximately 30,000 molecular weight. EF was soluble in 7% perchloric acid, was not destroyed by pneumococcal proteolytic enzymes in the presence of Trasylol, but was inactivated by heating to 56 degrees C for 30 min. EF was detectable in the exudate after 10 h and had reached its highest concentration after 20 h. When the effect of EF was expressed as a percent suppression of control eosinophil levels, there was a geometric dose response. Eosinopenia could not be ascribed to steroids present in the preparation, and the EF was effective in adrenalectomized animals. Eosinopenia was not induced by transfer of similarly treated heat-killed pneumococci, pneumococcal culture filtrate, or normal serum. The eosinopenia of acute infection may be the direct effect of a substance present at the site of acute inflammation. | [
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|
PMID:14892 | Magnesium-dependent adenosine triphosphatase as a marker enzyme for the plasma membrane of human polymorphonuclear leukocytes. | The adenosine triphosphatase (ATPase) activities of human polymorphonuclear leukocytes (PMNL) were studied with an assay that monitored the release of 32P-labeled inorganic pyrophosphate (32P1) from gamma-[32P]adenosine 5'-triphosphate (ATP). In cell homogenates, (Na+ + K+)-sensitive, ouabain-inhibitable ATPase comprised an insignificant fraction of the total ATPase activity. Additions of p-nitrophenyl phosphate and beta-glycerophosphate (substrates for nonspecific acid and alkaline phosphatases) and of tartrate (inhibitor of acid phosphatase) gave no indication of inhibition. This suggested that the assay was relatively specific for ATP hydrolysis. The activity was found to have a pH optimum of 8.7 and a Km for ATP of 0.6 mM. There was an absolute requirement for Mg2+, with other divalent cations substituting less efficiently. When the Mg2+-dependent ATPase activity of intact cells was compared with that in homogenized cells, no significant difference was observed. The activity in intact cells was linear with respect to incubation time up to at least l0 min. Trypan blue staining and lactate dehydrogenase assays revealed that greater than 92% of the PMNL remained intact and viable during the assay. No soluble ATPase was released from the cells under assay conditions. In following the distribution of gamma[32P]ATP and 32P2 counts became cell associated. Since the experimental evidence supports the observation that PMNL remain intact and viable and that ATP does not penetrate the cell under assay conditions, it is proposed that greater than 90% of the Mg2+-dependent ATPase of the human PMNL is associated with a plasma membrnae enzyme. This would qualify the enzyme for the role of a plasma membrane marker for future fractionation and isolation attempts. | Magnesium-dependent adenosine triphosphatase as a marker enzyme for the plasma membrane of human polymorphonuclear leukocytes. The adenosine triphosphatase (ATPase) activities of human polymorphonuclear leukocytes (PMNL) were studied with an assay that monitored the release of 32P-labeled inorganic pyrophosphate (32P1) from gamma-[32P]adenosine 5'-triphosphate (ATP). In cell homogenates, (Na+ + K+)-sensitive, ouabain-inhibitable ATPase comprised an insignificant fraction of the total ATPase activity. Additions of p-nitrophenyl phosphate and beta-glycerophosphate (substrates for nonspecific acid and alkaline phosphatases) and of tartrate (inhibitor of acid phosphatase) gave no indication of inhibition. This suggested that the assay was relatively specific for ATP hydrolysis. The activity was found to have a pH optimum of 8.7 and a Km for ATP of 0.6 mM. There was an absolute requirement for Mg2+, with other divalent cations substituting less efficiently. When the Mg2+-dependent ATPase activity of intact cells was compared with that in homogenized cells, no significant difference was observed. The activity in intact cells was linear with respect to incubation time up to at least l0 min. Trypan blue staining and lactate dehydrogenase assays revealed that greater than 92% of the PMNL remained intact and viable during the assay. No soluble ATPase was released from the cells under assay conditions. In following the distribution of gamma[32P]ATP and 32P2 counts became cell associated. Since the experimental evidence supports the observation that PMNL remain intact and viable and that ATP does not penetrate the cell under assay conditions, it is proposed that greater than 90% of the Mg2+-dependent ATPase of the human PMNL is associated with a plasma membrnae enzyme. This would qualify the enzyme for the role of a plasma membrane marker for future fractionation and isolation attempts. | [
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|
PMID:14893 | Levan and levansucrase of Actinomyces viscosus. | A levansucrase was demonstrated in the growth medium and in association with the cell surface of Actinomyces viscosus. The amount of enzyme produced relative to cell density is not significantly affected by the growth conditions. Sugar alcohols inhibit growth of the cells. The levansucrase hydrolyzes sucrose to produce free glucose and levan; some free fructose is also formed. There is no requirement for cofactors. The Km for sucrose is 12 mM. A variety of heavy metal ions and two disaccharides, lactose and cellobiose, inhibit the enzyme. The levansucrase was purified to homogeneity and has a specific activity of 90 micronmol of glucose release per min per mg. The enzyme has a molecular weight of 220,000 and is composed of subunits of molecular weight 80,000. The levan product contains both beta(2 leads to 1) and beta(2 leads to 6) linkages. The enzyme remains tightly bound to the levan product, resulting in the formation of high-molecular-weight polymer on the order of 10(8) daltons. The possible role of the levan and levansucrase of A. viscosus in the pathogenesis of periodontal disease is discussed. | Levan and levansucrase of Actinomyces viscosus. A levansucrase was demonstrated in the growth medium and in association with the cell surface of Actinomyces viscosus. The amount of enzyme produced relative to cell density is not significantly affected by the growth conditions. Sugar alcohols inhibit growth of the cells. The levansucrase hydrolyzes sucrose to produce free glucose and levan; some free fructose is also formed. There is no requirement for cofactors. The Km for sucrose is 12 mM. A variety of heavy metal ions and two disaccharides, lactose and cellobiose, inhibit the enzyme. The levansucrase was purified to homogeneity and has a specific activity of 90 micronmol of glucose release per min per mg. The enzyme has a molecular weight of 220,000 and is composed of subunits of molecular weight 80,000. The levan product contains both beta(2 leads to 1) and beta(2 leads to 6) linkages. The enzyme remains tightly bound to the levan product, resulting in the formation of high-molecular-weight polymer on the order of 10(8) daltons. The possible role of the levan and levansucrase of A. viscosus in the pathogenesis of periodontal disease is discussed. | [
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|
PMID:14896 | Tumorigenicity of human hematopoietic cell lines in athymic nude mice. | Human hematopoietic cell lines, which had been classified on the basis of studies on clonality, and morphological, chromosomal and functional parameters as lymphoblastoid cell lines (LCL) of presumed non-neoplastic origin, and lymphoma, myeloma and leukemia lines of proven malignant origin, were tested for tumorigenic potential on subcutaneous transplantation to nude mice and for capacity to grow in semi-solid medium in vitro. Recently established LCL failed to grow both in nude mice and in agarose. In contrast, some of the LCL which had developed secondary chromosomal alterations during continuous cultivation for periods exceeding several years were tumorigenic and/or had the capacity to form colonies in agarose. Most lymphoma lines formed colonies in agarose and tumors in the mice. One of the two myeloma lines formed subcutaneous tumor which, however, showed no progressive growth. The other myeloma line failed to grow. Both myeloma lines, however, formed colonies in agarose. The myeloid leukemia line was tumorigenic while two of the three tested lymphocytic leukemia lines failed to grow in the mice. All leukemia lines formed colonies in agarose. We conclude from this study that: (1) Of the two types of Epstein-Barr virus containing cell lines [LCL and Burkitt lymphoma (BL) lines], only BL lines were shown to form tumors when inoculated subcutaneously in nude mice and had the capacity to grow in agarose in vitro. This shows that EBV transformation per se does not necessarily render lymphocytes tumorigenic in nude mice. The capacity to form colonies in agarose is not acquired either. (2) Changes of the karyotype and several phenotypic characteristics which occur in the originally diploid LCL during prolonged cultivation in vitro may be accompanied by the acquisition of the potential to grow subcutaneously in nude mice and in agarose in vitro. (3) The inconsistency with regard to the capacity of come of the neoplastic cell lines to grow in nude mice or in agarose seems to underline that neither of the two tests is a reliable criterion for malignancy of human lymphoma, leukemia and myeloma cell lines. | Tumorigenicity of human hematopoietic cell lines in athymic nude mice. Human hematopoietic cell lines, which had been classified on the basis of studies on clonality, and morphological, chromosomal and functional parameters as lymphoblastoid cell lines (LCL) of presumed non-neoplastic origin, and lymphoma, myeloma and leukemia lines of proven malignant origin, were tested for tumorigenic potential on subcutaneous transplantation to nude mice and for capacity to grow in semi-solid medium in vitro. Recently established LCL failed to grow both in nude mice and in agarose. In contrast, some of the LCL which had developed secondary chromosomal alterations during continuous cultivation for periods exceeding several years were tumorigenic and/or had the capacity to form colonies in agarose. Most lymphoma lines formed colonies in agarose and tumors in the mice. One of the two myeloma lines formed subcutaneous tumor which, however, showed no progressive growth. The other myeloma line failed to grow. Both myeloma lines, however, formed colonies in agarose. The myeloid leukemia line was tumorigenic while two of the three tested lymphocytic leukemia lines failed to grow in the mice. All leukemia lines formed colonies in agarose. We conclude from this study that: (1) Of the two types of Epstein-Barr virus containing cell lines [LCL and Burkitt lymphoma (BL) lines], only BL lines were shown to form tumors when inoculated subcutaneously in nude mice and had the capacity to grow in agarose in vitro. This shows that EBV transformation per se does not necessarily render lymphocytes tumorigenic in nude mice. The capacity to form colonies in agarose is not acquired either. (2) Changes of the karyotype and several phenotypic characteristics which occur in the originally diploid LCL during prolonged cultivation in vitro may be accompanied by the acquisition of the potential to grow subcutaneously in nude mice and in agarose in vitro. (3) The inconsistency with regard to the capacity of come of the neoplastic cell lines to grow in nude mice or in agarose seems to underline that neither of the two tests is a reliable criterion for malignancy of human lymphoma, leukemia and myeloma cell lines. | [
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|
PMID:14897 | Studies on Epstein-Barr virus-related antigens. II. Biochemical properties of soluble antigen in Raji Burkitt lymphoma cells. | Biochemical properties of Epstein-Barr virus (EBV)-related soluble antigen in non-producer Raji Burkitt lymphoblastoid cells, assayed by the indirect single radial immunodiffusion (ISRD) test, were investigated. The soluble ISRD antigen retained activity even after exposure to a temperature of 80 degrees C. The antigen was precipitated in 40% saturated ammonium sulfate and ISRD activity was recovered from the precipitate when reconstituted into a solution. Isoelectric focusing and crossed immunoelectrophoretic analyses revealed that the esoelectric point of the ISRD antigen is pH 4.8 with an electrophoretic mobility similar to that of serum alpha-globulin. With Sephadex G-200 or Sepharose 6B gel filtration, ISRD activity was obtained as a single peak which corresponded to the activity absorbing anti-complement immunofluorescence. The molecular weight of the present EBV-related soluble antigen was estimated to be 220,000-2408000 daltons. | Studies on Epstein-Barr virus-related antigens. II. Biochemical properties of soluble antigen in Raji Burkitt lymphoma cells. Biochemical properties of Epstein-Barr virus (EBV)-related soluble antigen in non-producer Raji Burkitt lymphoblastoid cells, assayed by the indirect single radial immunodiffusion (ISRD) test, were investigated. The soluble ISRD antigen retained activity even after exposure to a temperature of 80 degrees C. The antigen was precipitated in 40% saturated ammonium sulfate and ISRD activity was recovered from the precipitate when reconstituted into a solution. Isoelectric focusing and crossed immunoelectrophoretic analyses revealed that the esoelectric point of the ISRD antigen is pH 4.8 with an electrophoretic mobility similar to that of serum alpha-globulin. With Sephadex G-200 or Sepharose 6B gel filtration, ISRD activity was obtained as a single peak which corresponded to the activity absorbing anti-complement immunofluorescence. The molecular weight of the present EBV-related soluble antigen was estimated to be 220,000-2408000 daltons. | [
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|
PMID:14898 | Purification of several proteolytic enzymes by tosyl- and carbobenzoxy-triethylene-tetramine-sepharoses. | Tosyl-triethylenetetramine-Sepharose (Tos-T-Sepharose) and carbenzoxytriethylenetetramine-Sepharose (Z-T-Sepharose) were found to be adsorbents utilizable in the purification of several microbial and animal proteases. The former Sepharose derivative adsorbed alpha-chymotrypsin, trypsin, subtilisin, thermolysin and neutral subtilopeptidase at neutral pH range, and acid proteases such as pepsin and Rhizopus niveus protease at pH 3.5-6.5. alpha-Chymotrypsin and trypsin were eluted with 0.1 N acetic acid and Rhizopus protease with 0.5 N acetic acid, thermolysin with 1 M guanidine-HCl or 33% ethyleneglycol, whilst pepsin was recovered by elution with 2 M guanidine-HCl at pH 3.5. The binding of neutral subtilopeptidase and subtilisin to this adsorbent was comparatively weak and both the enzymes were recovered by elution with 0.5 M NaCl at neutral pH. On the other hand, Z-T-Sepharose was found to bind tightly to these proteolytic enzymes except neutral subtilopeptidase. Trypsin and alpha-chymotrypsin were released from the adsorbent column with 1 M p-toluenesulfonate, and subtilisin with 1 M guanidine-HCl or 33% ethyleneglycol at neutral pH region. By these chromatographic procedures, the specific activities of these proteolytic enzymes increased effectively. Comparison of the binding abilities of acetyl-, benzoyl-, tosyl- and carbobenzoxy-T-Sepharoses to these enzymes suggests that hydrophobicity of tosyl and carbobenzoxy groups plays an important role in the enzyme-adsorbent interaction. | Purification of several proteolytic enzymes by tosyl- and carbobenzoxy-triethylene-tetramine-sepharoses. Tosyl-triethylenetetramine-Sepharose (Tos-T-Sepharose) and carbenzoxytriethylenetetramine-Sepharose (Z-T-Sepharose) were found to be adsorbents utilizable in the purification of several microbial and animal proteases. The former Sepharose derivative adsorbed alpha-chymotrypsin, trypsin, subtilisin, thermolysin and neutral subtilopeptidase at neutral pH range, and acid proteases such as pepsin and Rhizopus niveus protease at pH 3.5-6.5. alpha-Chymotrypsin and trypsin were eluted with 0.1 N acetic acid and Rhizopus protease with 0.5 N acetic acid, thermolysin with 1 M guanidine-HCl or 33% ethyleneglycol, whilst pepsin was recovered by elution with 2 M guanidine-HCl at pH 3.5. The binding of neutral subtilopeptidase and subtilisin to this adsorbent was comparatively weak and both the enzymes were recovered by elution with 0.5 M NaCl at neutral pH. On the other hand, Z-T-Sepharose was found to bind tightly to these proteolytic enzymes except neutral subtilopeptidase. Trypsin and alpha-chymotrypsin were released from the adsorbent column with 1 M p-toluenesulfonate, and subtilisin with 1 M guanidine-HCl or 33% ethyleneglycol at neutral pH region. By these chromatographic procedures, the specific activities of these proteolytic enzymes increased effectively. Comparison of the binding abilities of acetyl-, benzoyl-, tosyl- and carbobenzoxy-T-Sepharoses to these enzymes suggests that hydrophobicity of tosyl and carbobenzoxy groups plays an important role in the enzyme-adsorbent interaction. | [
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PMID:14899 | The acylation of bovine serum albumin with diacetylcycloserine. | The reaction of the amino groups of bovine serum albumin (BSA) with diacetylcycloserine (I) at pH 7.2-9.0 proceeded with both acylation by the diacetyl-beta-aminooxy-D-alanyl (DAA) group and acetylation. The number of DAA groups was determined by their conversion to cycloserine (III) which can be accurately measured in micromolar amounts. The method was developed using the model compound diacetyl-beta-aminooxy-D-alanine butyl amide (II) which was converted to beta-aminooxy-D-alanine butyl amide (IV) by methanolysis and then to cycloserine by basic ring-closure of IV. Calculations using results obtained by this method combined with the experimentally determined number of free amino groups in the modified BSA indicated that the reaction of excess I with BSA effected the acetylation of about 35 and the acylation with DAA groups of about 22 of the 59 amino groups. These findings were supported by experiments demonstrating that the amount of acetic acid formed by hydrolysis of the modified BSA was approximately that predicted from the results of the cycloserine analyses. | The acylation of bovine serum albumin with diacetylcycloserine. The reaction of the amino groups of bovine serum albumin (BSA) with diacetylcycloserine (I) at pH 7.2-9.0 proceeded with both acylation by the diacetyl-beta-aminooxy-D-alanyl (DAA) group and acetylation. The number of DAA groups was determined by their conversion to cycloserine (III) which can be accurately measured in micromolar amounts. The method was developed using the model compound diacetyl-beta-aminooxy-D-alanine butyl amide (II) which was converted to beta-aminooxy-D-alanine butyl amide (IV) by methanolysis and then to cycloserine by basic ring-closure of IV. Calculations using results obtained by this method combined with the experimentally determined number of free amino groups in the modified BSA indicated that the reaction of excess I with BSA effected the acetylation of about 35 and the acylation with DAA groups of about 22 of the 59 amino groups. These findings were supported by experiments demonstrating that the amount of acetic acid formed by hydrolysis of the modified BSA was approximately that predicted from the results of the cycloserine analyses. | [
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PMID:14900 | Iodinated radiological contrast media as radiosensitizers. | This paper describes the radiosensitizing effects of diatrizoic (DA) and iothalamic (ITA) acids and of iodipamide (IP) on the survival of E coli B/r irradiated with X-rays and with high-intensity electron pulses. All compounds at concentrations between 10 and 50 mM display a strong sensitizing effect in the presence of oxygen (DMF between 0-1 and 0-3) and are much less effective in nitrogen. In N2O the degree of sensitization is intermediate between oxygen and nitrogen. The situation is the same at pH 7 or 5-6. Solutions of DA, ITA and IP irradiated at pH lower than 6 become highly toxic to bacteria added after irradiation, for several hours after X-irradiation or several minutes after pulsed irradiation. The maximum toxic effect occurs with 2 krad of X-ray and with 6-8 krad of electrons. Oxygen must be present in order to observe the bactericidal activity. This is not affected by scavenging hydrated electrons with nitrate, but is completely cancelled by scavenging OH radicals with formate. It is also cancelled by adding thiosulphate to the irradiated solutions immediately before the bacteria. In the presence of nutrient broth, the radiosensitizing effect is absent after irradiation with pulsed electrons; whereas after X-irradiation it is reduced when the concentration of sensitizers is 50 mM. The experimental data appear to be compatible with a mechanism operated by short and long-lived transients resulting from the radiolysis of iodinated contrast media. | Iodinated radiological contrast media as radiosensitizers. This paper describes the radiosensitizing effects of diatrizoic (DA) and iothalamic (ITA) acids and of iodipamide (IP) on the survival of E coli B/r irradiated with X-rays and with high-intensity electron pulses. All compounds at concentrations between 10 and 50 mM display a strong sensitizing effect in the presence of oxygen (DMF between 0-1 and 0-3) and are much less effective in nitrogen. In N2O the degree of sensitization is intermediate between oxygen and nitrogen. The situation is the same at pH 7 or 5-6. Solutions of DA, ITA and IP irradiated at pH lower than 6 become highly toxic to bacteria added after irradiation, for several hours after X-irradiation or several minutes after pulsed irradiation. The maximum toxic effect occurs with 2 krad of X-ray and with 6-8 krad of electrons. Oxygen must be present in order to observe the bactericidal activity. This is not affected by scavenging hydrated electrons with nitrate, but is completely cancelled by scavenging OH radicals with formate. It is also cancelled by adding thiosulphate to the irradiated solutions immediately before the bacteria. In the presence of nutrient broth, the radiosensitizing effect is absent after irradiation with pulsed electrons; whereas after X-irradiation it is reduced when the concentration of sensitizers is 50 mM. The experimental data appear to be compatible with a mechanism operated by short and long-lived transients resulting from the radiolysis of iodinated contrast media. | [
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PMID:14906 | The activity and distribution of gamma-glutamyl transpeptidase (y-GT) in human foetal organs. | The activity and distribution of y-GT was investigated in a number of organs from human foetuses aged from 14 to 24 weeks post menstruationem. Over this period, enzyme activity increased in the kidney, pancreas and thymus, but decreased in the small intestine. No trend could be established for the liver, although activity was high. In the lung, spleen, brain and adrenals, y-GT was either detectable at very low levels or could not be demonstrated. The possible relationship between y-GT activity in some human tumours and the enzyme level in the corresponding foetal organs is discussed. | The activity and distribution of gamma-glutamyl transpeptidase (y-GT) in human foetal organs. The activity and distribution of y-GT was investigated in a number of organs from human foetuses aged from 14 to 24 weeks post menstruationem. Over this period, enzyme activity increased in the kidney, pancreas and thymus, but decreased in the small intestine. No trend could be established for the liver, although activity was high. In the lung, spleen, brain and adrenals, y-GT was either detectable at very low levels or could not be demonstrated. The possible relationship between y-GT activity in some human tumours and the enzyme level in the corresponding foetal organs is discussed. | [
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PMID:14904 | Subcellular distribution of pyridoxal kinase in ox retina. | PL kinase activity has been determined in primary and secondary subcellular fractions of ox retina. Enzymic activity is predominantly located in the soluble fraction (S3). About 65% of the recovered PL kinase activity of crude mitochondria is released from synaptosomal fraction after hypoosmotic treatment. PL kinase activity in supernatant fraction (S3) does not exceed the enzyme activity measured in the homogenate, excluding the presence of a particulate-bound inhibitor to PL kinase in the homogenate. The possible physiological significance of PL kinase cellular compartmentation has been considered. | Subcellular distribution of pyridoxal kinase in ox retina. PL kinase activity has been determined in primary and secondary subcellular fractions of ox retina. Enzymic activity is predominantly located in the soluble fraction (S3). About 65% of the recovered PL kinase activity of crude mitochondria is released from synaptosomal fraction after hypoosmotic treatment. PL kinase activity in supernatant fraction (S3) does not exceed the enzyme activity measured in the homogenate, excluding the presence of a particulate-bound inhibitor to PL kinase in the homogenate. The possible physiological significance of PL kinase cellular compartmentation has been considered. | [
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|
PMID:14919 | Purification and properties of glutamate synthase from Thiobacillus thioparus. | Glutamate synthase was purified about 250-fold from Thiobacillus thioparus and was characterized. The molecular weight was estimated as 280,000 g/mol. The enzyme showed absorption maxima at 280, 380, and 450 nm and was inhibited by Atebrin, suggesting that T. thioparus glutamate synthase is a flavoprotein. The enzyme activity was also inhibited by iron chelators and thiolbinding agents. The enzyme was specific for reduced nicotinamide adenine dinucleotide phosphate (NADPH) and alpha-ketoglutarate, but L-glutamine was partially replaced by ammonia as the amino donor. The Km values of glutamate synthase for NADPH, alpha-ketoglutarate, and glutamine were 3.0 muM, 50 muM, and 1.1 mM, respectively. The enzyme had a pH optimum between 7.3 and 7.8. Glutamate synthase from T. thioparus was relatively insensitive to feedback inhibition by single amino acids but was sensitive to the combined effects of several amino acids. Enzymes involved in glutamate synthesis in T. thioparus were studied. Glutamine synthetase and glutamate synthase, as well as two glutamate dehydrogenases (NADH and NADPH dependent), were present in this organism. This levels of glutamate synthase and glutamate dehydrogenase were similar in T. thioparus grown on 0.7 or 7.0 mM ammonium sulfate. The sum of the activities of both glutamate dehydrogenases was only 1/25 of that of glutamate synthase under the assay conditions. It was concluded that the glutamine pathway is important for ammonia assimilation in this autotrophic bacterium. | Purification and properties of glutamate synthase from Thiobacillus thioparus. Glutamate synthase was purified about 250-fold from Thiobacillus thioparus and was characterized. The molecular weight was estimated as 280,000 g/mol. The enzyme showed absorption maxima at 280, 380, and 450 nm and was inhibited by Atebrin, suggesting that T. thioparus glutamate synthase is a flavoprotein. The enzyme activity was also inhibited by iron chelators and thiolbinding agents. The enzyme was specific for reduced nicotinamide adenine dinucleotide phosphate (NADPH) and alpha-ketoglutarate, but L-glutamine was partially replaced by ammonia as the amino donor. The Km values of glutamate synthase for NADPH, alpha-ketoglutarate, and glutamine were 3.0 muM, 50 muM, and 1.1 mM, respectively. The enzyme had a pH optimum between 7.3 and 7.8. Glutamate synthase from T. thioparus was relatively insensitive to feedback inhibition by single amino acids but was sensitive to the combined effects of several amino acids. Enzymes involved in glutamate synthesis in T. thioparus were studied. Glutamine synthetase and glutamate synthase, as well as two glutamate dehydrogenases (NADH and NADPH dependent), were present in this organism. This levels of glutamate synthase and glutamate dehydrogenase were similar in T. thioparus grown on 0.7 or 7.0 mM ammonium sulfate. The sum of the activities of both glutamate dehydrogenases was only 1/25 of that of glutamate synthase under the assay conditions. It was concluded that the glutamine pathway is important for ammonia assimilation in this autotrophic bacterium. | [
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PMID:14920 | Pyrrolidone carboxylyl peptidase in Streptococcus cremoris: dependence on an interaction with membrane components. | A study of the distribution of pyrrolidone carboxylyl peptidase (PCP) activity among cell fractions of Streptococcus cremoris HP revealed that this enzyme is associated with a particulate fraction, which mainly consists of membrane material. This location could only be established using a gentle nonmechanical method for the disruption of spheroplasts under the conditions of which intracellular marker enzymes are released. The effect of monovalent anions and treatments, which do not destroy covalent binding, suggests an association of the enzyme with surrounding structures determined by both hydrophobic and electrostatic interactions. The activity of PCP associated with cells harvested from different growth phases and in the solubilized state was studied as a function of the temperature in the absence and in the presence of the membrane-interfering agent n-butanol. A decrease in the apparent activation energy, inherent to the solubilized enzyme, is induced in situ at a lower transition temperature. Only with logarithmic-phase cells is this transition followed (mid-logarithmic cells) or accompanied (late logarithmic cells) by a secondary decrease in the energy of activation. n-Butanol appeared to decrease the lower transition temperature of the enzyme activity in situ, and additionally it exerted an effect on the manifestation of the secondary transition. Thecorganization of membrane components, mainly the lipids. | Pyrrolidone carboxylyl peptidase in Streptococcus cremoris: dependence on an interaction with membrane components. A study of the distribution of pyrrolidone carboxylyl peptidase (PCP) activity among cell fractions of Streptococcus cremoris HP revealed that this enzyme is associated with a particulate fraction, which mainly consists of membrane material. This location could only be established using a gentle nonmechanical method for the disruption of spheroplasts under the conditions of which intracellular marker enzymes are released. The effect of monovalent anions and treatments, which do not destroy covalent binding, suggests an association of the enzyme with surrounding structures determined by both hydrophobic and electrostatic interactions. The activity of PCP associated with cells harvested from different growth phases and in the solubilized state was studied as a function of the temperature in the absence and in the presence of the membrane-interfering agent n-butanol. A decrease in the apparent activation energy, inherent to the solubilized enzyme, is induced in situ at a lower transition temperature. Only with logarithmic-phase cells is this transition followed (mid-logarithmic cells) or accompanied (late logarithmic cells) by a secondary decrease in the energy of activation. n-Butanol appeared to decrease the lower transition temperature of the enzyme activity in situ, and additionally it exerted an effect on the manifestation of the secondary transition. Thecorganization of membrane components, mainly the lipids. | [
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|
PMID:14921 | Partial purification of the Escherichia coli K-12 mec+ deoxyribonucleic acid-cytosine methylase: in vitro methylation completely protects bacteriophage lambda deoxyribonucleic acid against cleavage by R-EcoRII. | A procedure is described for the partial purification of the deoxyribonucleic acid (DNA)-cytosine methylases controlled by the RII plasmid and by the Escherichia coli mec+ gene. The two enzymes exhibit similar but distinct chromatographic behavior on diethylaminoethyl-cellulose and phosphocellulose. Preliminary studies on the two methylases indicate that they are indistinguishable with respect to their Km for S-adenosylmethionine and their pH (in tris (hydroxymethyl)aminomethane buffer) and NaCl concentration optima. In vitro methylation of various phage lambda DNA substrates by the mec'r RII enzyme modifies the DNA to a form that is completely resistant to double-stranded cleavage by the RII restriction endonuclease (R-EcoRII). These results are consistent with our earlier proposal that the mec8ethylase recognizes RII host specificity sites. | Partial purification of the Escherichia coli K-12 mec+ deoxyribonucleic acid-cytosine methylase: in vitro methylation completely protects bacteriophage lambda deoxyribonucleic acid against cleavage by R-EcoRII. A procedure is described for the partial purification of the deoxyribonucleic acid (DNA)-cytosine methylases controlled by the RII plasmid and by the Escherichia coli mec+ gene. The two enzymes exhibit similar but distinct chromatographic behavior on diethylaminoethyl-cellulose and phosphocellulose. Preliminary studies on the two methylases indicate that they are indistinguishable with respect to their Km for S-adenosylmethionine and their pH (in tris (hydroxymethyl)aminomethane buffer) and NaCl concentration optima. In vitro methylation of various phage lambda DNA substrates by the mec'r RII enzyme modifies the DNA to a form that is completely resistant to double-stranded cleavage by the RII restriction endonuclease (R-EcoRII). These results are consistent with our earlier proposal that the mec8ethylase recognizes RII host specificity sites. | [
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|
PMID:14922 | Sterol 24(28) methylene reductase in Saccharomyces cerevisiae. | Optimal conditions for the 24(28)methylene reductase were obtained. The enzyme assay provided for unusually high activity; the Km was determined to be 10.8 mum. The enzyme activity was increased in cells grown with ethanol as the substrate. | Sterol 24(28) methylene reductase in Saccharomyces cerevisiae. Optimal conditions for the 24(28)methylene reductase were obtained. The enzyme assay provided for unusually high activity; the Km was determined to be 10.8 mum. The enzyme activity was increased in cells grown with ethanol as the substrate. | [
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|
PMID:14923 | Ornithine transcarbamylase from Salmonella typhimurium: purification, subunit composition, kinetic analysis, and immunological cross-reactivity. | Ornithine transcarbamylase (OTCase) was purified to hemogeneity from a derepressed strain of Salmonella typhimurium. The optimal pH for enzyme activity is 8.0. The molecular weight of the enzyme was calculated to be 116,000, based on measurements of the sedimentation coefficient by sucrose gradient ultracentrifugation and the Stokes radius by gel filtration. Polyacrylamide gel electrophoresis of cross-linked OTCase in the presence of sodium dodecyl sulfate showed that the enzyme is composed of three identical subunits. The molecular weight of the monomer was determined to be 39,000. Steady-state kinetics indicate that the reaction mechanism is sequential. The limiting Michealis constants for carbamylphosphate and ornithine were determined to be 0.06 and 0.2 mM, respectively. The dissociation constant for carbamylphosphate was 0.02 mM. Product and dead-end inhibition patterns are consistent with an ordered Bi Bi mechanism, in which carbamylphosphate is the first substrate added and phosphate is the last product released. OTCase activity was inhibited by arginine, but relatively high concentrations were required for significant inhibition. The inhibition by arginine might be physiologically significant in the regulation of carbamlphosphate utilization; a single carbamylphosphate synthetase is responsible for the synthesis of carbamylphosphate for both arginine and pyrimidines in S. typhimurium and the inhibition by argine might serve to divert carbamlphosphate to the synthesis of pyrimidines when arginine is present at high concentrations. The crossreaction of OTCases from different microorganisms with purified antibodies raised against the homogeneous OTCase from S. typhimurium was investigated. The results of immunotitration and immunodiffusion experiments revealed a high degree of identity between the enzymes form S. typhimurium and Esherichia coli B and W. In these three cases, a single gen (argl) encodes OTCase. Wild-type E. coli K-12 and strain 3000 X 111, which carry two OTCase genes (argI, argF), also revealed similar cross-reactivity, supporting the hypothesis that argF is the product of a relatively recent duplication. The activity of OTCase from Bacillus subtilis was partially inhibited by antibodies against the enzyme from S. typhimurium, indicating unusual conservation of primary structure among widely different taxonomic groups. OTCase from Saccharomyces cerevisiae, whose molecular weight and primary structure are similar to those of the enzyme from S. typhimurium, was without detectable cross-reactivity. | Ornithine transcarbamylase from Salmonella typhimurium: purification, subunit composition, kinetic analysis, and immunological cross-reactivity. Ornithine transcarbamylase (OTCase) was purified to hemogeneity from a derepressed strain of Salmonella typhimurium. The optimal pH for enzyme activity is 8.0. The molecular weight of the enzyme was calculated to be 116,000, based on measurements of the sedimentation coefficient by sucrose gradient ultracentrifugation and the Stokes radius by gel filtration. Polyacrylamide gel electrophoresis of cross-linked OTCase in the presence of sodium dodecyl sulfate showed that the enzyme is composed of three identical subunits. The molecular weight of the monomer was determined to be 39,000. Steady-state kinetics indicate that the reaction mechanism is sequential. The limiting Michealis constants for carbamylphosphate and ornithine were determined to be 0.06 and 0.2 mM, respectively. The dissociation constant for carbamylphosphate was 0.02 mM. Product and dead-end inhibition patterns are consistent with an ordered Bi Bi mechanism, in which carbamylphosphate is the first substrate added and phosphate is the last product released. OTCase activity was inhibited by arginine, but relatively high concentrations were required for significant inhibition. The inhibition by arginine might be physiologically significant in the regulation of carbamlphosphate utilization; a single carbamylphosphate synthetase is responsible for the synthesis of carbamylphosphate for both arginine and pyrimidines in S. typhimurium and the inhibition by argine might serve to divert carbamlphosphate to the synthesis of pyrimidines when arginine is present at high concentrations. The crossreaction of OTCases from different microorganisms with purified antibodies raised against the homogeneous OTCase from S. typhimurium was investigated. The results of immunotitration and immunodiffusion experiments revealed a high degree of identity between the enzymes form S. typhimurium and Esherichia coli B and W. In these three cases, a single gen (argl) encodes OTCase. Wild-type E. coli K-12 and strain 3000 X 111, which carry two OTCase genes (argI, argF), also revealed similar cross-reactivity, supporting the hypothesis that argF is the product of a relatively recent duplication. The activity of OTCase from Bacillus subtilis was partially inhibited by antibodies against the enzyme from S. typhimurium, indicating unusual conservation of primary structure among widely different taxonomic groups. OTCase from Saccharomyces cerevisiae, whose molecular weight and primary structure are similar to those of the enzyme from S. typhimurium, was without detectable cross-reactivity. | [
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|
PMID:14924 | Properties of an inducible extracellular neuraminidase from an Arthrobacter isolate. | The elective isolation of a soil microorganism, tentatively assigned to the genus Arthrobacter, which produced an extracellular neuraminidase is described. The secretion of neuraminidase from washed cells in minimal medium required the presence of sialo-containing glycoproteins, whereas free N-acetyl-neuraminic asid of N-acetylmannosamine were poor inducers. No enzyme could be dected in the induction fitrated of cells, in the absence of inducer or in the culture filtrate of cells grown in a complete medium. The routine enzyme inducer was a hot-water extract of "edible bird's nest." Mild acid treatment (0.05 N H2SO4) of this extract increased enzyme activity two--to threefold and the specific activity about eightfold. Neuraminidase induction with acid-treated bird's nest was manifested at a linear rate for 6 h without increase in cell number. No other anticipated glycohydrolase or protease activities were foud. The amount of enzyme located within the cells was barely detectable as compared to that found in the induction filtrate. Experiments with chloramphenicol or chlortetracycline indicate that de novo protein synthesis was required for neuraminidase production and that this exoenzyme was not released from a preformed pool. Neuraminidase from this source has an apparent molecular weight of 87,000, a pH optimum of 5 to 6, and an apparent Km of 2.08 mg/ml for collocalia mucoid and 3.3 X 10(-3) M for N-acetylneuraminlactose and is insensitive both to Ca2+ ions and ethylenediaminetetraacetic acid. Preliminary studies indicate that the enzyme can hydrolyze alpha-2,3-, alpha-2,6-, or alph-2-8-N-acetylneuraminylglycosidic linkages. From total activity data and purification criteria, it would appear that this isolate can produce about 5 mg of enzyme per liter of induction medium. | Properties of an inducible extracellular neuraminidase from an Arthrobacter isolate. The elective isolation of a soil microorganism, tentatively assigned to the genus Arthrobacter, which produced an extracellular neuraminidase is described. The secretion of neuraminidase from washed cells in minimal medium required the presence of sialo-containing glycoproteins, whereas free N-acetyl-neuraminic asid of N-acetylmannosamine were poor inducers. No enzyme could be dected in the induction fitrated of cells, in the absence of inducer or in the culture filtrate of cells grown in a complete medium. The routine enzyme inducer was a hot-water extract of "edible bird's nest." Mild acid treatment (0.05 N H2SO4) of this extract increased enzyme activity two--to threefold and the specific activity about eightfold. Neuraminidase induction with acid-treated bird's nest was manifested at a linear rate for 6 h without increase in cell number. No other anticipated glycohydrolase or protease activities were foud. The amount of enzyme located within the cells was barely detectable as compared to that found in the induction filtrate. Experiments with chloramphenicol or chlortetracycline indicate that de novo protein synthesis was required for neuraminidase production and that this exoenzyme was not released from a preformed pool. Neuraminidase from this source has an apparent molecular weight of 87,000, a pH optimum of 5 to 6, and an apparent Km of 2.08 mg/ml for collocalia mucoid and 3.3 X 10(-3) M for N-acetylneuraminlactose and is insensitive both to Ca2+ ions and ethylenediaminetetraacetic acid. Preliminary studies indicate that the enzyme can hydrolyze alpha-2,3-, alpha-2,6-, or alph-2-8-N-acetylneuraminylglycosidic linkages. From total activity data and purification criteria, it would appear that this isolate can produce about 5 mg of enzyme per liter of induction medium. | [
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|
PMID:14925 | Motility of Mycoplasma pneumoniae. | Cell of Mycoplasma pneumoniae FH gliding on a glass surface in liquid medium were examined by microscopic observation and quantitatively by microcinematography (30 frames per min). Comparisons were made only within the individual experiments. The cells moved in an irregular pattern with numerous narrow bends and circles. They never changed their leading end. The average speed (without pauses) was relatively constant between o.2 and 0.5 mum/s. The maximum speed was about 1.5 to 2.0 mum/s. The movements were interrupted by resting periods of different lengths and frequency. Temperature, viscosity, pH, and the presence of yeast extract in the medium influenced the motility significantly; changes in glucose, calcium ions, and serum content were less effective. The movements were affected by iodoacetate, p-mercuribenzoate, and mitomycin C at inhibitory or subinhibitory concentrations. Sodium fluoride, sodium cyanide, dinitrophenol, chloramphenicol, puromycin, cholchicin, and cytochalasin B at minimal inhibitory concentrations did not affect motility. The movements were effectively inhibited by anti-M. pneumoniae antiserum. Studies with absorbed antiserum suggested that the surface components involved in motility are heat labile. The gliding of M. pneumoniae cells required an intact energy metabolism and the proteins involved seemed to have a low turnover. | Motility of Mycoplasma pneumoniae. Cell of Mycoplasma pneumoniae FH gliding on a glass surface in liquid medium were examined by microscopic observation and quantitatively by microcinematography (30 frames per min). Comparisons were made only within the individual experiments. The cells moved in an irregular pattern with numerous narrow bends and circles. They never changed their leading end. The average speed (without pauses) was relatively constant between o.2 and 0.5 mum/s. The maximum speed was about 1.5 to 2.0 mum/s. The movements were interrupted by resting periods of different lengths and frequency. Temperature, viscosity, pH, and the presence of yeast extract in the medium influenced the motility significantly; changes in glucose, calcium ions, and serum content were less effective. The movements were affected by iodoacetate, p-mercuribenzoate, and mitomycin C at inhibitory or subinhibitory concentrations. Sodium fluoride, sodium cyanide, dinitrophenol, chloramphenicol, puromycin, cholchicin, and cytochalasin B at minimal inhibitory concentrations did not affect motility. The movements were effectively inhibited by anti-M. pneumoniae antiserum. Studies with absorbed antiserum suggested that the surface components involved in motility are heat labile. The gliding of M. pneumoniae cells required an intact energy metabolism and the proteins involved seemed to have a low turnover. | [
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|
PMID:14926 | Purification and some properties of an extracellular alpha-amylase from Bacteroides amylophilus. | A medium was developed to obtain maximum yields of extracellular amylase from Bacteroides amylophilus 70. Crude enzyme preparation, obtained by ammonium sulfate precipitation of cell-free broth, contained six amylolytic isoenzymes that were detected by isoelectric focusing and polyacrylamide gel electrophoresis. One of these amylases was purified by diethylaminoethyl-Sephadex A-50 ion-exchange chromatography and Sephadex G-200 gel filtration techniques. Some properties of the purified extracellular alpha-amylase were: optimum pH, 6.3; optimum temperature, 43 degrees C: PH stability range, 5.8 to 7.5; isoelectric point, pH 4.6; molecular weight, 92,000 (by sodium dodecyl sulfatedisc gel electrophoresis); and sugars causing inhibition, cyclomaltoheptaose, cyclomaltohexaose, and alpha-d-phenylglucoside. In addition, Ca2+ and Co2+ were strong activators,and Hg2+ was a strong inhibitior; all other cations were slightly stimulatory. Dialysis against 0.01 M ethylenediaminetetraacetic acid caused a 58% loss of activity that was restored to 92% of the original by the addition of 0.04 M Ca2+. The enzyme affected a blue-value-reducing-value curve characteristic of alpha-type amylases. The relative rates of hydrolysis of amylose, soluble starch, amylopectin, and dextrin were 100, 97, 92, and 60%, respectively; Michaelis constants for these substrates were 18.2, 18.7, 18.2, and 16.7 mumol of d-glucosidic bond/liter, respectively. The enzyme degraded maize (corn) starch granules to some extent and had relatively little activity on potato starch granules. | Purification and some properties of an extracellular alpha-amylase from Bacteroides amylophilus. A medium was developed to obtain maximum yields of extracellular amylase from Bacteroides amylophilus 70. Crude enzyme preparation, obtained by ammonium sulfate precipitation of cell-free broth, contained six amylolytic isoenzymes that were detected by isoelectric focusing and polyacrylamide gel electrophoresis. One of these amylases was purified by diethylaminoethyl-Sephadex A-50 ion-exchange chromatography and Sephadex G-200 gel filtration techniques. Some properties of the purified extracellular alpha-amylase were: optimum pH, 6.3; optimum temperature, 43 degrees C: PH stability range, 5.8 to 7.5; isoelectric point, pH 4.6; molecular weight, 92,000 (by sodium dodecyl sulfatedisc gel electrophoresis); and sugars causing inhibition, cyclomaltoheptaose, cyclomaltohexaose, and alpha-d-phenylglucoside. In addition, Ca2+ and Co2+ were strong activators,and Hg2+ was a strong inhibitior; all other cations were slightly stimulatory. Dialysis against 0.01 M ethylenediaminetetraacetic acid caused a 58% loss of activity that was restored to 92% of the original by the addition of 0.04 M Ca2+. The enzyme affected a blue-value-reducing-value curve characteristic of alpha-type amylases. The relative rates of hydrolysis of amylose, soluble starch, amylopectin, and dextrin were 100, 97, 92, and 60%, respectively; Michaelis constants for these substrates were 18.2, 18.7, 18.2, and 16.7 mumol of d-glucosidic bond/liter, respectively. The enzyme degraded maize (corn) starch granules to some extent and had relatively little activity on potato starch granules. | [
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|
PMID:14927 | Formation of glutamine from [13n]ammonia, [13n]dinitrogen, and [14C]glutamate by heterocysts isolated from Anabaena cylindrica. | A method is described for the isolation of metabolically active heterocysts from Anabaena cylindrica. These isolated heterocysts accounted for up to 34% of the acetylene-reducing activity of whole filaments and had a specific activity of up to 1,560 nmol of C2H4 formed per mg of heterocyst chlorphyll per min. Activity of glutamine synthetase was coupled to activity of nitrogenase in isolated heterocysts as shown by acetylene-inhibitable formation of [13N]NH3 and of amidelabeled [13N]glutamine form [13N]N2. A method is also described for the production of 6-mCi amounts of [13N]NH3. Isolated heterocysts formed [13N]glutamine from [13N]NH3 and glutamate, and [14C]glutamine from NH3 and [14C]glutamate, in the presence of magnesium adenosine 5'-triphosphate. Methionine sulfoximine strongly inhibited these syntheses. Glutamate synthase is, after nitrogenase and glutamine synthetase, the third sequential enzyme involved in the assimilation of N2 by intact filaments. However, the kinetics of solubilization of the activity of glutamate synthase during cavitation of suspensions of A. cylindrica indicated that very little, if any, of the activity of that enzyme was located in heterocysts. Concordantly, isolated heterocysts failed to form substantial amounts of radioactive glutamate from either [13N]glutamine or alph-[14C]ketoglutarate in the presence of other substrates and cofactors of the glutamate synthase reaction. However, they formed [14C]glutamate rapidly from alpha-[14C]ketoglutarate by aminotransferase reactions, with various amino acids as the nitrogen donor. The implication of these findings with regard to the identities of the substances moving between heterocysts and vegetative cells are discussed. | Formation of glutamine from [13n]ammonia, [13n]dinitrogen, and [14C]glutamate by heterocysts isolated from Anabaena cylindrica. A method is described for the isolation of metabolically active heterocysts from Anabaena cylindrica. These isolated heterocysts accounted for up to 34% of the acetylene-reducing activity of whole filaments and had a specific activity of up to 1,560 nmol of C2H4 formed per mg of heterocyst chlorphyll per min. Activity of glutamine synthetase was coupled to activity of nitrogenase in isolated heterocysts as shown by acetylene-inhibitable formation of [13N]NH3 and of amidelabeled [13N]glutamine form [13N]N2. A method is also described for the production of 6-mCi amounts of [13N]NH3. Isolated heterocysts formed [13N]glutamine from [13N]NH3 and glutamate, and [14C]glutamine from NH3 and [14C]glutamate, in the presence of magnesium adenosine 5'-triphosphate. Methionine sulfoximine strongly inhibited these syntheses. Glutamate synthase is, after nitrogenase and glutamine synthetase, the third sequential enzyme involved in the assimilation of N2 by intact filaments. However, the kinetics of solubilization of the activity of glutamate synthase during cavitation of suspensions of A. cylindrica indicated that very little, if any, of the activity of that enzyme was located in heterocysts. Concordantly, isolated heterocysts failed to form substantial amounts of radioactive glutamate from either [13N]glutamine or alph-[14C]ketoglutarate in the presence of other substrates and cofactors of the glutamate synthase reaction. However, they formed [14C]glutamate rapidly from alpha-[14C]ketoglutarate by aminotransferase reactions, with various amino acids as the nitrogen donor. The implication of these findings with regard to the identities of the substances moving between heterocysts and vegetative cells are discussed. | [
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|
PMID:14928 | Measurement of the pH of frozen buffer solutions by using pH indicators. | A method was established to estimate the pH change of several buffers solutions on freezing by using a combination of pH indicators. Among more than 30 buffers solutions examined, almost half exhibited a pH change in the temperature range between freezing point and 220 degrees K; the results were tabulated. Glycerol was found to suppress the pH changes because of its "salt buffer" effect. | Measurement of the pH of frozen buffer solutions by using pH indicators. A method was established to estimate the pH change of several buffers solutions on freezing by using a combination of pH indicators. Among more than 30 buffers solutions examined, almost half exhibited a pH change in the temperature range between freezing point and 220 degrees K; the results were tabulated. Glycerol was found to suppress the pH changes because of its "salt buffer" effect. | [
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|
PMID:14929 | Mathematical analysis of ligand-induced monomerization and dimerization in the monomer dimer equilibrium of proteins. Application to D-amino acid oxidase. | We have mathematically analyzed ligand-induced monomerization and dimerization in a protein monomer-dimer equilibrium system, in which the monomer has one and the dimer two binding sites. These dimer sites have the same binding constants for the first ligand but may cooperatively interact when one of them is occupied by a ligand molecule. In this system, the apparent dimerization constant and the apparent molecular weight are functions of free ligand concentration, and depend on the intrinsic binding constants of the ligand molecule to the monomer and the dimer. The behavior of these functions is classified into 17 cases according to the values of the three intrinsic binding constants, and some calculated examples are shown graphically for selected parameters. The theory was also applied to D-amino acid oxidase [EC 1.4.3.3], a flavoprotein, and the pH dependence of the apparent dimerization constant and the apparent molecular weight in the presence of ligand, p-aminobenzoate, were studied theoretically using parameters obtained in our previous experiments (5). | Mathematical analysis of ligand-induced monomerization and dimerization in the monomer dimer equilibrium of proteins. Application to D-amino acid oxidase. We have mathematically analyzed ligand-induced monomerization and dimerization in a protein monomer-dimer equilibrium system, in which the monomer has one and the dimer two binding sites. These dimer sites have the same binding constants for the first ligand but may cooperatively interact when one of them is occupied by a ligand molecule. In this system, the apparent dimerization constant and the apparent molecular weight are functions of free ligand concentration, and depend on the intrinsic binding constants of the ligand molecule to the monomer and the dimer. The behavior of these functions is classified into 17 cases according to the values of the three intrinsic binding constants, and some calculated examples are shown graphically for selected parameters. The theory was also applied to D-amino acid oxidase [EC 1.4.3.3], a flavoprotein, and the pH dependence of the apparent dimerization constant and the apparent molecular weight in the presence of ligand, p-aminobenzoate, were studied theoretically using parameters obtained in our previous experiments (5). | [
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|
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