An investigation into nonscientist involvement in peer review: the National Institutes of Health example.
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Biomedical subjects
Publications and source records attributed to M Zucker.
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Serial haematological investigations were carried out in 5 patients with Fanconi's anaemia over periods of 6 months--11 years. All the patients were pancytopenic with a depression of the granulocytic and megakaryocytic elements of the bone marrow throughout the greater part of their illnesses. Erythropoietic acitvity was variable. The initial bone marrow examination revealed depressed erythroid function in 3 patients. The erythroid hypoplasia persisted in 2 of them, while in the third, erythroid activity increased with time, possibly as the result of therapy with oxymetholone. Erythroid hyperplasia was present in the remaining 2 patients, both at presentation and throughout the course of the illness. This could not be attributed to treatment in either patient. Six erythrokinetic studies were carried out in the 5 patients at variable intervals after the diagnosis had been made. In 2 studies erythroid activity was unequivocally depressed, while in a further 3 a significant, though probably suboptimal, erythroid marrow response was present. In the final study erythropoiesis was increased but was markedly ineffective in terms of the delivery of viable red cells into the circulation. In vivo counting suggested that some degree of ineffective erythropoiesis was also present in the other patients and studies with 51Cr indicated a shortened red cell survival in all subjects studied. In 2 of them significant splenic sequestration was present. Leukaemic transformation occurred in 2 patients. In 1 of them its development was heralded by the appearance of micromegakaryocytes in the bone marrow.
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The increase in phenylalanine ammonia-lyase (PAL) activity in strawberry (Fragaria vesca var. WSU-1232) leaf disks required wounding, sucrose, and light and was cycloheximide-sensitive. In injured leaves and in leaf disks, the highest PAL activity was detected nearest the wounded tissues. Without wounding, no increase in activity was observed when leaves were cultured in sucrose and light.The optimal concentration of sucrose for enzyme activity increase ranged from 0.15 m to 0.4 m. At the suboptimal sucrose concentration, the level of PAL activity was dependent upon the concentration of sucrose. A low but constant level of activity was detected in leaf disks maintained in 0.15 m sucrose and in darkness. Light accelerated the rate of PAL increase but did not change the total level of enzyme activity which was determined by the sucrose concentration.Enzyme activity disappeared rapidly when leaf disks cultured in sucrose and light were transferred to darkness or to water in light. Unlike in Xanthium leaf disks, cycloheximide could not completely inhibit the decay of enzyme activity, suggesting that an inactivating system was synthesized during the induction period, and the activity of the inactivating system increased as the induction period lengthened.The effect of light on accumulation of PAL activity appeared to be linked to photosynthesis. In the presence of 25 mum 3-(3,4-dichlorophenyl)-1,1-dimethylurea, the effect of light on enzyme increase was completely nullified. Addition of 25 mum 3-(3,4-dichlorophenyl)-1,1-dimethylurea to culture medium caused rapid decay of PAL activity from leaf disks which had been previously cultured in sucrose and light. The relation between effect of light and photosynthesis was further demonstrated by the action spectrum. Leaf disks incubated in sucrose and light of different wavelengths exhibited maximum accumulation of PAL activity at two wavelengths (475 nm and 625 nm). Action spectrum for protection against PAL decay exhibited a plateau at 475 to 525 nm and a peak at 625 nm. Action spectra for accumulation and protection against inactivation of PAL activity, therefore, appeared to be very similar to the action spectrum of photosynthesis.
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A continuous flow of chromium into apples occurs throughout their growth. This ready movement of chromium into the apple is similar to that reported for essential trace elements such as boron, zinc, iron, copper, and manganese but differs from the restricted movement reported for mercury and cadmium.
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An improved solid agar medium (MP medium) has been developed which allows detection of pectolytic activity in bacteria. Organisms tested exhibited a variety of regulatory controls governing pectate lyase synthesis. The medium contains mineral salts, pectin, and yeast extract. After growth of the organisms, the agar plate is flooded with a polysaccharide precipitant, and pectolytic activity is shown by clear zones around active colonies. High concentrations of phosphate are shown to be necessary for pectic enzyme formation on solid media. The medium has successfully been used to detect pectolytic organisms in soil, forest litter, and rotting vegetable samples.
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Inducible synthesis of extracellular pectate lyase occurs in Erwinia carotovora, a bacterial soft-rot pathogen of plants, and, to a lesser extent, in a nonpathogenic isolate of Pseudomonas fluorescens. A combination of pectin and a heat-labile factor in fresh potato tissue or acetone powders of the tissue provided the best carbon source for induction. Yields of inducible pectate lyase were much greater than those usually reported. The pathogen, but not the saprophyte, produced a small amount of constitutive enzyme when grown on glucose. The relatively low level or absence of constitutive synthesis in these bacteria did not result from catabolite repression. Attempts were made to relieve any existing catabolite repression by restricting growth through slow feeding of glucose or by growing the organisms on glycerol. These conditions did not significantly alter the differential rate of lyase synthesis compared with changes observed in the presence of inducers. Previous growth history did not affect induction in the pathogen. However, P. fluorescens previously cultured on glucose required 10 to 20 generations of growth on inducing medium before appreciable lyase synthesis occurred. Differences between the pathogen and nonpathogen suggest that regulation of pectate lyase synthesis is related to pathogenicity of soft-rot bacteria.
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A cycloheximide-sensitive increase in the activity of phenylalanine ammonia-lyase (EC 4.3.1.5) occurs in Xanthium leaf disks exposed to light. Radioactive ammonia-lyase has been isolated by means of sucrose density gradient centrifugation and starch gel electrophoresis from disks fed l-isoleucine-U-(14)C or l-arginine-U-(14)C. The incorporation of radioactive amino acids into phenylalanine ammonia-lyase together with the inhibitory effects of cycloheximide indicate that the observed increase in enzyme activity involves the induction of lyase synthesis.The light-dependent synthesis of the ammonia-lyase is completely inhibited by 50 mum 3-(4-chlorophenyl)-1,1-dimethylurea (CMU) indicating that photosynthesis is involved. Only a trace quantity of some photosynthetic product must be needed because half light saturation occurs at very low intensity (ca. 30 ft-c). Exogenous carbohydrate is also required for continuing enzyme synthesis over a 72 hr period. But carbohydrate does not replace the photosynthetic requirement in darkness. Enzyme formed in light disappears rapidly from disks placed in the dark. The decay of ammonia-lyase activity follows first order kinetics. The half-life of the lyase ranged from 6 to 15 hr in leaf material used. Cyoloheximide inhibits the decay of lyase activity. Thus the maintenance of turnover in Xanthium leaf disks requires de novo synthesis of protein. That turnover, i.e., degradation as well as synthesis of lyase protein occurs is suggested by the apparent loss of radioactive ammonia-lyase from leaf disks placed in darkness. Light-induced synthesis coupled with rapid turnover can produce a diurnal fluctuation of ammonia-lyase activity in Xanthium leaf disks. Alternating periods of enzyme synthesis and degradation were observed in disks exposed to 24 hr cycles of light and dark. The average level of enzyme activity maintained in the tissue was directly related to the length of the light period. Induction of lyase synthesis was also observed in excised leaves and to a lesser extent in leaves of whole plants.
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The light induced synthesis of phenylalanine ammonia-lyase in disks cut from potato tubers is very sensitive to cycloheximide. Synthesis is inhibited 50% in disks cultured on 5 mum cycloheximide instead of water and almost completely in disks aged in the presence of 10 mum inhibitor. Inhibition is irreversible. Fresh disks exposed only 1 hour to 10 mum cycloheximide do not synthesize enzyme during the subsequent 24 hours.Normally a maximal enzyme activity develops in disks about 24 hours after being cut from the tuber. Thereafter enzyme activity declines. The disappearance of enzyme is not affected by concentrations of cycloheximide sufficient to inhibit the synthesis of enzyme initially. No disappearance of enzyme is noted during the initial phase of induction if enzyme synthesis is inhibited by cycloheximide. However, enzyme does disappear from the tissue if more than half the maximal enzyme content is allowed to form before synthesis is inhibited. If cycloheximide at a concentration 10-fold that needed to inhibit synthesis completely is added to disks after they have attained a maximal enzyme level, then subsequent loss of enzyme activity from the tissue is prevented. The initial stability of the enzyme in the absence of further synthesis and the inhibition of enzyme disappearance by high concentrations of cycloheximide suggest A) that early phases of induction involve synthesis of enzyme protein in the absence of turnover, B) that a system capable of degrading or inactivating the lyase subsequently forms in the tissue, and C) that the formation of the degrading or inactivating system requires protein synthesis.The effect of cycloheximide on uptake and incorporation of l-isoleucine-U-(14)C into soluble and insoluble proteins of tuber disks was also examined. During induction the rate of uptake increased 3 to 4-fold, and the rate of incorporation into protein, corrected for change in uptake, increased 25-fold. Cycloheximide inhibited incorporation of isoleucine-(14)C into proteins of fresh disks more than 80%. It did not prevent activation of general protein synthesis during induction and inhibited incorporation in induced disks only 20%. At all times incorporation of amino acid into the soluble, lyase-rich, protein fraction was more sensitive to cycloheximide than the insoluble fraction.
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