bluuebunny/crossref_metadata_2025 huggingface dataset:
Effect of 17 β-estradiol on calcium response to phytohaemagglutinin in human lymphocytes
Authors: Bellini Tiziana , Degani Diana , Matteuzzi Maurizio , Dallocchio Franco
Published: February 2005

Pre-treatment of human lymphocytes with 17 β-estradiol diminishes the increase in concentration of cytosolic free calcium after stimulation with phytohaemagglutinin. The effect is dependent on 17 β-estradiol concentration and on the preincubation time. The effect is not due to an interaction between 17 β-estradiol and phytohaemagglutinin, but appears to be a consequence of the binding of the hormone to the cell surface. The effect is specific for 17 β-estradiol, since the α isomer and other steroid hormones (progesterone, testosterone, diethylstilbestrol and 5α-androstan), have no effect. Since the effect of the 17 β-estradiol can be suppressed by treatment of lymphocytes with ouabain, it appears that the effect of estradiol on the rise of cytosolic calcium induced by phytohaemagglutinin is mediated by the (Na, K)-ATPase.
Glycogenolytic effect of pancreastatin in the rat
Authors: Sanchez Victor , Calvo Juan R. , Goberna Raimundo
Published: February 2005

Pancreastatin is a novel 49-amino acid peptide with a C-terminal glycine amide. The peptide was isolated from porcine pancreatic extracts and shows a structural similarity to chromogranin A. The effect of synthetic porcine pancreastatin on blood glucose levels and hepatic glycogen content was investigated in rats in vivo. Pancreastatin (300 pmol/kg) produced a time-dependent decrease in glycogen content of liver and a slight hyperglycemia. Basal plasma insulin and glucagon levels were not modified by pancreastatin. We suggest that pancreastatin could play a biological role in the glucose metabolism through a glycogenolytic effect.
Effect of morphine and acetylcholine on contractile activity and cyclic AMP in guinea-pig ileum
Authors: Alguacil L. F. , Lopez-Ruiz M. P. , Prieto J. C. , Alamo C. , Cuenca E.
Published: February 2005

Neither acute nor prolonged exposure to morphine altered cAMP content or spontaneous movements of longitudinal muscle-myenteric plexus strips of the guinea-pig ileum. By contrast, exogenous acetylcholine or electrical stimulation of the strips elicited both a decrease of cAMP concentration and a twitch response. Atropine blocked the effects of stimulation on these parameters. Addition of morphine to electrically stimulated strips inhibited the twitch response but did not affect cAMP levels. Incubation with morphine led to the development of tolerance to the inhibitory effect on twitch activity and prevented the fall in cAMP normally elicited by electrical stimulation. These results suggest that muscarinic activation is associated with a reduction of cAMP content, an effect which would be impaired in opiate-tolerant tissues.
Bradykinin stimulates prostaglandin E2 formation in isolated human osteoblast-like cells
Authors: Ljunggren Östen , Rosenquist Jan , Ransjö Maria , Lerner Ulf H.
Published: February 2005

The effect of bradykinin on prostaglandin E2 formation in cells from human trabecular bone has been studied. The cells responded to parathyroid hormone with enhanced cyclic AMP formation and were growing as cuboidal-shaped, osteoblast-like cells. In these isolated human osteoblast-like cells, bradykinin (1 μmol/l) caused a rapid (5 min) stimulation of prostaglandin E2 formation. This finding indicates that human osteoblasts are equipped with receptors for bradykinin linked to an increase in prostaglandin formation.
Cytoplasmic Ca2+ concentration of single normal human and bovine parathyroid cells measured by dual wavelength microfluorometry
Authors: Johansson H. , Larsson R. , Nygren P. , Lindh E. , Rastad J. , åkerström G. , Gylfe E.
Published: February 2005

Dual wavelength microfluorometry was utilized to measure the cytoplasmic calcium concentration (Cai2+) of single parathyroid cells loaded with the indicator fura-2. The method enabled the first registrations of Cai2+ of normal human parathyroid cells, available only in minute numbers. At 0.5 mM extracellular Ca2+, the Cai2+ levels were similar in normal human and bovine cells. Both cell types responded with an initial Cai2+ transient followed by a sustained increase when raising extracellular Ca2+ to 3.0 mM. The sustained effect exhibited a sigmoidal relation to extracellular Ca2+ in the 0.5–3.0 mM range. Although the increase was somewhat greater in the human cells, the half maximal responses were obtained at almost identical extracellular Ca2+ concentrations. Whereas K+ depolarization decreased Cai2+, the Cai2+ channel blocker D-600 had dual actions, raising Cai2+at 0.5 mM Ca2+ and decreasing it at 3.0 mM Cai2+, and the effects were similar in the bovine and human cells. The present experimental approach verified the validity of utilizing bovine cells as controls in studies of human parathyroid tissue and it appears suitable for analysis of the role of different subpopulations of parathyroid cells in the abnormal parathyroid tissue of patients with hyperparathyroidism.
A solid phase assay for galactosyl transferases. Evidence for an active site of less than 14 kDa
Authors: Plancke Yves , Delpouve Béatrice , Montreuil Jean
Published: February 2005

Galactosyltransferase (GalTase) prepared from human milk was found to exist as a complex with e-lactalbumin as demonstrated by crossed immunoelectrophoresis against specific antibodies raised against the complex. GalTase activity was stable to proteolysis and, when subjected to gel filtration on Ultrogel AcA54, the enzyme activity eluted as a single peak. A second peak of activity was found to be adsorbed to the column matrix and was eluted with buffer containing 1 M NaC1. The hydrophobic fraction represented 5% of the total GalTase activity in human milk. After polyacrylamide gel electrophoresis the main enzyme activity peak was represented by polypeptides of 67kDa molecular weight and of 14kDa molecular weight. Electroblotting of these peptides onto a nitrocellulose membrane followed by determination of GalTase activity showed activity for 45–55 kDa and for 14 kDa peptides. The hydrophobic fraction from the AcA54 column was resolved into polypeptides of 110 kDa-45 kDa molecular weight, all of which contained GalTase activity after blotting. It is supposed that the GalTase from non-proteolyzed milk is composed of a 14 kDa polypeptide containing the active site together with another part of the polypeptide backbone which is involved in the regulation of GalTase activity by α-lactalbumin, a third part of the polypeptide is responsible for the membrane insertion.
Anti-phospholipase C antibodies inhibit the lectin-induced proliferation of human lymphocytes
Authors: Moraru I. I. , Manciulea Mioara , Cǎlugǎru Anca , Ghyka Gr. , Popescu L. M.
Published: February 2005

A novel approach was used to assess the role of phosphoinositide hydrolysis in the mitogenic action of phytohemagglutinin (PHA) or concanavalin A (ConA). The treatment of human peripheral blood leukocytes (PBL) with monospecific antibodies against phospholipase C (PLC) produced a dose-dependent inhibition (up to 100%) of PHA (10 μg/ml) or ConA (25 μg/ml) proliferative effects. Thus, the activation of membrane-bound PLC is a sine-qua-non condition for lectin-induced proliferation of T lymphocytes. The key-role of PLC versus protein kinase C (PKC) is stressed by the fact that the inhibition of PKC with Hidaka's compound H-7 (40 μM) produced only a partial blockade (about 25%) of lectin mitogenic effect.
Bilayer stabilizing peptides and the inhibition of viral infection: antimeasles activity of carbobenzoxy-Ser-Leu-amide
Authors: Epand Richard M. , Lobl Thomas J. , Renis H. E.
Published: February 2005

A number of carbobenzoxy-dipeptide-amides raise the bilayer to hexagonal phase transition temperature of dielaidoylphosphatidylethanolamine (stabilizes the bilayer). The potency of the peptides in stabilizing the bilayer phase is Z-Tyr-Leu-NH2>Z-Gly-Phe-NH2>Z-Ser-Leu-NH2>Z-Gly-Leu-NH2>Z-Gly-Gly-NH2. A linear correlation was found between the respective HPLC retention time parameter k′ for the peptide and the slope of the bilayer stabilization curve determined with model membranes by differential scanning calorimetry. One dipeptide, Z-Ser-Leu-NH2, reduces measles virus cytopathic effect (CPE) in Vero cells. The mechanism by which this peptide reduces the CPE is not known, although some peptides which raise the bilayer to hexagonal phase transition temperature of phospholipids inhibit membrane fusion.
Defective regulation of the cytosolic Ca2+ activity in parathyroid cells from patients with hyperparathyroidism
Authors: Larsson Rolf , Wallfelt Chris , Abrahamsson Håkan , Gylfe Erik , Ljunghall Sverker , Rastad Jonas , Rorsman Patrik , Wide Leif , Åkerström Göran
Published: February 2005

The parathyroid hormone (PTH) release and cytosolic Ca2+ activity were determined in normal bovine parathyroid cells and parathyroid cells obtained from patients with hyperparathyroidism (HPT). There was a sigmoid relation between the cytosolic Ca2+ activity and the extracellular calcium concentration between 0.5 and 6.0 mmol/l. The PTH release was inhibited in parallel with the rise in the cytosolic Ca2+ activity. Both the hormone release and the cytosolic Ca2+ activity were lower in cells from human adenomas and hyperplastic glands∼ and in comparison with the bovine preparations these ceils had higher set points for the cytosolic Ca2+ activity and PTH release. There was a close correlation between the individual set points for the cytosolic Ca2+ activity and PTH release in a material containing both normal and pathological cells. The results indicate that the abnormal PTH release characteristic of HPT is due to a defective regulation of the cytosolic Ca2+ activity.
Nucleotide sequence of the thioredoxin gene from Escherichia coli
Authors: Höög Jan-Olov , von Bahr-Lindström Hedvig , Josephson Staffan , Wallace Betty J. , Kushner Sidney R. , Jörnvall Hans , Holmgren Arne
Published: February 2005

The nucleotide sequence of the thioredoxin gene from Escherichia coli was determined. The structural gene was identified on a cloned 3-kb PvuII Iragment by hybridization with a synthetic oligodeoxyribonucleotide corresponding to a part of the amino acid sequence of thioredoxin. Restriction-enzyme fragments were used as templates in the dideoxy sequence method, directly and after subcloning into M13mp8. A segment of 450 nucleotides was determined using both strands7 alternatively, without extensive overlaps. The sequence contains the thioredoxin coding region, a potential ribosome-binding site, and a putative promotor region. The predicted amino acid sequence differs by two inversions from the previously given thioredoxin sequence. The revised sequence is presented and the results further show that thioredoxins from E. coli B and K12 are identical.
An encapsidated, subgenomic messenger RNA encodes the coat protein of carnation mottle virus
Authors: Harbison Sally-Ann , Wilson T. Michael A. , Davies Jeffrey W.
Published: February 2005

The translation strategy of carnation mottle virus (CarMV) in vitro has been generally assumed to involve internal initiation events on full-length, genomic RNA (4.3 kb). We suggest that this is, at least in part, incorrect. Encapsidated RNA, fractionated on denaturing sucrose gradients, or total RNA from CarMV-infected leaves, fractionated under non-denaturing conditions, was translated in an mRNA-dependent rabbit reticulocyte cell-free system. Evidence for subgenomic RNAs which encode a polypeptide of Mr 38 000 was found. This product was shown to be related to authentic CarMV coat protein by partial proteolysis with α-chymotrypsin and SDS/polyacrylamide-gel electrophoresis.
Expression of a Drosophila heat-shock gene in cells of the yeast Saccharomyces cerevisiae
Authors: Nicholson Richard C. , Moran Larry A.
Published: February 2005

A 3.52-kilobase (kb) segment of Drosophila melanogaster DNA carrying the 2.l5-kb transcribed sequence for the 70 000-dalton heat-shock protein hsp70) and l.l4-kb of the 5′ flanking sequence was inserted into an autonomously replicating chimeric plasmid and used to transform the yeast Saccharomyces cerevisiae. The Drosophila gene is efficiently transcribed in the transformed cells, yielding a transcript which is 21 nucleotides shorter than the normal Drosophila mRNA at the 5′ end. Significant increases in the amount of Drosophila-specific RNA occur when the transformed ceils are subjected to heat shock, indicating that the Drosophila gene is inducible in the yeast cells.
A tetraantennary glycopeptide from human Tamm-Horsfall glycoprotein inhibits agglutination of desialylated erythrocytes induced by leucoagglutinin
Authors: Serafini-Cessi Franca , Malagolini Nadia , Dall'olio Fabio
Published: February 2005

Complex-type glycopeptides from Human Tamm-Horsfall glycoprotein were fractionated by affinity chromatography on leucoagglutinin-agarose. An oligosaccharide species was retained by the lectin-gel, suggesting that it contains an α-mannose residue of the trimannosyl core substituted at C-2 and C-6 positions with β-N-acetylglucosamine, as in tetraantennary oligosaccharides. The carbohydrate composition supported this branching pattern. The agglutination of neuraminidase-treated human erythrocytes induced by leucoagglutinin was selectively inhibited by the tetraantennary glycopeptide fraction.
Changes in kidney transamidinase activity during development in male and female rats
Authors: Tormanen Calvin D. , Sutter Barbara E.
Published: February 2005

The developmental changes in the activity of kidney transamidinase in male and female rats were investigated. The activity in both sexes increased rapidly after birth, reaching adult levels at 4 days of age. After weaning, the activity in male rats remained constant, while in female rats it declined to 60% ol that in males. Thus, transamidinase is in the neonatal cluster of enzyme differentiation.
The effect of different hyperglucagonemic states on monooxygenase activities and isozymic pattern of cytochrome P-450 in mouse
Authors: Rouer E. , Beaune P. , Augereau C. , Leroux J. P.
Published: February 2005

The continuous infusion of a low dose of glucagon (35 μg/kg/d, for 5 d) constitutes, in view of glucose-6-phosphatase and phosphoenolpyruvate carboxykinase activities, a reliable experimental model of hyperglucagonemia. By conjunction of monooxygenase assays and immunoquantitation of specific isozymes of cytochrome P-450, the actual inducing ability of glucagon has been shown and it might explain some of the modifications of the drug metabolizing system in diabetic mice. The isozymic pattern of cytochrome P-430 of liver microsomes from diabetic mice appears very different from that produced by classical inducers.
Separation of human endothelial cells from fibroblasts by centrifugation in Percoll gradients
Authors: Sbarbati Rosella
Published: February 2005

Human endothelial cells from the umbilical vein and skin fibroblasts can be separated by means of centrifugation in a density gradient of Percoll. Cells show a good recovery in culture. Viability is not impaired.
Inhibition by ceruloplasmin of the cardiac sarcolemmal adrenochrome formation
Authors: Guarnieri Carlo , Ventura Carlo
Published: February 2005

The addition of ceruloplasmin to bovine cardiac sarcolemmal vesicles supplemented with NADPH was able to reduce the formation of adrenochrome from adrenaline. This inhibitory effect appears at 2.5 μM ceruloplasmin and it is almost complete at the level of 20 μM.
Cell-free translation of mitochondrial nicotinamide nucleotide transhydrogenase
Authors: Carlenor Elisabeth , Joste Vigg , Nelson B. Dean , Rydström Jan
Published: February 2005

Mammalian nicotinamide nucleotide transhydrogenase is translated as a 5000 daltons larger molecular weight precursor in a cell-free system programmed with rat liver polysomes. The mature rat liver enzyme had the same molecular weight as the purified beef heart enzyme, 115 000 daltons. The precursor was not processed in vitro by liver mitochondria or by a rat liver mitochondrial matrix fraction, nor did it appear to bind to mitochondria. In contrast, pre-FeS protein of the cytochrome bc1 complex was processed in the same samples by both mitochondria and matrix, suggesting an important difference in the processing mechanisms or in the efficiency of processing of the two precursors.
Rat liver L-threonine deaminase: Properties and purification
Authors: Leoncini R. , Pagani R. , Casella A. , Marinello E.
Published: February 2005

A new method of purification of rat liver L-threonine deaminase has been developed, and the results obtained are compared with values obtained by other authors. Some properties of this enzyme (pH optimum, temperature optimum, thermal stability, specificity, etc.) have been examined and we found that the enzyme is inhibited by carbonate ions, that L-cysteine (a competitive inhibitor) is also an inactivator of the enzyme and that it is bound to the enzyme in a ratio of 0.25 mole of cysteine per mole of enzyme, supporting the hypothesis that the enzyme consists of 4 subunits.
Biosynthesis of trehalose by Brevibacterium flavum: Use of long range13C-13C coupling data to characterize triose phosphate isomerase activity
Authors: London Robert E. , Walker Thomas E.
Published: February 2005

The13C isotopic labeling pattern in the disaccharide trehalose (1,1-α-α-D-glucose) produced by the micro-organismBrevibacterium flavum when grown on a medium containing [1-{au13}C]glucose has been determined. Long range coupling between C-1 and C-6 carbons of the glucose units can be observed in the excreted material. It is proposed that some of the13C isotopomers in the excreted trehalose reflect the labeling pattern in (unobserved) fructose 1,6-diphosphate. Analysis of the label distribution within the framework of a steady state kinetic model allows an analysis of the contributions of the hexose monophosphate shunt and the degree of equilibration of triose phosphate isomerase. Analogous measurements on excreted glucose could be carried out in other organisms.
The differentiation of HL-60 cells in the synthetic medium induced by GM3 ganglioside
Authors: Senoo Haruki , Momoi Takashi
Published: February 2005

GM3 ganglioside, added exogenously to a promyelocytic leukemia cell line (HL-60 cells) in serum-free synthetic medium, induced differentiation into macrophage-like cells. Macrophagic morphology and function of differentiation-induced cells were determined by cell growth behavior, May-GriJnwald-Giemsa staining, activities of nonspecific esterase, phagocytosis and nitroblue tetrazolium (NBT) reduction. GM3 ganglioside may play a role in triggering differentiation of HL-60 cells into macrophage-like cells.
Erythrocyte membrane Ca2+-pumping ATPase of hypertensive humans: Reduced stimulation by calmodulin
Authors: Olorunsogo O. O. , Okudolo B. E. , Lawal S. O. A. , Falase A. O.
Published: February 2005

The Ca2+-pumping ATPase of erythrocyte plasma membranes of hypertensive humans (HTN) show, in the absence of calmodulin, a low Vmax comparable to that of the enzyme of the erythrocyte membranes of normotensive humans (NTN). Although the addition of calmodulin (1.5 μgper ml) increased the maximum activity of the calcium pump of membranes of HTN and NTN individuals by at least 2-fold and 4-fold, respectively, the activator protein partially purifed from the erythrocytes of HTN individuals enhanced the activity of the enzyme in a fashion similar to that of the protein obtained from the haemolysate of NTN individuals. A determination of the dependence of the activity of the pump on concentration of ATP revealed that the Km (ATP) of the enzyme of membranes of HTN individuals is 52% higher than that of the enzyme of membranes of NTN individuals, while the Vmax (1.75 ± 0.28 μmol ATP mg protein-1 h-1) of the pump is 46% lower in the membranes of HTN humans than that of the enzyme of membranes of normal individuals (3.25 ± 0.42 μmol ATP mg protein-1 h-1). It seems likely from these results that elevated erythrocyte Ca2+ concentration associated with essential hypertension may be due to a defective interaction between the Ca2+-pumping ATPase and the calmodulin Ca2+ complex,
Cold exposure or chronic noradrenaline treatment induces an increase in the calmodulin-like immunoreactivity of brown adipose tissue of rats
Authors: Mory G.
Published: February 2005

Cell proliferation is often associated with an increase in calmodulin, the ubiquitous intracellular calcium receptor of non-muscle cells. A long lasting increase in the proliferative activity of brown adipose tissue is induced by cold exposure in the rat. The present work showed that this phenomenon is also associated with a rapid and long lasting increase in the calmodulin content of this tissue. It was equally shown that this increase can be reproduced by noradrenaline admini-stration.
The role of transmembrane sodium gradient in rabbit ileal smooth muscle: Utilization of harmaline
Authors: Suleiman M. S.
Published: February 2005

Decreasing extracellular sodium concentration was found to produce a contractile response of rabbit ileal smooth muscle. As the concentration decreases, the amplitude of contraction increases, thus producing a dose-dependent curve. Harmaline, a competitor for sodium, was found to inhibit the sodium gradient-dependent contractions in a dose-dependent manner. The results are interpreted as harmaline inhibiting a Na–Ca exchange mechanism present in ileal smooth muscle.
Novel application of quantitative immunoassays for screening seed globulins of cowpea varieties
Authors: Mawal M. R. , Ranade S. A. , Mawal Y. R. , Ranadive S. N. , Bhattacharya A. , Ranjekar P. K.
Published: February 2005

Using antibodies raised in rabbits, radioimmunoassay (RIA) and enzyme-linked immunosorbent assay (ELISA) are standardized for cowpea (var. Pusa Barsati) seed globulins. The RIA, when used to screen three stages of seed development, reveals that maximum globulins are detected at 28 days after flowering. When three different varieties of cowpea are assayed for their globulin content by RIA and ELISA, it is observed that the Bold Grain cowpea has the highest amount of related globulin as compared to two other varieties, namely Pusa Phalgun and Asparagus Bean.
A high-affinity folate binding protein in normal human leukocytes: Ligand binding characteristics, ionic charge and molecular size
Authors: Holm Jan , Hansen Steen Ingemann , Lyngbye J⊘rgen
Published: February 2005

High-affinity binding of [3H]folate to supernatant from homogenized human leukocytes containing large amounts of binding protein displayed apparent positive cooperativity. The DEAE-Sepharose® CL-6B chromatographic profile of the supernatant at pH 6.3 contained a major peak of folate binding (Mr approx. 25 000) in the front effluent and a smaller more acidic peak (Mr approx. 25 000) that emerged after a rise in NaCl from 30 mmol/l to 1 mol/l. Triton X-100 solubilized ceil sediment from the leukocyte homogenate contained some high-affinity folate binding activity (Mr approx 25 000), typically 5–10% of the total binding activity.
Effects of fatty acid chain length and saturation on gastric inhibitory polypeptide release in obese hyperglycaemic (ob/ob) mice
Authors: Kwasowski Piotr , Flatt Peter R. , Bailey Clifford J. , Marks Vincent
Published: February 2005

Plasma gastric inhibitory polypeptide (GIP) responses to equimolar intragastrically administered emulsions of fatty acids (2.62 mmol/7.5 ml/kg) were examined in 18 h fasted obese hyperglycaemic (ob/ob) mice. Propionic acid (C3:0), a saturated short-chain fatty acid, and capric acid (C10:0), a saturated medium chain fatty acid, did not signilicantly stimulate GIP release. However, the saturated long-chain fatty acid stearic acid (C18:0), and especially the unsaturated long-chain fatty acids oleic (C18:1), linoleic (C18:2) and linolenic (C18:3) acids produced a marked GIP response. The results show that chain length and to a lesser extent the degree of saturation are important determinants of fatty acid-stimulated GIP release. The GIP-release action of long-chain, but not short-chain, fatty acids may be related to differences in their intracellular handling.
Isolation by restriction endonuclease digestion and base-specific affinity chromatography of rat-embryo DNA sequences disproportionately enriched in virogenic bromodeoxyuridine
Authors: Schwartz Stephen A.
Published: February 2005

Control and bromodeoxyuridine-containing rat-embryo-cell DNA were digested by the restriction endonucleases Hpa II and Msp I and were subsequently analyzed by agarose-gel electrophoresis as well as DNA-affinity chromatography. By the former technique, it appeared that no substantial differences existed between the two DNA samples with respect to the amount or distribution of methylcytosine. On the other hand, it was obvious following base-specific DNA chromatography that the virogenic analog was markedly concentrated in particular nucleotide sequences which demonstrated a proportionately greater affinity for the (A-T)-specific adsorbent irrespective of digestion by either restriction endonudease.
Influence of pH and ionic strength on the lysis ofMicrococcus luteuscells by hen lysozyme at low (20°C) and high (physiological, 40°C) temperature
Authors: Saint-Blancard J. , Maurel J. P. , Constant J. F. , Berthou J. , Jolles P.
Published: February 2005

From isoactivity curves (showing activity as a function of pH and ionic strength) it was found that in the pH domain 6.7–8.6 frequently used in experiments involving hen lysozyme, the pH optimum of lysis of Micrococcus luteus ceils at low ionic strength (0.02–0.05) by the high-temperature form (40°C physiological temperature) was one to two pH units lower than that by the low-temperature form (20°C).
Uptake by rat peritoneal macrophages of125I-labelled polyvinylpyrrolidone entrapped within liposomes
Authors: Pratten Margaret K. , Millard Peter C. , Lloyd John B.
Published: February 2005

Rat peritoneal macrophages in vitro capture125I-labelled polyvinylpyrrolidone entrapped within either negatively or positively charged liposomes more rapidly than they do the free macromolecule. The uptake of negatively charged liposomes was linear with time over l0 h, whilst the uptake of positively charged ones, although more rapid, was more transient. Neither type of liposome was taken up in the presence of 2,4-dinitrophenol (100 μg/ml), and 5 mM calcium chloride increased the uptake of negatively charged liposomes. The enhanced uptake of 125I-labelled polyvinylpyrrolidone when presented in liposomes must have been a consequence of entrapment rather than of a simple interaction between lipid and polyvinylpyrrolidone, since the presence of the lipids employed or of empty liposomes had no effect on the uptake of unentrapped125I-labelled polyvinylpyrrolidone.
Improved technique for investigation of cell metabolism by31P NMR spectroscopy
Authors: Jacobson Lev , Cohen Jack S.
Published: February 2005

SIp NMR studies on microorganisms have been carried out with the cells embedded in agarose gel. The novel use of the gel for the NMR studies has advantages over the usual liquid suspensions in terms of improved reproducibility of data and cell viability, with no net loss of spectral quality. Polyphosphate formation in Escherichia coli was monitored continuously for up to 24 h and metabolic changes in yeast for 6 h. Changes of the intracellular pH during glycolysis in yeast were determined from the chemical shift of the internal Pi. NMR titration curves of Pi in the presence of Mg2+ indicate uncertainties in internal pH values estimated by this technique.
‘Enteroglucagon’: A specific effect on gastric glands isolated from the rat fundus. Evidence for an ‘oxyntomodulin’ action
Authors: Bataille D. , Gespach C. , Coudray A. M. , Rosselin G.
Published: February 2005

A newly isolated porcine glucagon-like biologically active intestinal peptide (‘enteroglucagon’) has been tested for its ability to stimulate the cyclic AMP system present in different tissue preparations from the rat: membranes prepared from liver or from isolated fundic glands as well as intact glands isolated from either the fundus or the antrum. Enteroglucagon (EG) was about l0 times less potent than pancreatic glucagon (G) in liver membranes, whereas it was about 20 times more potent than G in fundic membranes as well as in fundic glands, where it acts at doses as low as 3 × 10−10 M. EG and G were practically ineffective in antral glands. It is concluded that EG is an ‘under-glucagon’ in rat liver, whereas it is a ‘super-glucagon’ in rat fundic glands. Accordingly, we propose to call this peptide 'oxyntomodulin'.
A putative pathway of glyconeogenesis in skeletal muscle
Authors: Odedra Bhanu R. , Palmer T. Norman
Published: February 2005

Evidence is presented in support of a pathway in skeletal muscle of glyconeogenesis (glycogen biosynthesis de novo) from L-glutamate and related amino acids involving the enzyme phosphoenolpyruvate carboxykinase (PEP CK). In the rat hemidiaphragm in vitro, not only did L-[U-14C]glutamate exert a glycogen-sparing action, but14C-label was incorporated into glycogen. The incorporation is thought not to be simply via label randomization and was decreased by factors that increased glycolysis or pyruvate oxidation. 3-Mercaptopicolinate and amino-oxyacetate, specific inhibitors of PEP CK and aminotransferase-type enzymes, respectively, decreased14C-incorporation from L-[U-14C]glutamate into glycogen. No quantitative determination of apparent glyconeogenic flux was made, and it remains to be established whether glyconeogenesis via PEP CK and/or via PEP CK coupled with 'malic' enzyme (or pyruvate carboxylase) is functionally important in skeletal muscle.
Some aspects of calcium regulation in cell biology
Authors: Cavaggioni Andrea
Published: February 2005

The roles of intracellular calcium in the regulation of cell metabolism and cell membrane permeability are highlighted with examples taken from recent studies.
Recent advances in biological imaging
Authors: Saibil Helen , White Nicholas
Published: February 2005

Recent innovations in microscopy and digital image processing have greatly enhanced the power of biological imaging. Basic principles of several new methods in light and electron microscopy will be discussed, and examples presented of their application to cell and molecular biology.
A transient voltage-dependent outward current in cultured cerebellar granules
Authors: Robello Mauro , Carignani Corrado , Marchetti Carla
Published: February 2005

Granule cells were dissociated from rat cerebella with a procedure that yields a 98% pure cell population. Potassium currents in these cells were studied using the patch-clamp technique. Depolarizing pulses of 10 mV step and 100 ms duration from a holding potential of −80 mV elicited two different potassium outward currents: a transient, low-voltage activated component and a long lasting, high-voltage activated component. At +30 mV, the total current reached an amplitude of 2 nA (mean value of 15 experiments). The reversal potential of the transient current, estimated by measuring tail currents, was −77 mV, close to that predicted by the Nernst equation. The transient current was half inactivated with a holding potential of −78 mV and completely inactivated with −50 mV or more positive holding potentials. Finally, the current decay could be fitted by the sum of two exponentials with time constants of about 20 and 250 ms.
Doppler velocimetry measurements of phototactic response in flagellated algae
Authors: Cantatore G. , Ascoli C. , Colombetti G. , Frediani C.
Published: February 2005

A periodic shading mechanism (about 0.5 s period) is believed to be used by flagellated algae to detect light direction. Time changes in the orientation of a population of Haematococcus pluvialis under positive photoaxis conditions (cell samples were stimulated alternatively by two diametrically opposed beams of actinic light), have been investigated by analysing the Doppler shifts of laser light scattered by the cells by means of heterodyne detection techniques. This technique allows us to measure the mean value of the component of the cells' swimming velocities along the light stimulus axis. Preliminary data indicate that the time taken by a cell population to change orientation is about 1 s.
Motor responses of Halobacterium halobrium to sinusodial light stimuli
Authors: Lucia S. , Ascoli C. , Petracchi D. , Vanni L.
Published: February 2005

Sinusoidal light stimuli have been used to elicit photoresponses in Halobacterium halobium at various periods of the stimulus. Cycle histograms and histograms of the times between consecutive reversals are used to illustrate the responses, and the method used to obtain sinusoidally modulated intense light (with periods up to 1 s) is described.
Model membrane partition ESR study in the presence of α-tocopherol by a new spin probe
Authors: Severcan F. , Cannistraro S.
Published: February 2005

The effect of α-tocopherol (αT) on partitioning and fluidity changes occurring in phospholipid liposomes have been investigated by monitoring the X-band ESR spectrum of the high resolution amphiphilic spin probe perdeutero-di-t-butyl nitroxide (PDDTBN), which partitions in the lipid and water phase of liposomes, showing all the three resonances from each phase well resolved.
Ca2+ interaction with phospholipid bilayers studied by multifrequency phase fluorometry
Authors: Zolese G. , Curatola G.
Published: February 2005

Calcium interaction with phospholipid membranes containing phosphatidic acid is studied by multifrequency phase fluorometry, using DPH as fluorescent molecule. DPH decay is analysed by a continuous distribution of lifetimes. The results suggest an increase of membrane heterogeneity at low calcium concentrations, without changes in the polarity of the environment surrounding the probe.
Subcellular distribution of the enzymes of the malate-aspartate shuttle in rat heart and effect of experimental cardiac hypertrophy
Authors: Mavrides Charalampos , Korecky Borivoj
Published: February 2005

The effect of experimental cardiac hypertrophy on the enzymes of the malate – aspartate shuttle aspartate aminotransferase (AAT) and malate dehydrogenase (MDH) was studied. (1) Aortic constriction in adult rats resulted in 25% cardiac hypertrophy in 2½–3 weeks. Total DNA (mg per heart) did not change. (2) The proportions of mitochondrial and cytosolic isozymes of AAT and MDH did not change as a result of cardiac hypertrophy. About two-thirds of each enzyme occurred in the mitochondrial form and one-third in the cytosolic form. (3) Total AAT in hypertrophic hearts, in enzyme units per mg DNA, increased by 24% compared to AAT content in the hearts of sham-operated animals. Total MDH did not change. SoIubilized protein increased by 20%. Normal hearts contained 10 times more enzyme units of MDH than of AAT. (4) Cardiac growth stimulation induced in newborn rats did not result in specific changes of either enzyme. It is suggested that true cardiac hypertrophy acts as a specific stimulus for the possibly rate-limiting enzyme AAT of the shuttle.
Rapid large-scale purification of plasmid DNA by medium or low pressure gel filtration. Application: construction of thermoamplifiable expression vectors
Authors: Vo-Quang Tuyen , Malpiece Yves , Buffard Dominique , Kaminski P. Alexandre , Vidal Dominique , Strosberg A. Donny
Published: February 2005

This paper describes a new method of lasmid DNA purification which is fast and reliable enough for most purposes in recombinant DNA technology. The present method does not require the use of toxic chemicals such as phenol or ethidium bromide, costly ultra-centrifugation procedures or other processes which can modify the supercoiled structure of the plasmids, such as adsorption on glass fiber. This method is based on the principle of gel filtration chromatography, at low pressure (1 bar) or medium pressure (between 5 and 10 bars), using Sephacryl S1000 or Superose 6B. It permits recovery oI plasmids: (I) in preparative quantities (from 300 gg to 4 mg), (II) exempt from RNA, DNA and protein contamination, and (III) suitable for various common genetic engineering procedures immediately after purification. To test the reliability of the technique as well as the degree of purilication, the plasmids were used to construct thermoampliIiable vectors, carrying the tacUV5 promoter and the 5′ end of the β -gallactosidase gone with a single EcoRl site in each of the three possible translational phases. This set of vectors is designed for the expression of foreign genes as hybrid proteins in Escherichia coli.
Polypeptide compositions of amoebae of the cellular slime mould Dictyostelium discoideum separated by partitioning during development
Authors: Sharpe P. T. , Sharrard R. M. , Watts D. J.
Published: February 2005

Amoebae of the cellular slime mould Dictyostelium discoideum at 8 h or 10 h development were separated into two populations by countercurrent distribution in a dextran-poly(ethylene glycol), two-phase system. Two-dimensional, polyacrylamide-gel electrophoresis was then used to separate the polypeptides from the populations of amoebae. The two populations of amoebae at 8 h development differed sn polypeptide composition as did the populations separated at 10 h development. This confirms that cell differentiation is initated in D. discoideum prior to 8 h development.
Reconstitution of the complement channel into lipid vesicles and planar bilayers starting from the fluid phase complex
Authors: Menestrina Gianfranco , Pasquali Flavia
Published: February 2005

Proteolysis of the fluid phase complement complex SC5b-9 transforms it into an arnphiphilic molecule which resembles the membrane attack complex of complement and reconstitutes into lipid vesicles. Complement-containing vesicles prepared in this way can be made to fuse with planar lipid bilayers transferring their protein content to the host membrane. Massive conductance increases can thus be observed, which are due to the insertion of a large number of ionic channels into the membrane. Using low concentrations of vesicles, single channels can be studied.
Molecular cloning of potato spindle tuber viroid (PSTV) cDNA synthesized by enzymatic elongation of PSTV-specific DNA primers: A general strategy for viroid cloning
Authors: Tabler Martin , Schnölzer Martina , Sänger Heinz L.
Published: February 2005

Different cDNAs were synthesized by primer extension from the RNA of the severe strain KF 440 of potato spindle tuber viroid (PSTV) with the aid of reverse transcriptase using three PSTV-specific DNA molecules as primers. The cDNAs were made double-stranded and cloned into plasmid pBR 322. Various overlapping subgenomic DNA fragments were prepared from these clones and recombined in two different ways. In both cases a PSTV DNA copy was obtained which represented the entire PSTV RNA genome. The sequence of the DNA of one of the resulting full-length clones was identical with the original PSTV isolate, whereas the other clone showed one nucleotide change. On the basis of these results the advantages and problems of different strategies for the molecular cloning of the circular viroid RNA genome are discussed.
Thermogenic and metabolic consequences of thyroid hormone treatment in brown and white adipose tissue
Authors: Williams Christine M. , Ellis Rodney
Published: February 2005

Male rats were treated with triiodothyronine in the drinking water for 12 days. In vitro rates of isoprenaline stimulated lipolysis were significantly greater in brown but not white adipose tissue. Rates of [14C]glucose incorporation into triacylglycerols were significantly reduced in BAT (brown adipose tissue) and WAT (white adipose tissue) under basal and isoprenaline stimulated conditions, in a second experiment, hyperthyroid animals showed impaired weight gain, despite increased food intake during t9 days' treatment. Energy expenditure on days 5 and 12, and BAT core temperature differences (TBAT – TCORE) on day 19, were significantly greater than in control animals. Epididymal white fat pad weight was reduced and interscapular brown fat pad weight increased by triiodothyronine treatment.
Interaction of vasoactive intestinal peptide with rat small intestinal epithelial cells after intestinal resection
Authors: Diaz-Juarez José L. , Bodega Guillermo , Arilla Eduardo , Prieto Juan C.
Published: February 2005

Specific binding of vasoactive intestinal peptide (VIP) and VIP-stimulated cyclic AMP accumulation were studied in small intestinal epithelial cells (both of crypt and villous levels) 3, 7 and 14 d after a 60% resection of the small intestine. The affinity, but not the binding capacity, of VIP receptors decreased during the adaptive hyperplastic response. Basal cyclic AMP levels were similar in cells of both control and resected rats. Resection induced a decrease of potency, but not of efficiency, of VIP on the stimulation of cyclic AMP accumulation.
Accelerated degradation of glycogen phosphorylase in denervated and dystrophic mouse skeletal muscle
Authors: Butler P. Elaine , Cookson E. Jane , Beynon Robert J.
Published: February 2005

Pyridoxal phosphate, the cofactor of glycogen phosphorylase, fulfils the criteria needed of a turnover label for this enzyme. The decay of protein-bound label following administration of [3H]pyridoxine is a good index of the rate of degradation of the enzyme in vivo. This method has been applied to the study of catabolism of the enzyme in normal, denervated and dystrophic mouse skeletal muscle. In both of the pathological conditions the enzyme is degraded more rapidly than normal.
Transglutaminase and cellular motile events: Retardation of proinsulin conversion by glycine methylester
Authors: Alarcon C. , Valverde I. , Malaisse W. J.
Published: February 2005

Glycine methylester, an inhibitor of transglutaminase, decreased glucose-stimulated insulin release and delayed proinsulin conversion in rat pancreatic islets pulselabelle with L-[4-[3H]phenylalanine. Sarcosine methylester, which does not inhibit transglutaminase activity, failed to affect insulin release and proinsulin conversion. The incorporation of L-[4-3H]phenylalanine into islet peptides, the ratio of hormonal to total tritiated peptides and the insulin content of the islets failed to be affected by either of these methylesters. It is proposed that transglutaminase participates in the control of motile events involved in both the transfer of proinsulin from its site of synthesis to its site of conversion, and the translocation of insulin from its site of storage to its site of release.
Evidence that fasting can induce a selective loss of uncoupling protein from brown adipose tissue mitochondria of mice
Authors: Trayhurn P. , Jennings G.
Published: February 2005

The effects of fasting and refeeding on the concentration of uncoupling protein in brown adipose tissue mitochondria have been investigated in mice. Fasting mice for 48 h led to a large decrease in the total cytochrome oxidase activity of the interscapular brown fat pad. Mitochondrial GDP binding and the specific mitochondrial concentration of uncoupling protein also fell on fasting. After 24 h refeeding both GDP binding and the mitochondrial concentration of uncoupling protein were normalized, but there was no alteration in the total tissue cytochrome oxidase activity. Fasting appears to induce a selective loss of uncoupling protein from brown adipose tissue mitochondria, which is rapidly reversible on refeeding.
Conserved elements in the 3′ untranslated regions of c-fos and actin mRNAs
Authors: Renan Michael J.
Published: February 2005

In this study, the nucleotide sequences of the 3′ untranslated regions (UTR) of the mouse and human c-fos genes, and the rat and human β-actin genes were examined. It is shown (i) that the 3′ UTR of c-fos is highly conserved between mouse and man, (ii) that multiple copies of a 12 bp element occur, in clusters, in the 3′ UTR both of c-fos and of β-actin. This conserved 12 bp element is analogous to the putative repressor binding site previously identified (Renan, Bioscience Reports,5 (1985), 739–753). These findings provide additional support for the proposal that regulatory signals are located in the 3′ UTR's of certain genes.
A water-soluble aminated β1-3D-glucan derivative causes regression of solid tumors in mice
Authors: Seljelid Rolf
Published: February 2005

Meth A sarcoma, when inoculated in the skin, grew progressively in hybrid CB6 Fl(Balb/c×C57B1/6) mice. When water-soluble aminated β1-3D-glucan (AG) was injected intravenously or intraperitoneally on day 7 of tumor growth, the tumors underwent complete regression. When the injection was performed on day 3 there was regression of tumors in only about half of the cases. When the injection was performed on day 14 there was no apparent effect on tumor growth. Tumors in thymectonized animals did not appear to respond to treatment with AG on day 7. The relatively simple chemistry and low toxicity of AG, together with its solubility in biological fluids, makes it a promising tool in experimental—and possibly clinical—tumor therapy.
Enzymatic cleavage of RNA by RNA
Authors: Altman Sidney
Published: February 2005

The discovery and characterization of the catalytic RNA subunit of the enzyme ribonuclease P of Escherichia coli is described.
Direct oxidation of lymphocytes by chromic acid does not induce blastogenesis
Authors: Malinin George I. , Hornicek Francis J. , Malinin Theodore I.
Published: February 2005

Blastogenic and cytotoxic effects of hexavalent chromium were evaluated by direct, 2 and 20 min oxidation of lymphocytes by 10.0 to 0.0005 mM CrO3 at 0°C. Oxidized cells exhibited concentration-dependent cytotoxicity and the inhibition of tritiated thymidine incorporation rates. When lymphocytes were oxidized first by 1.0 mM periodic acid (H5IO6) and thereafter by 1.0 mM CrO3, the viability and [3H]-TdR incorporation rates of sequentially oxidized cells were identical to the corresponding indicators of lymphocytes oxidized only by CrO3. The reversal of the oxidation sequence restored [3H]-TdR incorporation to control levels and increased cell survival. It is therefore concluded that direct interaction of hexavalent CrO3 with plasma membrane of lymphocytes results in concentration-dependent cytotoxicity and the inhibition of [3H]-TdR incorporation, but it does not induce blastogenesis.
Receptor expression and oxidase activity in human neutrophils: Regulation by granulocyte-macrophage colony-stimulating factor and dependence upon protein biosynthesis
Authors: Edwards Steven W. , Watson Fiona , MacLeod Ronald , Davies John
Published: February 2005

Incubation of human bloodstream neutrophils with 50 u/ml recombinant granulocyte-macrophage colony-stimulating factor (rGM-CSF) “primed” the respiratory burst (as assessed by fMet-Leu-Phe stimulated luminol-dependent chemiluminescence) and resulted in a rapid (within 15 min) upregulation of expression of CD11b and CD18 (as measured by FACS analysis). This rapid “priming” and modulation of receptor expression was not inhibited by cycloheximide and hence appeared to be independent of de novo protein biosynthesis. When neutrophils were incubated for up to 5 h in culture, the fluorescence distributions of CD11b and CD18 declined indicating the loss of expression of these receptors as the neutrophils aged, but in rGM-CSF treated suspensions receptor expression was maintained. When neutrophils were incubated in the presence of cycloheximide, they progressively lost their ability to generate reactive oxidants in response to fMet-Leu-Phe so that by 5 h incubation with this inhibitor they could only generate about 25% of the oxidative response stimulated in untreated cells, and the expression of CD16 and CD18 was grossly impaired. Similar effects were observed in rGM-CSF treated suspensions except that cycloheximide required longer incubation times (typically 4–5 h) before impairment of function or receptor expression occurred. These data show that de novo protein biosynthesis is required for both the maintenance of neutrophil function and also for the continued expression of some plasma membrane receptors.