bluuebunny/crossref_metadata_2025 huggingface dataset:
A C9 related channel forming protein in the cytoplasmic granules of human large granular lymphocytes
Authors: Zalman Leora S. , Brothers Mary A. , Müller-Eberhard Hans J.
Published: February 2005

Large granular lymphocytes (LGL) from human blood main-tained in culture for 2 to 6 weeks with IL-2 were found positive in the K562 cell killing assay. The cytoplasmic granules of the LGL were isolated, lysed and the soluble proteins were passed over a Sepharose-anti-C9 column. The retained protein was eluted with NaC1 and found to consist by SDS polyacrylamide gel electrophoresis of essentially one component with a molecular weight of approximately 70 000. The protein did not give a positive precipitation test with anti-human C9 by Ouchterlony analysis, but it reacted reproducibly with anti-human C9 by Western blot analysis. By ELISA the cross reaction with human C9 was less than 1%. The C9 related lymphocyte protein lacked C9 hemolytic activity, but it formed functional pores in liposomes in presence of Ca++. These results suggest that the cytoplasmic granules of human LGL that are capable of killing NK target cells contain C9 related protein which is involved in the cellular cytotoxicity reaction.
Gamma-glutamyltranspeptidase induction by cortisol in liver parenchyma of unweaned rats
Authors: Tongiani Roberto , Paolicchi Aldo , Chieli Elisabetta
Published: February 2005

In contrast to many differentiated hepatic functions developing after birth, very little is known about in vivo glucocorticoid influences on postnatal expression of fetal liver enzymes, such as GGT. This study showed that cortisol markedly induces liver GGT activity in unweaned rats, but has no effectafter weaning. Enzyme induction was dose- and time-dependent and occurred in parenchymal cells, progressing with time from zone 1 to zone 2 of the liver acinus. Zone-3 hepatocytes were unresponsive even after a 5-day treatment. Lag-times for GGT induction in zones I and 2 of the liver acinus were 1 to 2 days and 2 to 3 days, respectively. From this, a permissive cell change, determined by the hormone administration itself, seems required for the hepatocyte GGT induction by cortisol in pre-weaning rats.
Catalysis by protein disulphide-isomerase of the assembly of trimeric procollagen from procollagen polypeptide chains
Authors: Forster Simon J. , Freedman Robert B.
Published: February 2005

Type-I procollagen, 14C-biosynthetically labelled, was reduced under denaturing and non-denaturing conditions. Reoxidation to disulphide-linked trimers occurred with non-denatured chains in the presence of an oxidant system containing oxidized and reduced glutathione. Dimeric intermediates were not detected. This reoxidation was accelerated by homogeneous beef liver protein disulphide-isomerase.
Effect of ruthenium red on the Ca2+ and Sr2+ efflux from rat liver mitochondria: Influence of nupercaine
Authors: Pezzi Luis
Published: February 2005

The rate of ruthenium-red-induced Ca2+ efflux depends on the time that the calcium interacts with the mitochondria prior to the addition of the inhibitor. This time-dependency is abolished in the presence of nupercaine; it does not occur in the case of Sr2+ efflux from mitochondria in which the endogenous Ca2+ has been substituted by strontium (strontium-treated mitochondria, STM). Ruthenium red inhibits the respiratory-inhibitor- or uncoupler-induced Sr2+ efflux from STM, but not the Ca2+ efilux from standard mitochondria. The influence of the calcium-induced mitochondrial damage upon the effect of ruthenium red is discussed.
Exercise endurance-training alters vitamin E tissue levels and red-blood-cell hemolysis in rodents
Authors: Aikawa K. M. , Quintanilha A. T. , de Lumen B. O. , Brooks G. A. , Packer L.
Published: February 2005

Muscle tissue levels of dl-α-tocopherol (vitamin E) were significantly lower in endurance-trained rats than in sedentary animals, whether the animals were fed on vitamin-E-deficient or control (vitamin-E-sufficient) diets. In vitamin-E-deficient rats, liver tissue levels of vitamin E were significantly lower in those that were endurance-trained than in those that were sedentary; this was not the case in control animals. In addition, for vitamin-E-deficient rats, the onset of red-blood-cell hemolysis in the sedentary animals occurred one week earlier than in the endurance-trained animals. Thus, it appears that training induces a protective effect against hemolysis despite vitamin E deficiency.
Rabbit antibodies against the low molecular weight folate binding protein from human milk. Use for immunological characterization of human folate binding proteins in an enzyme-linked immunosorbent assay (ELISA)
Authors: Høier-Madsen Mimi , Hansen Steen Ingemann , Holm Jan
Published: February 2005

Antibodies to a low molecular weight folate binding protein isolated from human milk were raised in rabbits and used for development of a two-site enzyme-linked immunosorbent assay (ELISA) for immunological characterization of human folate binding proteins (FBPs). The high and low molecular weight FBPs from human milk were immunologically indistinguishable. Furthermore, the FBPs in human urine and cerebrospinal fluid showed a cross-reactivity of 70% and 30%, respectively. No cross-reactivity of the FBP from cow's milk was observed.
Ciglitazone is not itself thermogenic but increases the potential for thermogenesis in lean mice
Authors: Thurlby P. L. , Wilson S. , Arch J. R. S.
Published: February 2005

Chronic dietary administration of the oral hypoglycaemic ciglitazone (3 g/day for 14–28 days) to lean, non-diabetic CD1 mice resulted in increased brown adipose tissue mitochondrial GDP binding and a marked increase in the thermic effect of the beta-adrenoceptor agonist BRL 26830A. However, ciglitazone was not itself thermogenic after an acute dose, nor did it raise resting metabolic rate during chronic dietary dosing.
Morphological transformation and tumorigenicity in C3H/10T1/2 cells transformed with an inducible c-Ha-ras oncogene
Authors: Trimble William S. , Boulianne Gabrielle L. , Hozumi Nobumichi
Published: February 2005

A C3H/10T1/2 cell line containing an inducible metallothionein-ras hybrid oncogene was conditionally and reversibly transformed upon exposure to zinc ions. Interestingly, although the cell line was fully malignant when expressing only low levels of ras, complete morphological transformation required much higher levels.
Hepatic uptake of gluconeogenic substrates in late-pregnant and mid-lactating rats
Authors: Casado Javier , Remesar Xavier , Pastor-Anglada Marçal
Published: February 2005

Lactate uptake by liver is markedly increased in late-pregnant and mid-lactating rats without concomitant changes in its availability. Glycerol contribution to the liver 3-C unit uptake is only significant at term gestation (50% of lactate uptake) but almost negligible at mid-lactation (10% of lactate uptake). Pyruvate is only taken up by the liver of 15-day lactating rats. As a general trend, the livers of either pregnant or lactating rats are provided with an enhanced capacity to take up gluconeogenic substrates.
Analysis by partitioning in aqueous two-phase systems of the loss of transferrin-binding capacity during maturation of rat reticulocytes
Authors: Mendieta J. , Herráez A. , Sancho P. , Luque J.
Published: February 2005

A decrease in the number of binding sites for125I-transferrin, without an apparent modification of the association constant, has been observed during the maturation of reticulocytes into erythrocytes. As an experimental model, different red cell populations from phenylhydrazinic anaemic rates (95% to 12% reticulocyte-rich) have been used. The fractionation by multiple partition in two-phase systems of these red cell populations has been applied here to show the relationship between number of transferrin receptors and rate of reticulocyte maturation.
Hepatic endothelial lipase activity in neonatal rat liver
Authors: Burgaya Ferran , Peinado Julia , Llobera Miquel , Ramirez Ignasi
Published: February 2005

Hepatic endothelial lipase (HEL) activity is as high in the neonatal (1-day old) rat liver as in adults. Most of the HEL activity is located at the capillaries since 75% of the total activity is released by heparin or collagenase perfusion. The residual activity (non-releasable) is located in hepatocytes and not in hemopoietic cells, which are the major cell type in neonatal liver. Per mg of protein, the HEL activity is 50% higher in neonatal than in adult hepatocytes. We suggest that neonatal hepatocytes have an increased capacity to synthesize and secrete HEL activity, so maintaining a high activity in the whole organ. it might contribute to the hepatic uptake of cholesterol from circulating lipoproteins, in a period in which endogenous cholesterol synthesis is known to be inhibited in the liver.
L-Tyrosine stimulates induction of tyrosinase activity by MSH and reduces cooperative interactions between MSH receptors in hamster melanoma cells
Authors: Slominski Andrzej , Jastreboff Pawel , Pawelek John
Published: February 2005

L-tyrosine, a precurosr to melanin, has recently been shown to be a regulator of the melanogenic pathway in some cultured melanoma cell lines. In this paper we demonstrated that L-tyrosine, besides increasing binding capacity for MSH, decreased cooperativity between MSH receptors and increased the level of tyrosinase induction by MSH. Apparently, regulation of MSH receptor activity by L-tyrosine involves specific changes in the interactions between the receptors and modification of the cellular responsiveness to MSH.
Islet cell surface antibodies in genetically obese hyperglycaemic (ob/ob) mice
Authors: Flatt Peter R. , Bone Adrian J. , Bailey Clifford J.
Published: February 2005

A quantitative method for circulating islet cell surface antibodies (ICSA), based on the binding of125I-protein A to insulin-producing RINm5F cells, was used to evaluate ICSA in plasma of 4- to 40-week-old Aston obese hyperglycaemic (ob/ob) mice and normal control (+/+) mice. RINm5F cells bound 2502 ± l196 c.p.m.125I-protein A per 105 cells (mean ± S.D., n = 54) after incubation with +/+ plasma. ICSA positive plasma (defined as125I-protein A binding, mean ± 2 S.D. of +/+ plasma) was detected in 3 out of 54+/+ mice and 3 out of 54 ob/ob mice. ICSA were not observed in ob/ob mice before the onset of diabetes (7 weeks of age), but were detected at 9, 20 and 40 weeks. At 20 weeks125I-protein A binding produced by ob/ob plasma was 35% greater than +/+ plasma (P < 0.05). The low occurrence of ICSA in ob/ob mice (6%) suggests that factors other than ICSA are responsible for B-cell dysfunction and eventual islet degeneration observed in Aston ob/ob mice.
The calmodulin content of normal and leukaemic lymphocytes
Authors: MacNeil sheila , Dawson Rebecca , Tucker William F. G. , Clegg Alan , Platts Andrew , Rees Robert C.
Published: February 2005

Normal human peripheral blood mononuclear cells (PBMC) and leukaemic cell lines (three of human and one of gibbon origin) were found to contain similar levels of calmodulin (CaM) when expressed relative to the total cell protein. Two of the cell lines examined further were found to contain much higher amounts of CaM per cell (up to 5-fold) than PBMC but this was readily explained by their much greater cell size. Variations in CaM levels were noted during culture of both PBMC and leukaemic cells which were apparently independent of the percentage of cells undergoing active division in these cultures. These results do not support, the contention that transformed cells contain a higher proportion of CaM than normal cells.
‘Small’-proteoglycan: collagen interactions: Keratan sulphate proteoglycan associates with rabbit corneal collagen fibrils at the ‘a’ and ‘c’ bands
Authors: Scott J. E. , Haigh M.
Published: February 2005

l. Proteoglycans (PGs) in rabbit corneal stroma and mouse sclera have been stained for electron microscopy with Cupromeronic blue in a critical electrolyte concentration (CEC) mode, with and without prior digestion of the tissue by keratanase or chondroitinase ABC to remove the keratan sulphate (KS) or chondroitin-dermatan sulphates (CS or DS) respectively.2. Two classes of PGs, located orthogonally to the corneal collagen fibrils at either the ‘step’ (band ‘a’ or ‘c’) or gap zone (band ‘d’ or ‘e’) are shown to be KS-PGs or DS-PGs respectively. Four separate and specific PG binding sites on Type I collagen fibrils have thus been identified.3. Rabbit corneal KS and DS PGs each contain two kinds of PG (Gregory JD, Coster L & Damle SP (1982) J. Biol. Chem.257, 6965–6970). We propose that each ‘small’ protein-rich PG is associated with a specific binding site on the collagen fibril.
Structural and functional studies on C4b-binding protein, a regulatory component of the human complement system
Authors: Chung L. Ping , Reid Kenneth B. M.
Published: February 2005

The binding and cofactor activities of C4b-binding protein were examined before and after limited proteolysis by pepsin, trypsin and chymotrypsin. The major fragments generated were characterized by amino acid sequencing, thus establishing the precise points of limited proteolysis. These studies allow a tentative assignment of the cofactor activity site to the residues 177–322 of the 549 amino acid long chain of C4b-binding protein but indicated that residues in the region 332–395 are important in the binding activity.
The antibody paradox: Trying on a pair of genes
Authors: Fleischman Julian B.
Published: February 2005

Rodney Porter's separation of antibody molecules into Fab and Fc fragments engendered the notion that a single antibody polypeptide chain might be coded by two or more genes. This concept profoundly influenced the development of molecular immunology over the past 25 years. Our current knowledge of antibody gene organization has enabled investigators to recombine antibody genes to create ‘chimeric’ antibodies with a number of potentially useful applications.
Human conglutinin-like protein
Authors: Jensenius Jens Chr , Thiel Steffen , Baatrup Gunnar , Holmskov-Nielsen Uffe
Published: February 2005

The presence in human plasma of a molecule homologous to bovine congtutinin is indicated by the results of biological and immunochemical analysis. The human conglutinin-like protein shows calcium-dependent binding to complement-treated solid phase IgG and immunological cross-reaction with chicken anti-bovine conglutinin. The binding of the human protein to complement-treated IgG was inhibited by N-acetyl-D-glucosamine but not by other sugars. Analysis by SDS-PAGE and Western blotting showed reaction of anticonglutinin with molecules of similar mobility to the monomer and hexamer of bovine conglutinin.
Quantitative expression of epitopes defined by murine monoclonal antibodies on DR molecules from different HLA haplotypes
Authors: Letarte Michelle , Addis Jane , Iturbe Sonia , Petsche Dieter
Published: February 2005

Monoclonal antibodies 50D6 and 2Ir5, reactive with human class II molecules, were analyzed quantitatively by flow cytometry and cellular radioimmunoassay for their binding to cells of different HLA-DR types. Monoclonal antibody 50D6 bound equally to cells of all DR types tested except DR7, where no reactivity was observed. Monoclonal antibody 2Ir5 was reactive with all cells. However, the percentage of DR molecules at the cell surface expressing 2Ir5 epitope varied with the DR type and increased as follows: DR3 = DR7 < DR2 < DR5 < DR4 < DR1. MAb 50D6 reacted with an epitope spatially related to but distinct from the 2lw4 epitope present on all DR molecules. The 50D6 epitope was shown to be present on isolated DR1 molecules.
A glycophospholipid covalently attached to the C-terminus of the Thy-1 glycoprotein
Authors: Williams Alan F. , Tse Albert G.-D.
Published: February 2005

The Thy-1 glycoprotein is a very abundant cell surface molecule of rat thymocytes and neuronal cells with the properties of a molecule that inserts into the lipid bilayer. The hydrophobicity is due to a glycophospholipid component covalently attached to the carboxy group of the C-terminal cysteine residue. The mature glycoprotein does not contain a stretch of hydrophobic amino acids that could traverse the membrane bilayer. These findings present a new mode of membrane attachment for a cell surface molecule that can mediate lymphokine release and cell division after cross-linking by antibodies.
Subcortical limbic 3H-N-propylnorapomorphine binding sites are markedly modulated by cholecystokinin-8 in vitro
Authors: Agnati L. F. , Fuxe K.
Published: February 2005

By means of the dopamine (DA) agonist radio ligand 3H-N-propyinorapomorphine (3H-NPA) the effects of cholecystokinin-8 (CCK-8) have been evaluated in vitro on the binding characteristics of the DA agonist sites in membrane preparations from the subcortical limbic forebrain containing mainly nucleus accumbens and tuberculum olfactorium. It was shown that CCK-8 (10−8 M) can produce a 40% increase in the KD value of the 3H-NPA binding sites and a significant I0% increase in the Bmax values of these sites. It is therefore suggested that there exist marked receptor-receptor interactions between the CCK-8 binding sites and DA agonist binding sites in the limbic forebrain. On the basis of these findings and in view of the fact that CCK peptides are comodulators in certain types of mesolimbic DNA neurons but cannot modulate DA release in these DNA synapses) the hypothesis is introduced that the presence of DA comodulators such as CCK-8 in the DA synapses makes possible a heterostatic regulation of the synapse. Thus, by means of receptor-receptor interactions, peptide comodulators may change the set point of the main transmission line without inducing homeostatic feedback responses on synthesis and release of the main transmitter, opening up a new way to modulate chemical transmission in general.
Monoclonal antibodies against the complement control protein Factor H (β1 H)
Authors: Sim E. , Palmer M. S. , Puklavec M. , Sim R. B.
Published: February 2005

Two mouse monoclonal antibodies against the human complement control protein, Factor H (β1H), are described. The antibodies are both IgG − γ1 - subclasses and are directed against different epitopes on the human Factor H molecule. One of the antibodies, MRC OX 24, increases the cofactor activity of Factor H in Factor I-mediated cleavage of soluble C3b. The second antibody, MRC OX 23, which has no effect alone, reduces the increase in cofactor activity observed in the presence of the first antibody. However, MRC OX 24 inhibits the binding of 125I-labelled Factor H to surface-bound C3b (EAC3b). Again MRC OX 23 alone does not have any effect but decreases the inhibition in 125I-Labelled Factor H binding to EAC3b observed with MRC OX 24. These studies show clearly that the interaction of Factor H with soluble C3b is different to its interaction with surface-bound C3b. In an indirect immunoprecipitation system using these monoclonal antibodies, single-chain molecules of 150 000 mol. wt. are specifically precipitated from human serum and also from the sera of other primates - rhesus monkey, cynomolgus monkey, and African green monkey. There was no precipitation from sera of cow, pig, sheep, chick, or rabbit. Using a radioimmunoassay with radiolabelled monoclonal MRC OX 23, the concentration of Factor H in human plasma was determined.
An investigation of the growth characteristics of oil-palm (Elaeis guineensis) suspension cultures using 31P NMR
Authors: Hughes W. A. , Bociek S. M. , Barrett J. N. , Ratcliffe R. G.
Published: February 2005

High-resolution 31P nuclear-magnetic-resonance (NMR) spectra are reported for oil-palm (Elaeis guineensis) cells in suspension culture. The spectra are a signicant improvement on the results that have appeared for other cultures and they are comparable with the spectra of the meristematic tissue in seedling roots. The NMR technique was used in parallel with other analytical methods to investigate the growth characteristics of the suspension culture, indluding the effect of 2,4-dichlorophenoxyacetic acid.
Maximal activities of hexokinase, 6-phosphofructokinase, oxoglutarate dehydrogenase, and carnitine palmitoyltransferase in rat and avian muscles
Authors: Blomstrand Eva , Challiss R. A. John , Cooney Gregory J. , Newsholme Eric A.
Published: February 2005

The maximum activities of 6-phosphofructokinase and oxoglutarate dehydrogenase in muscle provide quantitative indices of the maximum capacities of anaerobic glycolysis and the Krebs cycle (i.e. the aerobic capacity) respectively. These activities were measured in red, white, and cardiac muscle of birds and the rat. The activities in the white pectoral muscle of the domestic fowl suggest that the Krebs cycle plus electron transfer could provide only about 1% of the rate of ATP production provided by anaerobic glycolysis whereas in pigeon pectoral muscle the predicted maximal rates from the two processes are similar. In contrast to domestic-fowl pectoral muscle, the white rat muscle, epitrochlearis, contains a significant activity of oxoglutarate dehydrogenase, which indicates that the Krebs cycle could provide about 12% of the maximum rate of ATP formation. This may be explained by a higher proportion of type-I and -IIA fibres in the rat muscle compared to the avian muscle. In the aerobic muscles of the rat the maximum activities of carnitine palmitoyl transferase indicate that fatty-acid oxidation could provide a high rate of ATP formation.
Specificity of a ctyochemical bioassay for arginine-vasopressin and its validation for plasma measurement
Authors: Smith A. , McIntosh N.
Published: February 2005

The total Na+/K + ATP-ase activity of the thick ascending limb of the loop of Henlé may be stimulated by arginine-vasopressin (AVP). Lysine-vasopressin (LVP), oxytocin (OT), and arginine-vasotocin (AVT) produce less than 5% of the enzyme activity induced by the same concentration of AVP. Physiological concentrations of a mixture of other hormones with known activity on the kidney (T3, T4 aldosterone, angiotensin II, and OT) did not significantly increase total Na+/K + ATP-ase activity. Specific AVP antiserum consistently removed greater than 90% of the stimulatory effect of plasma. The concentration of AVP in plasmas from dehydrated subjects was greater than l0 times that of the same subjects hydrated. Intra-assay coefficient of variation was 35% and 52% from 200 μl and 20 μl of plasma respectively. The interassay coefficient of variation was 53% and 55% from plasma pools with high and low AVP content.
The structure of the α-keratin microfibril
Authors: Fraser R. D. B. , MacRae T. P.
Published: February 2005

Quantitative measurements of the intensity of the meridional reflections in the X-ray-diffraction pattern of α-keratin are shown to be consistent with a microfibril structure in which a surface lattice with an axially projected period around 200 Å is subject to a periodic interruption with an axially projected period of 470 Å. Taken in conjunction with recent evidence on the chemical structure of α-keratin and other inter-mediate filaments this finding enables an elaboration to be made of a model proposed earlier by RDB Fraser, TP MacRae, & E Suzuki (3. Mol. Biol. 108, 435–452, 1976.) for the α-helical framework of the microfibrii. The disposition and connectivity of the helical segments suggested here provides a straightforward explanation of a number of recent physicochemical and electron-microscopical observations on intermediate filaments and provides a starting point for the development of models for the framework of other intermediate filaments.
Inhibition of Na,K-dependent adenosine triphosphatase by adenosine in intact pigeon erythrocytes
Authors: Leskovac Vladimir
Published: February 2005

In intact pigeon erythrocytes, adenosine is a potent inhibitor of Na,K-dependent adenosine triphosphatase. In purified cell-membrane preparations, adenosine is only a weak competitive inhibitor of Na,K-ATPase, with respect to ATP. This indicates that adenosine must not be a direct inhibitor of the sodium pump in intact red cells per se; instead, adenosine exerts its inhibitory effect via endogenous cell factors.
Modern theories of metabolic control and their applications
Authors: Westerhoff Hans V. , Groen Albert K. , Wanders Ronald J. A.
Published: February 2005

Existing, qualitative notions with respect to the way in which enzyme properties control metabolism are discussed in the light of the control analysis developed by H. Kacser and J. A. Burns ((1973) in: Rate Control of Biological Processes, Davies DD, ed., Cambridge University Press, pp. 63–104) and R. Heinrich and T. A. Rapoport ((1974) Eur. 3. Biochem. 42, 89–95), and recent experimental data. Points at which the existing notions should be adjusted are: (i) Metabolic control is shared by enzymes rather than confined to one rate-limiting enzyme per pathway. (if) Whether an enzyme exercises strong control on a flux cannot be deduced solely from its own properties, nor is it directly related to its distance from equilibrium. With respect to metabolic control, enzymes should be classified into four groups, rather than two (reversible versus irreversible). (iii) The distribution of control among the enzymes depends on the metabolic conditions. (iv) Control structures of metabolic pathways probably differ with the function of that pathway.
Alterations in the components of ribonucleotide reductase in hydroxyurea-resistant hamster cells
Authors: Wright Jim A. , Cory Joseph G.
Published: February 2005

Two components of mammalian ribonucleotide reductase have been separated by blue dextran-Sepharose chromatography from a hydroxyurea-resistant cell line, NcR-30A2, and its parental wild type. Analysis of reductase activity in these cells and the enzyme components reveals that there are three alterations involving ribonucleotide reductase activity in NcR-30A2 cells. There is an elevation in the effector-binding (EB) component, an elevation in the non-heine-ironcontaining (NHI) component, and an alteration in the NHI component that renders the enzyme less sensitive to inhibition by hydroxyurea. These findings easily account for the resistance of NcR-30A2 cells to the antitumor agent hydroxyurea, and to other drugs with a similar mode of action.
Mitochondria of adult human brown adipose tissue contain a 32 000-Mr uncoupling protein
Authors: Bouillaud F. , Combes-George M. , Ricquier D.
Published: February 2005

Using both immunohistological study and photoaffinity labelling with radioactive azido-ATP, evidence is presented that the mitochondrial membranes of the brown adipose tissue of the human adult contain a 32 000-Mr uncoupling protein, which is probably similar to the uncoupling protein of BAT of rodents.
Calmodulin in human serum and the specific release of calmodulin from calmodulin-rich platelets
Authors: Mac Neil Sheila , Walker Simon W. , Seid Janet , Tomlinson Stephen
Published: February 2005

Calmodulin(CaM)-like activity was detected in human serum and foetal calf serum, with an activity i0 times more than that detectable in plasma. Serum CaM was largely accounted for by release from human platelets as confirmed by both radioimmunoassay and sodium-dodecyl-sulphate/polyacrylamide-gel electrophoresis of CaM partially purified from platelet releasate obtained in response to thrombin. Lactate dehydrogenase release was unaffected by thrombin. Platelet CaM content was very variable (1.3 to 11.3 pg/mg protein; n = 15). It is suggested that intact platelets are rich in CaM and that release of CaM during preparation explains the variation in CaM content reported here and in the literature.
Sequence of ionophore conformational changes induced by a simulated membrane/water interface
Authors: Brasseur Robert , Deleers Michel , Ruysschaert Jean-Marie
Published: February 2005

We demonstrate how the progressive passage through an interface simulated by a linear increase in dielectric constant may mediate a reversible transconformation of Na+/X337A (lasalocid A) from a cyclic to an extended structure, During the passage through the interface, the complex adopts progressively a more extended conformation favorable to the Na+ complexation or decomplexation at the interface.
Species differences in the expression of carbohydrate differentiation antigens on mammalian blood cells revealed by immunofluorescence with monoclonal antibodies
Authors: Thorpe Susan J. , Feizi Ten
Published: February 2005

Following recent observations using monoclonal antibodies that carbohydrate structures behave as differentiation antigens of man and mouse, we have made a preliminary survey of the expression of 8 monoclonal antibody-defined carbohydrate antigens on blood cell smears of man, baboon, mouse, rat, rabbit, pig, and dog. There are considerable species differ-ences in the patterns of antigen expression. However, certain generalizations can be made as follows: the i and I antigens, associated with linear and branched carbohydrate chains consisting of repeating N-acetyl-lactosamine sequences (GalB1-4GIcNAc, termed Type-2 backbone sequences) are widely distributed among granulocytes and lymphocytes of all the species studied, and on erythrocytes, monocytes, and piateiets of some of them. Substantial amounts of Type-1 backbone sequences (GalB1-3GlcNAc) may occur on rabbit lymphocytes. The N-acetylneuraminic acid-containing antigens, Pr 2 and Gd, are also expressed to varying degrees on blood cells. On the other hand, antigens based on mono- and diiucosylated N-acetyllactosamine, termed SSEA-1 (or X-hapten) and C14 (or Y-hapten) are predominantly granulocyte/monocyte-associated antigens. The former antigen is expressed in overt form only on untreated human granulocytes but occurs in cryptic state, masked by sialic acid, on human monocytes, and on the granulocytes and monocytes of baboon, rabbit, and dog but not on those of mouse, rat, and pig. The latter antigen is expressed on human granulocytes and on neuraminidase-treated monocytes and granulocytes oi dog. Lymphocytes of dog are unusual in their expression of C14 antigen, in cryptic state, masked by sialic acid residues. Although the physiological roles of these various carbohydrate structures, in vivo, are not yet known, they seem excellent candidates as determinants of species and cell-type differences in susceptibilities to infective agents.
S-Adenosylhomocysteine hydrolase activity during differentiation of HL-60 cells
Authors: Chiba Peter , Plas A. , Wessels J. M. C. , De Bruyn C. H. M. M.
Published: February 2005

Early changes in S-adenosylhomocysteine (SAH) hydrolase activity during DMSO-induced granulocytic differentiation of HL-80 ceils were followed. Within 24 h a decrease of activity of SAH hydrolase could be detected in induced cultures but not in control cultures. This decrease could be shown to be associated with G1 phase of the cell cycle and was detected prior to phenotypic changes of the ceils.
A new member of the troponin C superfamily: Comparison of the primary structures of rat oncomodulin and rat parvalbumin
Authors: Mac Manus J. P. , Watson D. C. , Yaguchi M.
Published: February 2005

When the amino-acid sequence of the 108-residue, rat tumour calcium-binding protein, oncomodulin, was aligned with that of rat muscle parvalbumin, 55 homologous positions were found, with an additional 33 single base-pair substitutions. This extensive homology, with virtual identity of the calcium-binding domains, signalled oncomodulin to be a member of the troponin C superfamily. The presence of Cys-18 and Phe-66 in oncomodulin, plus its isoelectric point of 3.9) suggest that this tumour protein is a 8-parva Jbumin, rather than a muscle α-parvalbumin.
A cell-free analysis of the endocytic pathway
Authors: Davey John
Published: February 2005

The molecular control of the endocytic pathway is poorly understood. To obtain this information requires the use of cell-free systems which faithfully recreate the various endocytic events as they occur in the intact cell. Here I describe our approach to elucidating the mechanism which controls the fusion between different vesicles on the pathway.
Effects of purified myosin light chain kinase on myosin light chain phosphorylation and catecholamine secretion in digitonin-permeabilized chromaffin cells
Authors: Lee Sung A. , Holz Ronald W. , Hathaway David R.
Published: February 2005

Many non-muscle cells including chromaffin cells contain actin and myosin. The 20,000 dalton light chain subunits of myosin can be phosphorylated by a Ca2+/calmodulin-dependent enzyme, myosin light chain kinase. In tissues other than striated muscle, light chain phosphorylation is required for actin-induced myosin ATPase activity. The possibility that actin and myosin are involved in catecholamine secretion was investigated by determining whether increased phosphorylation in the presence of [γ-32P]ATP of myosin light chain by myosin light chain kinase enhances secretion from digitonin-treated chromaffin cells. In the absence of exogenous myosin light chain kinase, 1 μM Ca2+ caused a 30–40% enhancement of the phosphorylation of a 20 kDa protein. This protein was identified on 2-dimensional gels as myosin light chain by its comigration with purified myosin light chain. Purified myosin light chain kinase (400 μg/ml) in the presence of calmodulin (10 μM) caused little or no enhancement of myosin light chain phosphorylation in the absence of Ca2+ in digitonin-treated cells. In the presence of 1 μM Ca2+, myosin light chain kinase (400 μg/ml) caused an approximately two-fold increase in myosin light chain phosphorylation in digitonin-treated cells in 5 min. The phosphorylation required permeabilization of the cells by digitonin and occurred within the cells rather than in the medium. Myosin light chain kinase-induced phosphorylation of myosin light chain was maximal at 1 μM. Ca2+. Under identical conditions to those of the phosphorylation experiments, secretion was unaltered by myosin light chain kinase. The experiments indicate that the phosphorylation of myosin light chain by myosin light chain kinase is not a limiting factor in secretion in digitonin-treated chromaffin cells and suggest that the activation of myosin is not directly involved in secretion from the cells. The experiments also demonstrate the feasibility of investigation of effects of exogenously added proteins on secretion in digitonin-treated cells.
Secretory processes in lymphocyte function
Authors: Henkart Pierre , Henkart Maryanna , Hodes Richard , Taplits Michael
Published: February 2005

The secretion of immunoglobulin by plasma cells has been considered a classical example of the “non-regulated” pathway of protein secretion, in which newly synthesized protein is processed by the Golgi, packaged into small vesicles, and immediately secreted without intracellular storage. In the case of lymphokine secretion by T lymphocytes, it is generally not clear whether this non-regulated pathway is also being used, as opposed to the “regulated” pathway which has been proposed to operate in the cytotoxic lymphocyte mechanism. In this case, as in mast cells and endocrine cells, proteins are synthesized and then stored in cytoplasmic granules. The secretion is triggered (regulated) by a membrane receptor-ligand interaction, which for the cytotoxic lymphocytes is part of the target cell binding process. In cytotoxic T lymphocytes, this secretion process can be measured by following the appearance of a granule serine protease in the medium, and it has been shown to be triggered by target cells or by immobilized antibodies which bind the T celt receptor complex. In addition to cytotoxic lymphocytes, cloned T helper cells contain this serine protease in cytoplasmic granules with a low internal pH. Helper lymphocytes secrete this enzyme in response to (1) soluble antigen which has been processed by cells bearing the appropriate MHC antigens; (2) immobilized antibodies against the T cell receptor complex; (3) a combination of phorbol ester and calcium ionophore. Thus in both helper and cytotoxic lymphocytes, the regulated pathway of protein secretion clearly operates after triggering by the T cell antigen receptor.
A hypothesis for the local control of osteoclast function by Ca2+, nitric oxide and free radicals
Authors: Towhidul Alam A. S. M. , Huang Christopher L.-H. , Blake David R. , Zaidi Mone
Published: February 2005

Several important conclusions have recently emerged from in vitro studies on the resorptive cell of bone, the osteoclast. First, it has been established that osteoclast function is modulated locally, by changes in the local concentration of Ca2+ caused by hydroxyapatite dissolution. It is thought that activation by Ca2+ of a surface membrane Ca2+ receptor mediates these effects, hence providing a feedback control. Second, a number of molecules produced locally by the endothelial cell, with which the osteoclast is in intimate contact, have been found to affect bone resorption profoundly. For instance, the autocoid nitric oxide strongly inhibits bone resorption. Finally, reactive oxygen species have been found to aid bone resorption and enhance osteoclastic activity directly. Here, we will attempt to integrate these control mechanisms into a unified hypothesis for the local control of bone resorption.
ConA induced changes in energy metabolism of rat thymocytes
Authors: Buttgereit F. , Brand M. D. , Müller M.
Published: February 2005

The influence of ConA on the energy metabolism of quiescent rat thymocytes was investigated by measuring the effects of inhibitors of protein synthesis, proteolysis, RNA/DNA synthesis, Na+K+-ATPase, Ca2+-ATPase and mitochondrial ATP synthesis on respiration. Only about 50% of the coupled oxygen consumption of quiescent thymocytes could be assigned to specific processes using two different media. Under these conditions the oxygen is mainly used to drive mitochondrial proton leak and to provide ATP for protein synthesis and cation transport, whereas oxygen consumption to provide ATP for RNA/DNA synthesis and ATP-dependent proteolysis was not measurable. The mitogen ConA produced a persistent increase in oxygen consumption by about 30% within seconds. After stimulation more than 80% of respiration could be assigned to specific processes. The major oxygen consuming processes of ConA-stimulated thymocytes are mitochondrial proton leak, protein synthesis and Na+K+-ATPase with about 20% each of total oxygen consumption, while Ca2+-ATPase and RNA/DNA synthesis contribute about 10% each. Quiescent thymocytes resemble resting hepatocytes in that most of the oxygen consumption remains unexplained. In constrast, the pattern of energy metabolism in stimulated thymocytes is similar to that described for Ehrlich Ascites tumour cells and splenocytes, which may also be in an activated state. Most of the oxygen consumption is accounted for, so the unexplained process(es) in unstimulated cells shut(s) off on stimulation.
Lysis of erythrocytes by Trichomonas vaginalis
Authors: Potamianos Spyros , Mason Peter R. , Read John S. , Chikunguwo Silas
Published: February 2005

The in vitro hemolytic activity of 4 isolates of Trichomonas vaginalis was investigated. Repetitive hemolysis assays of any one isolate showed cyclical fluctuations in hemolytic activity, varying over 24 hr of continuous culture. Maximal hemolytic activity was detected using trichomonads in the lag phase of the growth cycle. Investigations showed that hemolysis was a contact-dependent phenomenon and microscopic investigation of samples showed a significant correlation between hemolysis and attachment of erythrocytes to the trichomonad surface. Quantitative data from cytoadherence assays using [51Cr]-labeled erythrocytes were consistent with these observations. It is suggested that hemolytic activity is dependent upon adherence of red blood cells to the surface of T. vaginalis.
Carboxyterminal telopeptide of type I collagen, ICTP, as a marker of matrix degradation in neonatal mouse calvarial bones, in vitro
Authors: Ljunggren Östen , Ljunghall Sverker
Published: February 2005

Bone resorption, in vitro, is often measured as the release of prelabelled45Ca from neonatal mouse calvarial bones, or from fetal rat long bones. In this report we describe a technique to measure the breakdown of bone-matrix, in vitro. We also describe a new way to dissect neonatal mouse calvarial bones, in order to obtain large amounts of bone samples.

Twelve bone fragments were dissected out from each mouse calvaria and were thereafter cultured in CMRL 1066 culture medium in serum-free conditions in 0.5 cm2 multiwell culture dishes. Matrix degradation after treatment with parathyroid hormone was assessed by measuring the amount of carboxyterminal telopeptide of type I collagen (ICTP) by RIA. The data on matrix degradation was compared to the release of prelabelled45Ca from neonatal mouse calvarial bones. We found that the dose-responses for parathyroid hormone-induced release of prelabelled45Ca and ICTP were identical.

In conclusion: RIA-analysis of the ICTP-release is an easy and accurate method to measure degradation of bone-matrix, in vitro. Furthermore, the new dissection technique, described in this report, makes it easy to obtain large amounts of bone samples and thus to perform extensive experiments, e.g. dose-responses for agents that enhance bone resorption.
Inhibitors such as staurosporine, H-7 or polymyxin B cannot be used in skeletal muscle to prove the role of protein kinase C on insulin action
Authors: Gumà Anna , Muñoz Purificación , Camps Marta , Testar Xavier , Palacín Manuel , Zorzano Antonio
Published: February 2005

The precise role of protein kinase C in insulin action in skeletal muscle is not well defined. Based on the fact that inhibitors of protein kinase C block some insulin effects, it has been concluded that some of the biological actions of insulin are mediated via protein kinase C. In this study, we present evidence that inhibitors of protein kinase C such as staurosporine, H-7 or polymyxin B cannot be used to ascertain the role of protein kinase C in skeletal muscle. This is based on the following experimental evidences: a) staurosporine, H-7 and polymyxin B markedly block in muscle the effect of insulin on System A transport activity; however, this effect of insulin is not mimicked in muscle by TPA-induced stimulation of protein kinase C, b) H-7 and polymyxin B block insulin action on System A transport activity in an additive manner to the inhibitory effect of phorbol esters, c) staurosporine, H-7 and polymyxin B block the effect of insulin on lactate production, a process that is activated by insulin and TPA in an additive fashion, and d) staurosporine completely blocks the tyrosine kinase activity of insulin receptors partially purified from rat skeletal muscle.
A sensitive spectorophotometric assay for pyruvate dehydrogenase and oxoglutarate dehydrogenase complexes
Authors: Gohil K. , Jones D. A.
Published: February 2005

The activities of pyruvate dehydrogenase and oxo-glutarate dehydrogenase can be reliably measured by coupling the production of NADH to the reduction of added cytochrome c. Maximum activities required the addition of NADH-cytochrome c reductase activity prepared from rat heart mitochondria. Compared to other spectrophotometric assays this method provides an eight-fold increase in sensitivity and is particularly suitable for use with small tissue samples such as needle-biopsy samples of human skeletal muscle. Measurements of activities in rat tissues showed them to be in the order skeletal muscle < liver < heart ≤ brown adipose tissue. Activities in normal human skeletal muscle were similar to those of rat muscle, tn the rat tissues specific differences were seen in the relative activities of the two complexes and cytochrome c oxidase suggesting tissue-specific differences in the activities of the dehydrogenases and components of the electron-transport chain.
Lipogenesis in response to an oral glucose load in fed and starved rats
Authors: Sugden Mary C. , Watts David L. , Marshall Christopher E.
Published: February 2005

Lipogenesis in livers of fed but not of starved rats is increased after intragastric feeding with glucose. In contrast, lipogenesis in brown adipose tissue increases in both fed and starved animals. These observations suggest that lipogenesis in brown adipose tissue is regulated by mechanisms in addition to, or other than, those operating in liver. The fate of newly synthesized lipid in brown adipose tissue is not known. However, the formation of palmitoyl-carnitine from palmitoyl-CoA and carnitine by mitochondria from brown fat was inhibited by malonyl-CoA. Although inhibition was not 100%, it is implied that mitochondrial uptake of the newly synthesized fat by the carnitine acyltransferase system is restricted under conditions of increased lipogenesis.
Structure-function relationships in insulin
Authors: Saunders D. J.
Published: February 2005

A new interpretation of structure-function relationships in the insulin molecule is presented. Negative cooperativity is postulated to arise from a dimerization event occurring between two receptor-bound molecules. The receptor-binding surface of insulin can necessarily not involve residues involved in dimerization as has been generally accepted. Support for this interpretation is based on published data.
Malate dehydrogenase: Isolation from E. coli and comparison with the eukaryotic mitochondrial and cytoplasmic forms
Authors: Fernley Ross T. , Lentz Steven R. , Aradshaw Ralph A.
Published: February 2005

Escherichia coli malate dehydrogenase has been isolated in homogeneous form by a procedure employing chromatography on DEAE-cellulose, 5′-AMP-Sepharose, and Sephacryl-200. It is composed of two identical polypeptide chains each of Mr = 32 500. Like porcine mitochondrial malate dehydrogenase, it is devoid of tryptophan, but otherwise it is not particularly more similar in composition to one of the eukaryotic isozymes than to the other. However, amino-terminal sequence analysis of the first 36 residues shows remarkable similarity of the bacterial and mitochondrial enzymes (69% identical residues) in contrast to the cytoplasmic form (27%). The two porcine heart enzymes are identical in 24t% of the positions compared. These results clearly establish that all three forms of malate dehydrogenase have evolved from a common precursor and that the prokaryotic and mitochondrial forms have retained sequences that are much closer to the ancestral one than the cytoplasmic enzyme. These findings appear to further substantiate the endosymbiotic hypothesis for the origin of the mitochondrion.
Nuclear magnetic resonance Fourier transform spectroscopy
Authors: Ernst Richard R.
Published: February 2005

The development and current status of Fourier transform spectroscopy is described.
Effects of parathyroid hormone on cyclic AMP-formation and cytoplasmic free Ca2+ in the osteosarcoma cell line UMR 106-01
Authors: Ljunggren Östen , Johansson Hans , Lerner Ulf H. , Lindh Erik , Ljunghall Sverker
Published: February 2005

The effects of parathyroid hormone (PTH) on cytoplasmic free CA2+ (Cai2+) and cAMP-formation were investigated in the rat osteosarcoma cell line UMR 106-01.

In fura-2 loaded adherent single cells bPTH 1–34 (10 nM−1μM) induced a rapid transient increase in Cai2+ in 11% of the studied cells. In fura-2 tracings from UMR 106-01 cells in suspension, bPTH 1–34 (0.1 μM) induced a transient increase in Cai2+ in 20% of the experiments. The transient increase in Cai2+ seen in suspensions of cells was not abolished by addition of EGTA (2.5 mM) prior to challenge with PTH, suggesting that the increase in Cai2+ was derived from intracellular stores.

A marked rapid increase in cAMP-formation was observed in all experiments with cells in suspension, also in the experiments where PTH did not affect Cai2+.

These data show that PTH causes a release of Ca2+ from intracellular stores in a small percentage of osteosarcoma UMR 106-01 cells, and that PTH is capable of inducing an increase in cAMP-formation without affecting Cai2+ in osteoblasts.