bluuebunny/crossref_metadata_2025_split huggingface dataset:
Efficacy of oral rofecoxib versus intravenous ketoprofen as an adjuvant to PCA morphine after urologic surgery
Authors: Cabrera M. C. , Schmied S. , Derderian T. , White P. F. , Vega R. , Santelices E. , Verdugo F. 
Published: September 2004

Background:  Adjunctive use of nonsteroidal anti‐inflammatory drugs has become increasingly popular in the perioperative period because of their opioid‐sparing effects. This randomized, controlled, double‐dummy study was designed to evaluate the cost‐effectiveness of using oral rofecoxib as an alternative to intravenous ketoprofen for pain management in patients undergoing urologic surgery.Methods:  Seventy patients were randomly assigned to receive either a placebo (Control) or rofecoxib 50 mg po (Rofecoxib) 1 h prior to surgery. After a standardized spinal anesthetic, patients in the Control group received ketoprofen 100 mg IV q 8 h for 24 h, while the Rofecoxib group received an equivolume of saline at 8‐h intervals for 24 h. Both groups were allowed to self‐administer morphine (1 mg IV boluses) using a PCA delivery system. The need for ‘rescue’ analgesic medication, as well as pain scores [using an 11‐point verbal rating scale (VRS) (0 = none to 10‐severe)], were recorded at 1, 2, 6, 12, and 24‐h intervals after surgery. In addition, the incidences of side‐effects were recorded at the end of the study period.Results:  Total amount of morphine required in the initial 24‐h postoperative period was nonsignificantly reduced in the Rofecoxib group (29 ± 2 vs. 37 ± 4 mg). More importantly, the percentage of patients reporting moderate‐to‐severe pain (VRS score ≥4) during the study period was lower in the Rofecoxib group (12 vs. 22%, P < 0.05). The daily cost of rofecoxib (USD 1.14 for 50‐mg dose) was also significantly less than ketoprofen (USD 3.06 for three 100‐mg doses).Conclusion:  Premedication with oral rofecoxib (50 mg) is a cost‐effective alternative to the parenteral nonselective NSAID, ketoprofen (100 mg q 8 h), when used as an adjuvant to PCA morphine for pain management after urologic surgery.
Peripheral venous pressure is an alternative to central venous pressure in paediatric surgery patients
Authors: Anter A. M. , Bondok R. S. 
Published: September 2004

Background:  Peripheral venous pressure (PVP) is easily and safely measured. In adults, PVP correlates closely with central venous pressure (CVP) during major non‐cardiac surgery. The objective of this study was to evaluate the agreement between CVP and PVP in children during major surgery and during recovery.Methods:  Fifty patients aged 3–9 years, scheduled for major elective surgery, each underwent simultaneous measurements of CVP and PVP at random points during controlled ventilation intraoperatively (six readings) and during spontaneous ventilation in the postanaesthesia care unit (three readings). In a subset of four patients, measurements were taken during periods of hypotension and subsequent fluid resuscitation (15 readings from each patient).Results:  Peripheral venous pressure was closely correlated to CVP intraoperatively, during controlled ventilation (r=0.93), with a bias of 1.92 (0.47) mmHg (95% confidence interval = 2.16–1.68). In the postanaesthesia care unit, during spontaneous ventilation, PVP correlated strongly with CVP (r = 0.89), with a bias of 2.45 (0.57) mmHg (95% confidence interval = 2.73–2.17). During periods of intraoperative hypotension and fluid resuscitation, within‐patient changes in PVP mirrored changes in CVP (r = 0.92).Conclusion:  In children undergoing major surgery, PVP showed good agreement with CVP in the perioperative period. As changes in PVP parallel, in direction, changes in CVP, PVP monitoring may offer an alternative to direct CVP measurement for perioperative estimation of volume status and guiding fluid therapy.
Impact of a critical care outreach team on critical care readmissions and mortality
Authors: Garcea G. , Thomasset S. , Mcclelland L. , Leslie A. , Berry D. P. 
Published: September 2004

Aims:  The aim of a critical care outreach team is to facilitate discharges from critical care beds, educate ward staff in the management of deteriorating patients, facilitate transfer to critical care and reduce readmission rates to critical care. Although intuitively a good idea, there are few data to support outreach in terms of reducing the readmission rate to critical care and subsequent patient mortality.This retrospective observational study attempted to determine the change in the critical care readmission rate, an indicator of the quality of critical care, critical care mortality and in‐hospital mortality following the introduction of a critical care outreach team in a major teaching hospital.Methods:  A retrospective review of 1380 discharges from critical care was undertaken and the readmissions identified (n = 176). Readmission rate, mortality and other demographic data were compared between the pre and postoutreach periods.Results:  Critical care mortality, in‐hospital mortality and 30‐day mortality were all reduced in the postoutreach period amongst readmissions to critical care. There was also a decease in the overall mortality of all patients admitted to critical care. There were no apparent causative factors for this reduction in mortality before and following outreach.Conclusions:  There are many confounding factors in assessing the impact of outreach teams in hospitals. This study tentatively concludes that outreach teams may have a favourable impact on mortality rate amongst readmissions to critical care, but more data is needed from multicentre trials.
Purification and Properties of Human Placental ATP Diphosphohydrolase
Authors: Christoforidis Savvas , Papamarcaki Thomais , Galaris Dimitrios , Kellner Roland , Tsolas Orestes 
Published: September 2004

ATP diphosphohydrolase activity (ATP‐DPH) has been previously identified in the paniculate fraction of human term placenta [Papamarcaki, T. & Tsolas, O. (1990) Mol. Cell. Biochem. 97, 1–8]. In the present study we have purified to homogeneity and characterized this activity. A 260‐fold purification has been obtained by solubilization of the particulate fraction and subsequent chromatography on DEAE Sepharose CL‐6B and 5′‐AMP Sepharose 4B. The preparation has been shown to be free of alkaline phosphatase even though the placental extract is rich in this activity.The purified enzyme is a glycoprotein and migrates as a single broad band of 82 kDa on SDS/PAGE. The same band is obtained after photoaffinity labeling of the enzyme with 8–azido‐[α‐32P]ATP. The enzyme has a broad substrate specificity, hydrolyzing triphosphonucleosides and diphosphonucleosides but not monophosphonucleosides or other phosphate esters. The activity is dependent on the addition of divalent cations Ca2+ or Mg21. The Km values for ATP and ADP were determined to be 10μM and 20 μM, respectively. Maximum activity was found at pH 7.0–7.5 with ATP as substrate, and pH 7.5–8.0 with ADP. The enzymic activity is inhibited by NaN3 NaF3 adenosine 5′‐[β,γ‐imido]triphosphate and adenosine 5′‐[α,β‐methylene]triphosphate.Protein sequence analysis showed ATP‐DPH to be N‐terminally blocked. Partial internal amino acid sequence information was obtained after chymotryptic cleavage and identified a unique sequence with no significant similarity to known proteins.ATP‐DPH activity has been reported to be implicated in the prevention of platelet aggregation, hydrolysing ADP to AMP and thus preventing blood clotting.
Purification and Biochemical Characterization of Myomesin, a Myosin‐Binding and Titin‐Binding Protein, from Bovine Skeletal Muscle
Authors: Obermann Wolfgang M. J. , Plessmann Uwe , Weber Klaus , Fürst Dieter O. 
Published: September 2004

We report a method for isolating homogeneous myomesin from mammalian skeletal muscle. The identity of the purified bovine protein was confirmed by its reactivity with myomesin‐specific monoclonal antibodies and with polyclonal antibodies raised against peptides derived from the amino‐terminal and carboxy‐terminal ends of the sequence predicted by the human myomesin cDNA. All partial sequences obtained from bovine myomesin can be aligned along the human sequence predicted by its cloned cDNA. Electron microscopy of myomesin revealed short flexible rods with a molecular length of about 50 nm. Circular dichroism spectra showed a high degree of β structure as expected for a member of the immunoglobulin superfamily of proteins. Alignment of the sequences of the class I and II domains of myomesin with the sequences of domains of known three‐dimensional structure provides a more detailed model of myomesin. In agreement with this view, the cleavage sites observed by limited proteolysis locate primarily between individual domains.In a solid‐phase overlay assay myomesin specifically bound to the myosin rod and to light meromyosin (LMM), but not to the carboxy‐terminal 30‐kDa fragment of LMM. The myosin‐binding site seemed to be confined to the amino‐terminal 240 residues of the molecule. The cross‐reactivity of myomesin with the phosphorylation‐dependent monoclonal neurofilament antibody NE14 [Shaw, G. E., Debus, E. & Weber, K. (1984) Eur. J. Cell Biol. 34, 130–136] was analyzed. NE14 reactivity of myomesin was abolished by alkaline phosphatase. Reactivity of the antibody on stable proteolytic fragments of myomesin showed that the phosphorylation site must reside within the carboxy‐terminal 60 residues.
The Affinity of Human Erythrocyte Porphobilinogen Synthase for Zn2+ and Pb2+
Authors: Simons Timothy J. B. 
Published: September 2004

Porphobilinogen synthase activity has been measured in human erythrocyte lysates supplemented with metal‐ion buffers to control free Zn2+ and Pb2+ concentrations. The enzyme is activated by Zn2+ with a Km, of 1.6 pM and inhibited by Pb2+ with a K1 of 0.07 pM. Pb2+ and Zn2+ appear to compete for a single metal‐binding site. The half‐time for loss of Zn2+ from the active site, or replacement of Pb2+ by Zn2+, were in the 10–20‐min range at 37°C. Zn2+ did not affect the affinity for the substrate 5‐aminolevulinate, but Pb2+ reduced it non‐competitively.All the experiments were conducted with a blood sample of the common 1–1 phenotype [Astrin, K. H., Bishop, D. P., Wetmur, J. G., Kaul, B., Davidow, B. & Desnick, R. J. (1987) Ann. NY Acad. Sci. 514, 23–29]
Tissue Distribution and Intracellular Localisation of the 75‐kDa Inositol Polyphosphate 5‐Phosphatase
Authors: Speed Caroline J. , Matzaris Maria , Bird Phillip I. , Mitchell Christina A. 
Published: September 2004

The 75‐kDa inositol polyphosphate 5‐phosphatase (75‐kDa 5‐phosphatase) hydrolyses several important mediators of intracellular calcium homeostasis, including inositol 1,4,5‐trisphosphate [Ins(1,4,5)P3], inositol 1,3,4,5‐tetrakisphosphate [Ins(1,3,4,5)P4] and phosphatidylinositol 4,5‐bisphosphate [PtdIns(4,5)P2].Northern analysis of various human tissues revealed the 75‐kDa 5‐phosphatase has a ubiquitous expression, where differential splicing may occur in specific tissues. Prominent expression of a 4.4‐kb transcript was noted in human lung, thymus, testes and placenta, and a 4.6‐kb transcript was observed in heart, brain, kidney, ovary and colon. Determination of the intracellular location of the enzyme by indirect immunofluorescence, demonstrated that the 75‐kDa 5‐phosphatase was associated with mitochondrial and cytosolic cellular compartments. Immunoprecipitation of the total cell homogenate of human lung carcinoma cells (A549) with anti‐(recombinant 75‐kDa 5‐phosphatase) antibodies revealed that the 75‐kDa 5‐phosphatase is the major PtdIns(4,5)P2 5‐phosphatase in this cell line. Analysis of PtdIns(4,5)P2 5‐phosphatase activity in subcellular fractions of A549 cells revealed peak 75‐kDa 5‐phosphatase enzyme activity in the cytosolic and mitochondrial enriched fractions. Immunoblot analysis further confirmed the mitochondrial location of the enzyme.This study demonstrates the tissue distribution and intracellular location of the 75‐kDa 5‐phosphatase and reveals a novel location for an enzyme involved in phosphatidylinositol turnover.
Characterization of a cDNA Encoding a Cytosolic Peptidylprolyl Cis‐Trans ‐Isomerase from Blattella Germanica
Authors: Martinez‐Gonzalez José , Hegardt Fausto G. 
Published: September 2004

Cyclophilins are an abundant and ubiquitous class of proteins first identified by their high affinity for the immunosuppressive drug cyclosporin A. Cyclophilins have peptidylprolyl cis/trans ‐isomerase activity in vitro, and thus may be involved in protein folding and trafficking in vivo.In this study, we report the cloning and characterization of a Blattella germanica cyclophilin cDNA. Analysis of this 846‐bp cDNA reveals an open reading frame coding for a polypeptide of 164 amino acid residues with a molecular mass of 17934 Da. This B. germanica cyclophilin shares a central peptidylprolyl cis/trans ‐isomerase and a cyclosporin‐A‐binding domain with other cyclophilin sequences. The B. germanica cyclophilin amino acid sequence shares 83% identity with the cytosolic cyclophilin isoform from Drosophila melanogaster (Cyp‐1). This identity suggests that B. germanica cyclophilin is a member of the cytosolic cyclophilin A (CyPA) family.From the alignment of cyclophilin sequences, we have found that 62 residues (positional identity of 40%) have remained invariant in eukaryotes for more than 1 billion years of divergence. We calculated a unit evolutionary period of 30.9 million years for the cytosolic isoform.Northern‐blot analyses show that B. germanica CyPA mRNA is abundant, and present in all insect organs tested. The highest values for B. germanica cyclophilin mRNA tissue content were found in 6‐day‐old ovary, followed by brain, testis, and gut (15–30% the content of ovary). The muscle, fat body, and colleterial gland contained the lowest cyclophilin mRNA level (1–5% the content of ovary). There is a developmental pattern of gene expression affecting the embryo stages. These results suggest that this ubiquitously expressed B. germanica cyclophilin is subject to a differential regulation in tissues and during development. Southern‐blot analysis of B. germanica DNA shows that only one copy of the CyPA gene is present per genome, whereas at least 20 genes or pseudogenes were detected in the mammalian genome.
Involvement of Phosphorylase Kinase Inhibition in the Effect of Resorcinol and Proglycosyn on Glycogen Metabolism in the Liver
Authors: Van Schaftingen Emile 
Published: September 2004

The purpose of this study was to identify the mechanism by which proglycosyn and resorcinol decrease the phosphorylase a content and the fructose 2,6‐bisphosphate concentration in isolated hepatocytes. The intracellular concentrations of the glucuronide derivatives of proglycosyn and resorcinol have been measured by HPLC in hepatocytes incubated for 5 min or 30 min with different concentrations of these agents. At both times, there was a reciprocal relationship between the phosphorylase a content and the intracellular concentration of the glucuronidated metabolites, half‐maximal inactivation being observed at about 2 μmol/g protein and 0.25 μmol/g protein for resorcinylglucuronide and proglycosyn‐glucuronide, respectively. Glycogen synthase was not significantly activated by these agents after 5 min but was well activated after 30 min.Preincubation of hepatocytes with 1 mM resorcinol or with 100 μM proglycosyn resulted in a decrease in the rate at which phosphorylase was activated following the addition of glucagon, vasopressin, the protein phosphatase inhibitor calyculin A or the calcium ionophore A 23187, but did not reduce the rate of synthase inactivation. Proglycosynglucuronide and resorcinylglucuronide inhibited phosphorylase kinase in liver Sephadex filtrates, with Ki values of about 0.75 mM and 4 mM, respectively. Preincubation of the filtrates with ATP and cAMP decreased the sensitivity of phosphorylase kinase to resorcinylglucuronide by about fourfold. It is concluded that the effect of resorcinol and proglycosyn on the phosphorylase a content is due, at least partly, to an inhibition of phosphorylase kinase by their glucuronidated metabolites.Resorcinol and proglycosyn caused a parallel decrease in the concentration of fructose 2,6–bisphosphate and of hexose 6‐phosphates, without significantly changing the activity of 6‐phosphofructo‐2‐kinase. The decrease in the fructose 2,6‐bisphosphate concentration appears therefore to be secondary to the decrease in the hexose 6‐phosphate concentration.
Reaction Mechanism of Thioredoxin: 3′‐Phospho‐adenylylsulfate Reductase Investigated by Site‐Directed Mutagenesis
Authors: Berendt Uwe , Haverkamp Thomas , Prior Antje , Schwenn Jens D. 
Published: September 2004

Properties of purified recombinant adenosine 3′‐phosphate 5′‐phosphosulfate (PAdoPS) reductase fromEscherichia coliwere investigated. The Michaelis constants for reduced thioredoxin andPAdoPSare 23 μM and 10 μM, respectively; the enzyme has aVmaxof 94–99 (μmol min−1mg−1and a molecular activity/catalytically active dimer of 95 s−1. Adenosine 3′,5′‐bisphosphate (PAdoP) inhibits competetively (Ki4 μM) with respect toPAdoPS; adenosine 2′,5′‐bisphosphate and sulfite are not inhibitory. Alkylation by SH‐group inhibitors irreversibly inactivates the enzyme.The structural gene (cysH) encodes for a small polypeptide with a single Cys residue located in a conserved cluster (KXECGI/LH) of amino acids. Involvement of the only Cys and of Tyr209 in the reduction ofPAdoPSto sulfite was investigated by site‐specific mutagenesis:cysHwas mutated by single‐strand‐overlay extension PCR; the mutated genes were cloned in pBTac1 and expressed inE. coliRL 22 (ΔcysHIJ). Homogenous Cys239Ser and Tyr209Phe mutantPAdoPSreductases were investigated for altered catalytic properties. Mutation of the single Cys reducedVmaxby a factor of 4.5×103(Vmax= 0.02–0.013 μmol min−1mg−1) with marginal effects onKmforPAdoPS(19 μM) and reduced thioredoxin (14 μM). Mutation of Tyr209 drastically affected saturation with thioredoxin (Km1.5 μM) and decreasedVmax(0.22–0.25 μmol min−1mg−1) in addition to a small increase inKmforPAdoPS(31 μM).Chromophores as prosthetic groups were absent from recombinantPAdoPSreductase. Difference absorption spectra between reduced and oxidized forms of wild‐type and mutated proteins indicated that, in addition to Cys239 and Tyr209, an unidentified Trp (Δλmax292 nm) appears to be involved in the reduction.The data suggest a special ping‐pong mechanism withPAdoPSreacting with the reduced enzyme isomer in a Theorell‐Chance type mechanism.
Effects of Ionic Strength and pH on the Binding of Medium‐Chain Fatty Acids to Human Serum Albumin
Authors: Pedersen Anders Overgaard , Mensberg Karen‐Lise Dons , Kragh‐Hansen Ulrich 
Published: September 2004

Binding equilibria for the interactions of the medium‐chain fatty acid anions, laurate and myristate, with defatted human serum albumin have been investigated under varying environmental conditions such as ionic strength and pH. Since these ligands bind strongly to albumin (Kass≈ 107 M−1), conventional equilibrium dialysis is not a feasible method for these investigations. Consequently, we employed a dialysis method, allowing determination of very low concentrations of unbound ligand by measuring the rate of exchange of labelled ligand across a dialysis membrane under conditions of chemical equilibrium.Over a range of ionic strength, 8–68 mM, the binding of the first few molecules of laurate to albumin was weakened with increasing ionic strength, whereas the binding of subsequent molecules seemed to proceed independently of ionic strength. The binding of myristate by albumin, however, appeared to be independent of ionic strength in the observed range of concentrations.The influence of pH in the range 5.1–9.0 on the binding of the two fatty acid anions by albumin was more complicated. The first molecule of laurate appeared to bind with a slightly weaker affinity to albumin at low pH, compared to pH 7 and high pH, while the trends for the following molecules varied. The binding of myristate (irrespective of concentration) seemed to strengthen monotonously with pH, but this conclusion depends critically on the interpretation of the kinetic behaviour of the myristate anion.We have previously shown [Pedersen, A. O., Honoré, B. & Brodersen, R. (1990) Eur. J. Biochem. 190, 497–502] that the strength of binding of the first few molecules of the two fatty acid anions to albumin decreases with increasing temperature, whereas binding of subsequent molecules seems to proceed independently of temperature.We explain these findings as follows. The binding of the first few (3 or 4) molecules of the C12 laurate anion is clearly driven by formation of ionic bonds between the fatty acid anion and positively charged groups, such as lysine residues, in the albumin molecule, whereas the binding of subsequent molecules of laurate seems to depend more on hydrophobic interactions. In the case of the C14 myristate anion, the binding of the first few (only 1 or 2) molecules may depend on ionic forces, but binding of the following molecules of myristate seems to depend on hydrophobic interactions only. Thus, multiple binding of both ligands is considered to be an association between fatty acid anions stacked on the protein molecule and/or it may take place in hydrophobic regions of the albumin molecule. This type of binding will contribute entropy rather than enthalpy to the total binding energy.
The Myristoylated Alanine‐Rich C‐Kinase Substrate (MARCKS) is Sequentially Phosphorylated by Conventional, Novel and Atypical Isotypes of Protein Kinase C
Authors: Herget Thomas , Oehrlein Silke A. , Pappin Dairyl J. C. , Rozengurt Enrique , Parker Peter J. 
Published: September 2004

The myristoylated alanine‐rich C‐kinase substrate (MARCKS) is the major protein kinase C (PKC) substrate in many cell types including fibroblasts and brain cells. Here we describe the phosphorylation of MARCKS and the site specificity for different PKC isotypes. Conventional (c)PKC β1, novel (n)PKC δ and nPKC ɛ efficiently phosphorylated the MARCKS protein in vitro. The Km values were extremely low, reflecting a high affinity between kinases and substrate. The apparent affinity of nPKC δ (Km= 0.06 μM) was higher than that of nPKC ɛ and cPKC β1 (Km= 0.32 μM). The rate of substrate phosphorylation was inversely correlated with affinity and decreased in the order nPKC ɛ > cPKC β1 > nPKC δ. Atypical (a)PKC ζ did not phosphorylate the intact MARCKS protein. However, a 25‐amino‐acid peptide deduced from the MARCKS phosphorylation domain, was efficiently phosphorylated by aPKC ζ as well as by the other three PKC.Site analysis revealed that only serine residues S152, S156 and S163 were phosphorylated, with S163 phosphorylated highest, followed by S156 and S152; in contrast, S160 and S167 were not phosphorylated. No further PKC phosphorylation sites could be detected in MARCKS. The phosphorylation pattern was independent of the type of PKC isotype used. Kinetic analysis showed, that MARCKS is sequentially phosphorylated in the order S156 > S163 > S152 by cPKC, nPKC and aPKC. There was no dramatic difference in the sequential phosphorylation of MARCKS detectable when comparing the four PKC isotypes. The results are discussed in the context of the functional significance of MARCKS phosphorylation.