bluuebunny/crossref_metadata_2025_split huggingface dataset:
Electrophoretic analysis of four high molecular weight sialoglycoproteins produced by metastatic human colon carcinoma cells
Authors: Irimura T. , Carlson D. A. , Ota D. M. 
Published: December 2004

AbstractWe have found that polyacrylamide gel electrophoresis in 3% gels in the presence of sodium dodecyl sulfate is suitable for the separation of cellular glycoproteins having molecular weights ranging from 200,000 to 1,000,000. The gels secured on a rigid support (Gelbond) allow blotting techniques with lectins and antibodies for the detection of glycoproteins. Using these methods we have separated lysates of HT‐29 human colon carcinoma cells and detected at Jeast four distinct high molecular weight Sialoglycoproteins having molecular weights of 900,000, 740.000, 560,000, and 450,000. The expression of the 9000,000 component, as revealed by wheat germ agglutinin binding, was much higher in a subline of HT‐29 cells established from liver metastases in a nude mouse than it was in the parental cells. The relative intensity of wheat germ agglutinin binding to these four sialoglycoprotein components differs depending upon their growth phase in vitro. These glycoproteins were also detectable by the binding of peanut agglutinin, provided the glycoproteins were previously treated in the gels with mild acid to remove the sialic acid from their carbohydrate chains, suggesting that mucin‐type carbohydrate chains are present on these glycoproteins. The same set of glycoproteins can be detected by metabolic labeling of the cells with [3H] glucosamine in tissue culture. Very similar glycoprotein profiles are revealed by metabolic labeling of fresh colon carcinoma tissues with [3H] glucosamine in vitro.
Characterization of mono‐, di‐, and tri‐O‐acetylated sialic acids on human cells
Authors: Hutchins Jeff T. , Reading Christopher L. 
Published: December 2004

AbstractThe presence of mono‐, di‐, and tri‐O‐acetylated sialic acids on human cells was demonstrated by using radiochromatographic and chemical techniques. Human melanoma cells and fresh colon tissue were biosynthetically labeled with 6‐ (3H) glucosamine. Radiolabeled sialic acids were hydrolytically removed from cellular glycoconjugates, purified by ion‐exchange chromatography, and separated by paper chromatography on the basis of the number of O‐substitutions on each sialic molecule, This analytical technique characterized radiolabeled sialic acids that migrated with the same Rf as synthetic mono‐, di‐, and tri‐O‐acetylated 14C‐labeled sialic acids. The mono‐O‐acetylated sialic acids were characterized by their sensitivity to sodium periodate oxidation and a crude mouse liver esterase preparation. The di‐ and tri‐O‐acetylated sialic acids were characterized by their resistance to sodium periodate oxidation and sensitivity to the action of crude mouse liver esterase. Chromatographically separated di‐ and tri‐O‐acetylated sialic acids from normal human colon tissue were characterized by their respective ion molecular weights by using fast‐atom bombardment‐mass spectrometry. Using these methods, we chemically characterized mono, di‐, and tri‐O‐acetylated sialic acids expressed on human cells. Aberrant expression of O‐acetylated sialic acids was associated with adenocarcinoma of the colon, leading to a nearly complete loss of di‐ and tri‐O‐acetylated sialic acids.
Major O‐glycosylated sialoglycoproteins of human hematopoietic cells: Differentiation antigens with poorly understood functions
Authors: Gahmberg Carl G. , Autero Matti , Hermonen Jorma 
Published: December 2004

AbstractAll human hematopoietic cells seem to contain a major, heavily O‐glycosylated sialoglycoprotein. Glycophorin A is specific for the erythroid lineage of cells, and leukocytes have a major sialoglycoprotein, also called leukosialin or sialophorin. Cell differentiation results in patterns of O‐glycosylation in these proteins, which reflect the stage of differentiation within a cell lineage as well as lineage specificity. The altered carbohydrate compositions may influence the interactions of the cells with external ligands. Healthy individuals lacking glycophorin A in their red cells are known, whereas a deficiency of the leukocyte sialoglycoprotein may result in immunological disease. Although little is known about the physiological functions of these proteins, they form interesting models for studies on regulation of glyco‐sylation, biosynthesis of O‐glycosylated glyoproteins, and function of cell surface receptors.
Endogenous lectins as mediators of tumor cell adhesion
Authors: Lotan Reuben , Raz Avraham 
Published: December 2004

AbstractEndogenous carbohydrate‐binding proteins have been found in various normal tissues and cells. Although lectins with different sugar‐binding specificities have been described, the most prevalent ones are those that bind β‐galactosides. The ability of some normal and malignant cells to bind exogenous carbohydrate‐containing ligands suggested that lectinlike activity is associated with the cell surface and that carbohydrate‐binding proteins might mediate intercellular recognition and adhesion. We found that extracts of various cultured murine and human tumor cells exhibit a galactoside‐inhibitable hemagglutinating activity. This activity was associated with two proteins of molecular weights of 34,000 and 14,500 daltons, which were purified by affinity chromatography by using immobilized asialofetuin. That these lectins are present on the cell surface was indicated by the binding of monoclonal antilectin antibodies to the surface of various tumor cells and by the immunoprecipitation of 125I‐labeled lectins from solubilized cell‐surface iodinated cells by polyclonal anti‐lectin antibodies. That these cell surface lectins are functional was demonstrated by the ability of the galactose‐terminating asialofetuin to enhance cell aggregation and of asialofetuin glycopeptides to block this homotypic aggregation as well as to suppress cell attachment to substratum, and by the inhibition of both asialofetuin‐induced cell aggregation and cell attachment to substratum by the binding of monoclonal antilectin antibodies to the cell surface. These findings implicate cell surface lectins as mediators of cell‐cell and cell‐substratum adhesion. Some of these cellular interactions might be important determinants of tumor cell growth and metastasis.
(2′‐5′)An‐dependent endoribonuclease: Enzyme levels are regulated by IFNβ, IFNγ, and cell culture conditions
Authors: Floyd‐Smith Georgia 
Published: December 2004

AbstractThe levels of a (2′‐5′) An‐dependent endonuclease (RNase L) were determined in extracts prepared from murine L cells and Ehrlich ascites tumor (EAT) cells by measuring specific binding of protein to a labeled derivative of (2′‐5′)An, (2′‐5′)A3[32P]pCp. RNase L levels were found to depend both on interferon (IFN) treatment and on cell growth conditions. Treatment of murine L cells and EAT cells with 100–2,000 IRU IFNβ or IFNγ resulted in a similar 2–4‐fold increase in the levels of RNase L when cells were present at low density. The levels of RNase L were also shown to increase 2–3‐fold as cells approached saturation density. Serum‐starved cells also displayed relatively high levels of RNase L. RNase L levels in cells maintained at high cell density did not change appreciably following treatment with IFNβ or IFNγ. Regulation of RNase L levels by cell growth conditions as well as by IFNβ or IFNγ treatment suggests that RNase L may play an important role in regulating the levels of cellular mRNAs as well as acting to degrade viral RNAs.
Cation‐independent mannose 6‐phosphate receptor contains covalently bound fatty acid
Authors: Westcott Keith R. , Rome Leonard H. 
Published: December 2004

AbstractThe cation‐independent mannose 6‐phosphate receptor (215,000 daltons) was isolated from embryonic bovine tracheal cells and embryonic human skin fibroblasts labelled with [3H]palmitic acid, the tritium label was detected in the protein upon fluorographic analysis of SDS‐polyacrylamide gels of the purified receptor. The label was not sensitive to hydroxylamine, methanolic KOH, or β‐mercaptoethanol, but labelled fatty acid was recovered from the protein by acidic methanolysis. Labelled receptor protein could not be isolated from cells grown in the presence of [3H]myristic acid. The results suggest the presence of amide‐linked palmitic acid in the structure of the cation‐independent mannose 6‐phosphate receptor.
Cytochemical, histological, and phylogenetic distribution of a 38,000‐dalton protein associated with transverse tubules
Authors: Tidball James G. , Gadus Michelle V. 
Published: December 2004

AbstractA major protein in detergent extracts of skeletal muscle appears at 38,000 daltons in electrophoretic separations. Previous investigations have provided indirect evidence that a 38‐kD skeletal muscle protein is membrane associated, and this inference has served as the basis for speculations on 38‐kD protein function. In the present study, affinity purified, polyclonal antisera against 38‐kD protein from skeletal muscle are produced for immunolocalization and biochemical assays. Immunoblots of two dimensional electrophoretic separations show that this protein is heterogenously charged at pI ∼6.4. This 38‐kD protein is not extracted from muscle in low ionic strength or high ionic strength buffers, in isotonic buffers from pH 4 to pH 8 or in buffers containing 5 mM EGTA. The 38‐kD protein is extracted, however, by isotonic, pH 7.0 buffer containing 1.0% Triton‐X. Light microscope observations using indirect immunofluorescence of anti‐38‐kD labeled tissue show the protein distributed in a reticular pattern within cross‐sectioned muscle but not at the cell surface. Longitudinal sections show the protein concentrated in periodic, transverse bands. Purified fractions of muscle plasma membrane analyzed by immunoblotting contain 38‐kD protein. Immunoblots using anti‐38 kD show that this protein is present in all vertebrate skeletal muscle examined, however, the protein is present only in cardiac muscle that contains transverse tubules. The antibody does not recognize aldolase, another 38‐kD striated muscle protein.
Characterization of a bombesin receptor on Swiss mouse 3T3 cells by affinity cross‐linking
Authors: Sinnett‐Smith James , Zachary Ian , Rozengurt Enrique 
Published: December 2004

AbstractWe have previously identified by chemical cross‐linking a cell surface protein in Swiss 3T3 cells of apparent Mr 75,000–85,000, which may represent a major component of the receptor for peptides of the bombesin family in these cells [1]. Because bombesin‐like peptides may interact with other cell surface molecules, it was important to establish the correlation between receptor binding and functions of this complex and further characterize the Mr 75,000–85,000 cross‐linked protein. Detailed time courses carried out at different temperatures demonstrated that the Mr 75,000–85,000 affinity‐labelled band was the earliest cross‐linked complex detected in Swiss 3T3 cells incubated with 125I‐labelled gastrin‐releasing peptide (125I‐GRP). Furthermore, the ability of various nonradioactive bombesin agonists and antagonists to block the formation of the Mr 75,000–85,000 cross‐linked complex correlated extremely well (r = 0.994) with the relative capacity of these peptides to inhibit 125I‐GRP specific binding. Pretreatment with unlabelled GRP for up to 6 h caused only a slight decrease in both specific 125I‐GRP binding and the affinity labelling of the Mr 75,000–85,000 protein. We also show that the cross‐linked complex is a glycoprotein. First, solubilized affinity labelled Mr 75,000–85,000 complex applied to wheat germ lectin‐sepharose columns was cluted by addition of 0.3 M N‐acetyl‐D‐glucosamine. Second, treatment with endo‐β‐N‐actylglucosaminidase F reduced the apparent molecular weight of the affinity‐labelled band from 75,000–85,000 to 43,000, indicating the presence of N‐linked oligosaccharide groups.
Interferon‐α‐(IFN) producing CHO cell lines are resistant to the antiproliferative activity of IFN: A correlation with gene expression
Authors: van Heuvel M. , Govaert‐Siemerink M. , Bosveld I. J. , Zwarthoff E. G. , Trapman J. 
Published: December 2004

AbstractCHO cell lines that constitutively produce the murine interferon‐α (IFN‐α) subspecies α4 and α6 were constructed. The producer cell lines were protected against viral (vesicular stomatitis virus) infection by the IFN species secreted, but were resistant to the growth inhibitory activity of the IFN species. As compared with α4, the α6 protein displayed a high antiproliferative activity when added to normal CHO cells, which correlates completely with the high antiviral activity of a6 on these cells. Three messenger ribonucleic acid (mRNA) species, which are normally induced in CHO cells by IFN treatment (1–8, 2–5A synthetase, and ISG 15) were constitutively present in CHO producer cell lines. The level of another mRNA (ISG 54), however, was very low in the producer cells as compared with its expression in short‐term IFN‐treated cells. These data indicate that 1–8, 2–5A synthetase and ISG 15 are not involved in the antigrowth activity of IFN in this system, but rather suggest a function of ISG 54 in this respect.
Tissue plasminogen activator and acute pulmonary embolism
Authors: Goldhaber Samuel Z. , Kessler Craig M. , Heit John , Markis John E. , Sharma G. V. R. K. , Dawley Douglas L. , Meyerovitz Michael F. , Vaughan Douglas E. , Parker J. Anthony , Come Patricia C. , Kim Ducksoo , Selwyn Andrew P. , Loscalzo Joseph , Braunwald Eugene 
Published: December 2004

AbstractWe assessed the efficacy and safety of peripheral intravenous recombinant human tissue‐type plasminogen activator (rt‐PA) in 47 patients with angiographically documented pulmonary embolism (PE). We administered 50 mg/2 h and, if necessary, an additional 40 mg/4 h. By 6 hours, 94% of the patients had angiographic evidence of clot lysis that was slight in 5, moderate in 12, and marked in 27 patients. Among the 34 patients with pulmonary hypertension prior to treatment, average pulmonary artery pressure decreased from 43/17 (27) to 31/13 (19) mm Hg (P < 0.0001). The average lung scan perfusion defect decreased from 37% before therapy to 16% (P < 0.01) after therapy among the 19 patients who had pre‐ and post‐treatment lung scans. Of 7 patients with pre‐ and post‐treatment imaging and Doppler echocardiograms, hypokinetic right ventricular wall movement (mild in 1, moderate in 2, and severe in 4) normalized in 5 and improved to mild hypokinesis in 2. Right ventricular diameter decreased from 3.9 ± 1.0 to 2.0 ± 0.5 cm (P < 0.005). Fibrinogen decreased 33% from baseline at 2 h and 42% from baseline at 6 h. However, patients with the greatest degree of angiographic clot lysis at 2 h had a preponderance of fibrinogenolysis over fibrinolysis, demonstrated by a lower ratio of cross‐linked fibrin degradation products to fibrin(ogen) degradation products (0.14 ± 0.09 vs. 0.54 ± 0.82) (P < 0.04). Among selected patients, peripheral intravenous rt‐PA is associated with rapid lysis of PE, improved pulmonary perfusion, and improved right ventricular function.
Cytotoxicity mediated by tumor necrosis factor in variant subclones of the ME‐180 cervical carcinoma line: Modulation by specific inhibitors of DNA topoisomerase II
Authors: Coffman Frederick D. , Green Lora M. , Godwin Angela , Ware Carl F. 
Published: December 2004

AbstractThe mechanism of tumor necrosis factor (TNF)‐induced cytotoxicity has been investigated using two clonal variants of the ME‐180 human cervical carcinoma cell line. The clonal lines were characterized with respect to their expression of TNF receptors, kinetics of cell death, and their ability to communicate intercellularly through gap junctions. The ME‐180.4 and ME‐180.8 clones were identified by their relative sensitivity to TNF induced lysis in a 24‐h assay. The dose of TNF required to kill 50% of the target cells was 60 pM for the sensitive ME‐180.4 and 2.5 nM for the ME‐180.8. However, when assay times were extended, the dose response for both clones was the same, indicating that a difference in the kinetics of cell death and not absolute TNF sensitivity existed between the ME‐180.4 and ME‐180.8 clones. Both clones were gap junction deficient as judged by their inability to transfer Lucifer yellow or 6‐carboxyfluorescein, a characteristic phenotype of cells sensitive to cytotoxicity by TNF. The level of surface receptor expressed on these clones was nearly identical with a Kd = 0.3 nM and 5,000 binding sites per cell. Measurement of the kinetics of cell death revealed that the time between the addition of TNF and the onset of observed cell death (induction phase) was much shorter for the ME‐180.4 (32–55 h) than for the resistant ME‐180.8 (55–80 h). Mitomycin C, a DNA alkylating agent, significantly reduced the length of the induction phase for both clones, although the kinetic difference between the clones remained unchanged. Two epipodophyllotoxins, VP‐16 and VM‐26, which specifically inhibit the rejoining activity of DNA topoisomerase II, showed a 10–100‐fold synergistic effect when combined with TNF as shown by isobologram analysis. VM‐26 when added to the resistant ME‐180.8 clones decreased the length of induction phase and abolished the kinetic difference observed with the ME‐180.4 clone. These results indicate that the variance in the TNF response of these two clones was closely associated with DNA topoisomerase II, and suggest that this enzyme may play an important role in TNF mediated cytotoxicity.
Role for interleukin‐1 in the pathogenesis of hypersensitivity diseases
Authors: Dinarello Charles A. , Endres Stefan 
Published: December 2004

AbstractInterleukin‐1 (IL‐1), a polypeptide product of various cells, is one of the key mediators of the body's response to microbial invasion, inflammation, immunological reactions, and tissue injury. IL‐1 is a prominent member of a group of polypeptide mediators now called “cytokines.” Current evidence suggests that IL‐1 is not produced in health but that any perturbation such as inflammation or even slight injury triggers the expression of IL‐1 genes. The biological effects of IL‐1 are manifested in nearly every tissue and organ. These include various proinflammatory effects such as increased production of arachidonate metabolites, synovial cell proteases, activation of basophils, eosinophils and neutrophils, endothelial cell adhesiveness, and stimulation of lymphocyte responses. Control of IL‐1 synthesis in certain diseases is often appropriate. Although corticosteroids reduce both the transcription and translation of IL‐1, we have recently investigated the effect of dietary supplementation with N‐3 (omega‐3) fatty acids in human volunteers. The results indicate that increasing the amount of N‐fatty acids in the diet decreases the ability of blood mononuclear cells to synthesize IL‐1 in vitro. It is suggested that the ameliorative effects of N‐3 fatty acid dietary supplements in patients with hypersensitive diseases may be, in part, the result of decreased IL‐1 production.
Modulation of IL‐4‐induced human IgE production in vitro by IFN‐γ and IL‐5: The role of soluble CD23 (s‐CD23)
Authors: Pène J. , Chrétien I. , Rousset F. , Brière F. , Bonnefoy J. Y. , de Vries J. E. 
Published: December 2004

AbstractIL‐4 specifically induced IgE production by peripheral blood lymphocytes or by tonsil or spleen cells from healthy donors. IL‐4‐induced IgE synthesis was dependent on CD4+ T cells and monocytes and was blocked by IFN‐γ, IFN‐α, and prostaglandin E‐2 (PGE‐2). These substances also inhibited IL‐4‐induced CD23 expression and subsequent release of soluble CD23 (s‐CD23). In addition, IgE production was blocked by F(ab′)2 fragments of an mAb against CD23. In contrast, IL‐5 enhanced IL‐4‐induced IgE production, provided IL‐4 was added at nonsaturating concentrations. This increase in IgE production correlated quantitatively with an enhanced release of s‐CD23. Collectively, these results indicate that there is a correlation between s‐CD23 release and IgE production. However, s‐CD23 fractionated from supernatants of the lymphoblastoid cell line RPMI‐8866 was ineffective in inducing IgE production in the absence of IL‐4, but acted synergistically with suboptimal concentrations of IL‐4. In addition, it is demonstrated that alloreactive T‐cell clones produced varying concentrations of IL‐4, IL‐2, or IFN‐γ upon stimulation. Only supernatants of 2/4 of these T‐cell clones induced a low degree of IgE synthesis, but in the presence of anti‐IFN‐γ antibodies, all four supernatants induced a strong induction of IgE production. This IgE synthesis was blocked specifically by anti‐IL‐4 antibodies, indicating that IL‐4 is the sole inducer of IgE synthesis. Our findings demonstrate that IL‐4‐induced IgE production involves complex interactions of T cells, B cells, and monocytes and is positively modulated by IL‐5 and s‐CD23 but down‐regulated by IFN‐γ, IFN‐α, and PGE‐2, respectively.
An expression system for trypsin
Authors: Vasquez John R. , Evnin Luke B. , Higaki Jeffrey N. , Craik Charles S. 
Published: December 2004

AbstractThe eukaryotic serine protease, rat anionic trypsin, and various mutants created by site‐directed mutagenesis have been heterologously expressed in Escherichia coli. The bacterial alkaline phosphatase (phoA) promoter was used to control the expression of the enzymes in an induced or constitutive fashion. The DNA coding for the eukaryotic signal peptide of pretrypsinogen was replaced with DNA coding for the phoA signal peptide. The phoA signal peptide successfully directs the secretion of the mammalian trypsinogen to the periplasmic space of E. coli. Active trypsin was expressed in the periplasm of E. coli by deleting the DNA coding for the activation hexapeptide of the zymogen. The activity of trypsin in the periplasm suggests that the enzyme is correctly activated and has folded such that the 12 cysteine residues involved in the six disulfide bonds of rat anionic trypsin have paired correctly.A transcription terminator increased the level of expression by a factor of two. However, increasing the copy number of the plasmid decreased the levels of expression. Localization of the active enzyme in the periplasm allows rapid screening of modified trypsin activities and facilitates the purification of protein to homogeneity and subsequently to crystallinity.