bluuebunny/crossref_metadata_2025_split huggingface dataset:
The sub‐membrane reticulum of the human erythrocyte: A scanning electron microscope study
Authors: Hainfeld James F. , Steck Theodore L. 
Published: May 2005

AbstractA web‐like reticulum underlying the human erythrocyte membrane was studied at a resolution of 5–10 nm by means of a scanning electron microscope. The network was visualized in isolated membranes (ghosts) torn open to reveal their interior space and in residues derived from ghosts extracted with Triton X‐100. It formed a continuous (rather than patchy) cover over the entire cytoplasmic surface, except where lifted off or torn away. Filaments (5–40 nm in diameter), annular figures (40–60 nm in diameter), and nodes (30–100 nm in diameter) were prominent in different networks. The dimensions of the filaments and the interstices in the reticulum varied with conditions, suggesting that the network has elastic properties. This reticulum is probably related to the erythrocyte membrane proteins spectrin and actin.
Defective transport of thymidine by cultured cells resistant to 5‐bromodeoxyuridine
Authors: Lynch Thomas P. , Cass Carol E. , Paterson Alan R. P. 
Published: May 2005

AbstractA line of HeLa cells resistant to 5‐bromo‐2′‐deoxyuridine (BUdR) was established by continuous culture in growth medium containing BUdR; during the selection period, BUdR concentrations, initially 15 μM, were gradually increased to 100 μM. Cells of a clone (HeLa/B5) established from this line were also resistant to 5‐fluoro‐2′‐deoxyuridine (FUdR), but not to the free base, 5‐fluorouracil. Although extracts of HeLa/B5 cells exhibited levels of thymidine kinase activity comparable to those of parental cells, rates of uptake of BUdR, FUdR, and thymidine into intact cells were much reduced. The kinetics of uptake of uridine and adenosine, nucleosides which appear to be transported independently of thymidine in HeLa cells, were similar for HeLa/B5 and the parental line (HeLa/0). Relative to thymidine uptake by HeLa/0 cells, that by HeLa/B5 cells was distinctly less sensitive to nitrobenzlthionosine (NBMPR), a specific inhibitor of nucleoside transport in various types of animal cells. Despite this difference in NBMPR sensitivity, both cell lines possessed the same number of high affinity NBMPR binding sites per mg cell protein. The altered kinetics of thymidine uptake and the NBMPR insensitivity of that function in HeLa/B5 cells suggest that resistance to BUdR is due to an altered thymidine transport mechanism.
Association of (Ca+Mg)‐ATPase activity with ATP‐dependent Ca uptake in vesicles prepared from human erythrocytes
Authors: Quist Eugene E. , Roufogalis Basil D. 
Published: May 2005

AbstractGhost membranes prepared from human erythrocytes exhibit 2 distinct (Ca+Mg)‐ATPase1 activities (Quist and Roufogalis, Arch Biochem Biophys 168:240, 1975). (Ca+Mg)‐ATPase activity dependent on a water soluble protein fraction is selectively lost from ghost membranes during preparation of vesicles under low ionic strength, slightly alkaline conditions. In this study, the Ca2+ dependence of the remaining membrane bound (Ca+Mg)‐ATPase activity and ATP‐dependent Ca uptake in vesicles were compared. The C2+ activation curves for (Ca+Mg)‐ATPase activity and Ca uptake into vesicles were parallel over a Ca2+ range of 0.3–330 μM, and both curves have 2 apparent KA values for Ca2+ of 0.45 and 100 μM. Addition of a concentrated soluble protein fraction containing predomintly spectrin to the vesicles increased (Ca+Mg)‐ATPase activity over twofold but did not affect the rate of Ca uptake. These findings suggest that the (Ca+Mg)‐ATPase activity remaining in vesicles after extraction of the water soluble proteins is associated with the Ca pump whereas (Ca+Mg)‐ATPase activity dependent on the soluble protein fraction is associated with some other function.
Methods for rapidly altering the permeability of mammalian cells
Authors: Heppel Leon A. , Makan Nizar 
Published: May 2005

AbstractVarious agents alter mammalian cells so that they rapidly become nonspecifically permeable to substances that ordinarily do not penetrate intact cells. Thus, toluene renders liver cells permeable to nucleotides and macromolecules. Tween 80 and Tween 60 act in similar fashion, and the effect is reversible. Dextran sulfate reversibly alters the permeability of Ehrlich ascites tumor cells, which offers a tool for studying the control of macromolecular syntheses and other processes. Brief exposure to external ATP alters the permeability of certain transformed mouse cells but not of untransformed cells. The effect of ATP is rapidly reversible.
L‐arabinose transport and the L‐arabinose binding protein of escherichia coli
Authors: Hogg Robert W. 
Published: May 2005

AbstractThe active accumulation of L‐arabinose by arabinose induced cultures of Escherichia coli is mediated by 2 independent transport mechanisms. One, specified by the gene locus araE, is membrane bound and possesses a relatively “low affinity.” The other, specified in part by the genetic locus araF, contains as a functional component the L‐arabinose binding protein and functions with a “high affinity” for the substrate. The L‐arabinose binding protein has been purified, partially characterized, crystallized, and sequenced.
Microbial carbohydrate specific antibodies distinguish between different stages of differentiating mouse cerebellum
Authors: Trenkner Ekkhart , Sarkar Siddhartha 
Published: May 2005

AbstractHigh titered anticarbohydrate antibodies were used to identify cell surface carbohydrates during different stages in histogenesis of mouse cerebellum in a micro tissueculture system which mimics selected features of in vivo cerebellum development. Blockage of fiber formation within the first few days in vitro and inhibition of cell migrations by carbohydrate‐specific antibodies served as an assay system for possible contributions of surface carbohydrates to the behavior of developing cerebellar cells. Microbial strains were selected on the basis of carbohydrate structures of their cell wall antigens, and anticarbohydrate antibodies were raised against treated whole bacteria and yeast in rabbits. We found that antibodies to mannan were active at all stages of development tested (embryonic day 13, E13; the day of birth, P0; and postnatal day 7, P7). Antibodies to sialic acids prepared against strains B and C of Neisseria meningitidis distinguish different subterminal structures: anti‐B reacted with E 13 and P0 cerebellar cells, and anti‐C mostly with cells older than P7. Antifetuin antibody recognized E 13 and P0 but not P7 cell populations. Pneumococcus C strain R36A‐specific antibodies were effective only after coating cells to C type carbohydrate before application of the antibody. The results demonstrate that antimicrobial carbohydrate antibodies cross‐react with mammalian cell surface carbohydrate structures and therefore can be used as a powerful tool in tissue culture to analyze those structures which might control cell behaviors pertinent to cerebellar development.
Subcellular fractionation of brown adipose tissue
Authors: Giacobino J.‐P. 
Published: May 2005

AbstractThe present study proposes a technique, using Metrizamide, which permits the preparation of brown adipose tissue plasma membranes from the crude mitochondria as well as from the crude microsome fraction. These plasma membranes have high relative specific activities of their marker enzyme, 5′‐nucleotidase (15 ± 3 and 14 ± 2 respectively) and, particularly those originating in the crude microsomes, are relatively free of mitochondria contamination. This study also shows the influence of the mode of cell disruption on microsome integrity. When cell disruption was achieved by grinding in liquid nitrogen the purified microsome NADPH cytochrome c reductase specific activity was found to be 3.5 times greater than that of microsomes obtained after homogenization of the tissue.
Alkaline phosphatase of basal lateral and brush border plasma membranes from intestinal epithelium
Authors: Hanna S. D. , Mircheff A. K. , Wright E. M. 
Published: May 2005

AbstractThe alkaline phosphatases present on isolated brush border and basal lateral membranes of rat duodenal epitheilum were examined by means of a variety of biochemical assays and physical methods. The two alkaline phosphatases have similar pH optima of 9.6–9.8, similar substrate km's for p‐nitrophenyl phosphate (PNPP) of 71 micromolar, similar responses to the inhibitors 2‐mercaptoethanol, theophylline, phenylalanine, and ethylenediaminetetraacetic acid (EDTA), similar sensitivities to calcium, magnesium, zinc, sodium, and potassium, and similar insensitivities to digestion with trypsin or papain. The two enzymes also exhibit similar molecular weights on SDS‐polyacrylamide gels in the range 124,000–150,000, and both enzymes show an Rf value of 0.092 on Triton X‐100 polyacrylamide gels, indicating similar intrinsic charges. The Vmax of the brush border enzyme is ten times greater than that of the basal lateral enzyme, 140 μmoles/mg‐h as opposed to 14 μmoles/mg‐h. The differences in Vmax are a reflection of the known distribution of alkaline phosphatase in rat duodenum, there being more alkaline phosphatase activity present on the brush border than on the basal lateral surface. One other major difference was observed between the two enzymes, the stimulation of the basal lateral and not the brush border alkaline phosphatase by SDS, Triton X‐100, or cholate. We conclude that the enzymes are very similar to one another and probably perform similar membrane functions.
Growth rate and chromosome number of tumor cell lines with different metastatic potential
Authors: Cifone Maria A. , Kripke Margaret L. , Fidler Isaiah J. 
Published: May 2005

AbstractWe investigated whether the metastatic potential of various tumor cell lines was related to chromosome counts or to rate of growth in vitro or in vivo. Clones of known metastatic potential derived from a C3H− fibrosarcoma induced by UV radiation (UV–2237) and from C57BL/6 B16 melanoma were tested for these characteristics. No correlation was found between the growth rate of these clones in monolayer culture or at a subcutaneous site and their ability to produce metastases. The cells from clones of UV‐2237 were mainly in the diploid range with only one exception, and the B16 clones were all hyperploid. Thus, there was also no correlation between malignant behavior of the clones and gross changes in chromosome number.
Ganglioside patterns of metastatic and non‐metastatic transplantable hepatocellular carcinomas of the rat
Authors: Kloppel Thomas M. , Morré D. James , Jacobsen Linda B. 
Published: May 2005

AbstractIn previous investigations, we correlated levels of sialic acid, gangliosides, and ganglioside glycosyltransferases with tumorigenesis over a 24‐week continuum of growth of hepatocellular neoplasms of the rat induced by the carcinogen N‐2‐fluorenylacetamide. However, metastatic tumors developed only rarely and were not analyzed. To investigate surface changes associated with metastasis, well‐differentiated and poorly differentiated hepatocellular carcinomas were transplanted to syngeneic recipient rats. From those, several metastatic and nonmetastatic isolates were obtained and compared. Both total and ganglioside sialic acid amounts in transplantable hepatomas were elevated above control liver values but were significantly lower for metastatic lines than for nonmetastatic lines. The nonmetastatic lines were characterized by ganglioside patterns depleted in the precursor ganglioside GM3 (sialic acid‐galactose‐glucose‐ceramide) and elevated in the products of the monosialoganglioside pathway. In contrast, metastatic isolates exhibited a restoration of GM3 and nearer normal amounts of other gangliosides. The findings point to differences in sialic acid‐containing glycolipids, comparing metastatic and nonmetastatic hepatocellular carcinomas, and further extend the concept that ganglioside alterations do not cause tumorigenesis but are the end result of a cascade of events which apparently continue beyond the onset of metastasis.
Selection of malignant melanoma variant cell lines for ovary colonization
Authors: Brunson Kenneth W. , Nicolson Garth L. 
Published: May 2005

AbstractMurine melanoma line B16‐F1, which shows some specificity for metastatic organ colonization of lung but rarely metastasizes to ovary, was used to select variant cell lines with increased preference for experimental ovary metastasis. Ovary‐colonizing melanoma cell lines were sequentially selected in syngeneic C57BL/6 mice by repeated intravenous administration and surgical recovery of ovarian melanoma tumors for tissue culture. After ten selections for experimental ovary metastasis, line B16‐010 was established which formed experimental metastatic ovary tumors in almost every test animal. In tissue culture B16–010 cells grew in circular in circular colonies with rounded, smooth cell peripheries compared to B16‐F1 cells which were flatter, grew in irregular patterns, and exhibited long cellular projections. Ovary‐selected B16 lines contained less melainin pigment (B16‐010 < B16‐05 < B16‐01 ≅ B16‐F1) compared to the parental melanoma line. Together with previous cloning and selection data, these results are consistent with the preexistence of highly malignant cells in the parental tumor population that possess the ability to metastasize to specific organs.
Alteration of insulin binding and cytoskeletal organization in cultured fibroblasts by tertiary amine local anesthetics
Authors: Raizada Mohan K. , Fellows Robert E. 
Published: May 2005

AbstractTertiary amine local anesthetics cause a time‐ and dose‐dependent, reversible increase in insulin binding sites in cultured chick embryo fibroblasts. Incubation of fibroblasts with 0.2 mM dibucaine for 3 h at 37°C results in a twofold to threefold increase in insulin binding, with an increase in average number of binding sites (Ka = 3.0 × 107M−1) from 9 × 103 to 29 × 103 per cell. Trypsin or ethylenegly coltetraacetic acid (EGTA) alone increases insulin binding twofold to threefold, but fails to further increase 125I‐insulin binding in cells pretreated with dibucaine. Transformation of chick embryo fibroblasts with Rous sarcoma virus causes a threefold to fivefold increase in insulin binding, which is not further increased by incubation with dibucaine. As demonstrated by transmission electron microscopy, dibucaine and trypsin also induce changes in the cytoskeleton of chick embryo fibroblasts, characterized by disorganization and disappearance of microfilament and microtubule bundles. These alterations are accompanied by gross morphologic changes, including rounding of cells and appearance of numerous ruffles and blebs on the cell surface. These observations are consistent with the hypothesis that expression of surface receptors in cultured chick embryo fibroblasts is related to the organization and disorganization of cytoskeletal structures.
Production of monoclonal antibodies against a cell surface concanavalin a binding glycoprotein
Authors: Starling James J. , Simrell Charles R. , Klein Paul A. , Noonan Kenneth D. 
Published: May 2005

AbstractConcanavalin A‐binding (Con A)‐binding cell surface glycoproteins were isolated, via Con A‐affinity chromatography, from Triton X‐100‐solubilized Chinese hamster ovary (CHO) cell plasma membranes. The Con A binding glycoproteins isolated in this manner displayed a significantly different profile on sodium dodecyl sulfate‐polyacrylamide gels than did the Tritonsoluble surface components, which were not retarded by the Con A‐Sepharose column. [125I]‐Con A overlays of the pooled column fractions displayed on sodium dodecyl sulfate‐polyacrylamide gel electro‐phoresis (SDS‐PAGE) demonstrated that there were virtually no Con A receptors associated with the unretarded peak released by the Con A‐Sepharose column, whereas the material which was bound and specifically eluted from the Con A‐Sepharose column with the sugar hapten α‐methyl‐D‐mannopyranoside contained at least 15 prominent bands which bound [125I]‐Con A.In order to produce monoclonal antibodies against various cell surface Con A receptors, Balb/c mice were immunized with the pooled Con A receptor fraction. Following immunization spleens were excised from the animals and single spleen cell suspensions were fused with mouse myeloma P3/X63‐Ag8 cells. Numerous hybridoma clones were subsequently picked on the basis of their ability to secrete antibody which could bind to both live and glutaraldehyde‐fixed CHO cells as well as to the Triton‐soluble fraction isolated from the CHO plasma membrane fraction. Antibody from two of these clones was able to precipitate a single [125I]‐labeled CHO surface component of ∼265,000 daltons.
Modulation of cell surface iron transferrin receptors by cellular density and state of activation
Authors: Larrick James W. , Cresswell Peter 
Published: May 2005

AbstractThis report describes investigations of plasma membrane transferrin receptors on a variety of lymphoid cell lines and normal peripheral blood lymphocytes during activation and cell growth cycles. Transformed lymphoid cell lines have as many as 1,000 times the number of receptors found on normal resting lymphocytes. The number of iron transferrin receptors on continuous cell lines as well as normal human fibroblasts is down‐regulated during the transition from log‐phase growth to stationary plateau growth. When normal lymphocytes are transformed by mixed lymphocyte culture or mitogens, they rapidly express a 50‐fold increase in the number of transferrin binding sites. This appearance of iron transferrin receptors anticipates nuclear changes during cell activation and subsequent mitosis of normal cells.
The role of colony‐stimulating factor in granulopoiesis
Authors: Shadduck Richard K. , Pigoli Giuseppe , Waheed Abdul , Boegel Florence 
Published: May 2005

AbstractThe proliferation and maturation of granulocytic‐monocytic stem cells appears to be controlled by a series of closely related glycoproteins termed “colony‐stimulating factors” (CSFs). Recently, we devised a 6‐step scheme for the purification of murine fibroblast (L‐cell)‐derived CSF. Ten liter pools of conditioned media were concentrated by ultrafiltration, precipitated by ethanol, and separated on DEAE cellulose, Con‐A Sepharose, and Sephadex G 150. The CSF was separated from trace contaminants, including endotoxin, by density gradient centrifugation. The purified material was radioiodinated and used to define the serum half‐life and in vivo distribution. Following IV injection there was a biphasic serum clearance with a t½ of 24–40 min and 2–2½ hours in the first and second phases. Approximately 25% of the tracer was excreted in the urine at 6 h; however, urinary radioactivity was due to low molecular weight peptides. Simultaneous studies by radioimmunoassay showed a similar rapid serum clearance of unlabeled CSF but virtually no urinary CSF activity. Thus, assays for urinary CSF may not provide useful measures of in vivo CSF activity. Further in vitro studies have defined the interaction of CSF with responsive cells in the marrow. Varying doses of CSF were incubated with 107 marrow cells for intervals of 24–48 h. The major increment in cell‐associated radioactivity occurred between 6 and 16 h. The reaction was saturable with 1–2 ng/ml CSF. Binding was prevented by cold CSF, but not by other proteins. Irradiation yielded only a minimal reduction in CSF binding. The interaction of CSF with marrow cells appeared to require new protein synthesis, as binding was completely inhibited by cycloheximide and puromycin. Irradiated mice injected with antibodies to CSF showed an inhibition of granulopoiesis by marrow cells in peritoneal diffusion chambers; however, granulopoiesis in the intact bone marrow was unaffected. Granulpoiesis in long‐term marrow cultures was also unaffected by anti‐CSF. These different responses may be due to accelerated clearance of injected CSF in nonirradiated mice or to extensive stromal interactions that modulate and perhaps control granulocytic differentiation in the intact bone marrow microenvironment.
Direct linkage of EGF to its receptor: Characterization and biological relevance
Authors: Linsley Peter S. , Fox C. Fred 
Published: May 2005

AbstractA small portion of the 125I‐EGF that binds specifically to intact cells or isolated membranes from a variety of sources becomes directly and irreversibly linked to EGF receptors. This provides a simple technique for affinity labeling the EGF receptor. Membranes isolated from the human epidermoid carcinoma cell line A431, which posesses extraordinarily high numbers of EGF receptors, gave rise to three major direct linkage complexes of MW = 160,000, 145,000, and 115,000. The time course for formation of each is similar, showing that 125I‐EGF can form direct linkage complexes with several preexisting forms of the EGF receptor. The direct linkage of EGF to receptor is slow in comparison to 125I‐EGF binding, but both processes have similar susceptibilities to competition by unlabeled EGF.EGF was modified chemically with the amino site‐specific reagent, N‐hydroxysuccinimidyl biotin. The biotinyl‐EGF had a reduced capacity to engage in direct linkage complex formation with no concomitant reduction in its ability to bind to EGF receptors. Since native and biotinyl EGF have identical abilities to stimulate the uptake of 3H‐thymidine into DNA when incubated with cultured murine 3T3 cells, the direct linkage of EGF to its receptor does not appear to play an important role in EGF‐stimulated mitogenesis.
Controlled proteolysis of EGF receptors: Evidence for transmembrane distribution of the EGF binding and phosphate acceptor sites
Authors: Linsley Peter S. , Fox C. Fred 
Published: May 2005

AbstractA small quantity of the 125I‐EGF (epidermal growth factor) bound specifically to EGF receptors on the human epidermoid carcinoma cell line A431 associates covalently. The direct linkage complex formed migrates during gel electro‐phoresis as a single diffuse band of MW = 160,000–170,000. In contrast, direct linkages complexes of 160,000, 145,000, and 115,000 daltons are formed when EGF is incubated with membranes isolated from these cells; these arise from EGF receptor modification during membrane isolation. None of these modifications affected the affinity of the EGF binding site for 125I‐EGF.The electrophoretic mobilities of the MW = 160,000 and 145,000 direct linkage complexes were similar to those of the major 32Pi‐labeled products of the EGF‐stimulated phosphorylation reaction described by Carpenter et al [Nature 276:409–410, 1978], indicating that proteolytic fragments of EGF receptors are the major phosphate acceptors in this reaction. EGF receptors on intact A431 cells accepted phosphate effectively from γ‐32Pi‐ATP only when the cells were permeabilized with lysolecithin. This shows that the EGF binding and phosphate acceptor sites lie on opposing faces of the membrane. When the 145,000 dalton form of receptor is labeled with EGF or 32Pi and the labeled peptides subjected to tryptic hydrolysis under identical conditions, all phosphates is lost from high molecular weight products under conditions where the EGF‐receptor covalent complex is converted largely to a 115,000 dalton form. This suggests that the phosphate acceptor site lies on the cytoplasmic side of the membrane on a region of receptor extending 30,000 daltons from the 115,000 dalton fragment containing the EGF binding site.