bluuebunny/crossref_metadata_2025_split huggingface dataset:
Ionophore‐induced disassembly of blood platelet microtubules: Effect of cyclic AMP and indomethacin
Authors: Kenney Dianne M. , Chao Francis C. 
Published: February 2005

AbstractCytoplasmic calcium levels are believed to be important in blood platelet activation. Upon activation, the discrete marginal microtubule band, which maintains the discoid shape of non‐activated platelets, becomes disrupted. Present studies demonstrate that the extent of assembly of the marginal microtubule band is related to cytoplasmic calcium levels. The divalent cationophore, A23187, causes platelet aggregation, secretion, and contraction by promoting calcium transport from intraplatelet storage sites into the cytoplasm. A23187 caused disassembly of platelet microtubules. Quantitation of electron micrographs revealed that numbers of microtubules were reduced by approximately 80% after A23187 treatment. Secondly, assembled microtubules in homogenates of platelets, in which microtubules were stabilized prior to homogenization, were decreased in favor of free tubulin in A23187‐treated platelets. Thirdly, A23187 increased 14C‐colchicine binding by intact platelets; this also indicated a shift in the microtubule subunit equilibrium to favor free, colchicine‐binding tubulin subunits. In control experiments, A23187 did not affect the stability of platelet tubulin, the colchicine binding reaction, or the total tubulin content of platelets. Stimulation of colchicine binding depended on A23187 concentration (0.05–0.5 μM) and did not require extracellular calcium. A23187‐stimulation of colchicine binding was blocked by dibutyryl cyclic AMP (0.80 mM) and/or 3‐isobutyl‐1‐methylxanthine (50 μM) and by indomethacin (10 μM). Cyclic AMP or indomethacin also interferes with A23187‐induced platelet activation, but indomethacin is not likely to completely inhibit the perturbation of intraplatelet calcium gradients by A23187. It is suggested that A23187‐induced microtubule disassembly may be an indirect effect of calcium on microtubules.
Colony formation in agar by adult bone marrow multipotential hemopoietic cells
Authors: Johnson G. R. 
Published: February 2005

AbstractErythroid colony formation in agar cultures of CBA bone marrow cells was stimulated by the addition of pokeweed mitogen‐stimulated spleen conditioned medium (SCM). Optimal colony numbers were obtained when cultures contained 20% fetal calf serum and concentrated spleen conditioned medium. By 7 days of incubation, large burst or unicentric erythroid colonies occurred at a maximum frequency of 40–50 per 105 bone marrow cells. In CBA mice the cells forming erythroid colonies were also present in the spleen, peripheral blood, and within individual spleen colonies. A marked strain variation was noted with CBA mice having the highest levels of erythroid colony‐forming cells.In CBA mice erythroid colony‐forming cells were mainly non‐cycling (12.5% reduction in colony numbers after incubation with hydroxyurea or 3H‐thymidine). Erythroid colony‐forming cells sedimented with a peak of 4.5 mm/hr, compared with CFU‐S, which sedimented at 4.25 mm/hr. The addition of erythropoietin (up to 4 units) to cultures containing SCM did not alter the number or degree of hemoglobinisation of erythroid colonies.Analysis of the total number of erythroid colony‐forming cells and CFU‐S in 90 individual spleen colonies gave a correlation coefficient of r = 0.93 for these two cell types.In addition to benzidine‐positive erythroid cells, up to 40% of the colonies contained, in addition, varying proportions of neutrophils, macrophages, eosinophils, and megakaryocytes. Taken together with the close correlation between the numbers of CFU‐S in different adult hemopoietic tissues, including individual spleen colonies, the data indicate that the erythroid colony‐forming cells expressing multiple hemopoietic differentiation are members of the hemopoietic multipotential stem cell compartment.
Polyadenylate‐deficient analogues of poly(A)‐containing mRNA sequences in cultured AKR mouse embryo cells
Authors: Siegal G. P. , Hodgson C. P. , Elder P. K. , Stoddard L. S. , Getz M. J. 
Published: February 2005

AbstractFive to six percent (by mass) of AKR‐2B mouse embryo cell polysomal RNA consists of messenger RNA sequences which may exist in polyadenylated form. In the steady state, however, only 30–40% of these molecules are retained by extensive passage over oligo(dT)‐cellulose, the remainder being present in the form of poly(A)‐deficient analogues. Within experimental limits, these poly(A)‐deficient analogues contain representatives of all poly(A)‐containing mRNA sequences in these cells. An analysis of the kinetics of hybridization of cDNA probes enriched for either abundant or rare poly(A)‐containing mRNA sequences suggests that the frequency distributions of poly(A)‐containing and poly(A)‐deficient analogues are dissimilar, and that a relationship exists between the intracellular frequency of a given mRNA sequence and the number of poly(A)‐deficient analogues of that sequence. High frequency sequences appear to be enriched in the poly(A)‐containing fraction, while low frequency sequences are predominately associated with the poly(A)‐deficient fraction, thus, poly(A) may play a role in the regulation of mRNA frequency in the cytoplasm.
Cell‐surface glycosaminoglycans: Turnover in cultured human embryo fibroblasts (IMR‐90)
Authors: Vogel Kathryn G. , Kendall Vaughan F. 
Published: February 2005

AbstractAs constituents of both extracellular matrix and the cell surface, glycosaminoglycans are in a strategic position to influence several basic cell features. The localization and turnover of glycosaminoglycans was investigated in cultured normal human embryo fibroblasts of lung origin (IMR‐90). Attention was directed particularly toward that compartment of the culture which could be released by gentle proteloysis (trypsin, 0.1 mg/ml, 15 min) and is considered to represent the cell surface. In the presence of Na2SO4, sulfated glycosaminoglycans (S‐GAGs) of the cell surface were labeled rapidly, but within 30 min some 35S‐GAG appeared in the extracellular medium. The intracellular pool of S‐GAGs labeled during a 10‐min period was lost during the first hr of chase with a half‐life of 18 min, compared with 16 hr for S‐GAGs labeled over a 48‐hr period. Pulse‐labeled S‐GAGs of the surface turned over with an initial half‐life of 60 min, compared with 7 hr for surface material labeled over a 48‐hr period. These rapid movements of the early chase period were followed by similar movement at a much slower rate. The results are consistent with a model in which most of the S‐GAGs synthesized in the cell move rapidly to the surface. The surface GAGs are then released immediately to the medium or accumulate at the membrane to be shed more slowly at a later time or to be degraded. The S‐GAG which left the cell layer most rapidly during chase was dermatan sulfate, while heparan sulfate made up an increasing percentage of the cell layer as chase progressed. These cultures produce a fibrillar matrix of fibronectin, but the kinetics of this study suggest that the S‐GAGs of the surface are membrane‐bound, and an extracellular glycosaminoglycan matrix does not form.
Transmembrane potential and amino acid transport in rat hepatocytes in primary monolaver culture
Authors: Wondergem Robert , Harder David R. 
Published: February 2005

AbstractTransmembrane potential (Em) and alpha‐aminoisobutyric acid (AIB) transport were measured in primary monolayer cultures of rat hepatocytes obtained from unoperated control rats and from rats 12 hr following partial hepatectomy. Measurements were performed 20–24 hr after plating the cells. The capacity of both kinds of cells to concentrate AIB depended upon extracellular sodium; however, the steady‐state accumulation in regenerating cells was twice that of control cells. Transmembrane potentials, recorded with glass microelectrodes, were –13 ± 0.6 mV and –27 ± 1.6 mV in control and regenerating cells, respectively. Ouabain (1 mM) depolarlized regenerating cells to –18 ± 1.0 mV, but it had no effect on control cells. The initial rates of 1 mM AIB transport into control and regenerating cells were 1.2 ± 0.1 and 3.1 ± 0.1 nanomoles/mg protein × 4 min, respectively. Ouabain (1 mM) reduced the initial rate of AIB transport into regenerating cells to 2.7 ± 0.1 nanomoles/mg protein × 4 min, but it had no effect on AIB transport into control cells. Glucagon (10−7 M) added to control cells 12 hr before measurements hyperpolarized Em to –31 ± 1.3 mV and increased AIB transport rate to 3.1 nanomoles/mg protein × 4 min. The results suggest a relationship between increases in Em and increases in AIB transport in rat hepatocytes. An electrogenic Na‐K pump may be involved in both of these events.
Age‐dependent decrease of growth responsiveness in density inhibited 3T3 cell populations and their interactions with SV 40‐3T3 cells
Authors: van Der Bosch Jürgen 
Published: February 2005

AbstractAn aging process has been detected in stationary 3T3 cell cultures, especially in the presence of plasma‐derived serum (PDS) from adult bulls. It leads to irreversible conversion of an increasing percentage of initially responsive cells of a stationary population into cells unresponsive to growth stimulation by newborn calf serum (NBCS) or reseeding at low cell density in the presence of NBCS. These unresponsive cells are viable in the sense that, following trypsinization, they reattach and spread on a new culture plate and can be maintained for many days. The conversion process is accelerated by increasing PDS concentration. It is antagonized by NBCS. It is accompanied by enhancement of growth‐inhibiting interactions exerted by stationary 3T3 cell populations on SV 40–3T3 cells.
Control and virus‐transformed baby hamster kidney cells resistant to ethidium bromide. I. Characterization and the respiratory enzymes
Authors: Soslau Gerald , Zavodny Paul J. 
Published: February 2005

AbstractCell lines resistant to ethidium bromide have been developed from cultured mammalian BHK21/C13 cells and these same cells transformed by Rous sarcoma virus (C13/B4). Cells resistant to 2 μg ethidium bromide per milliliter have been cloned. One clone of the control and one of the virus‐transformed cell lines has been employed for characterization. The resistant cells, in the presence of 2 μg ethidium bromide/ml, grow at approximately the same rate as the untreated parental cells. The control cells possess a “normal” karyotype (44 chromosomes), while the corresponding ethidium bromide mutant has a reduced chromsome number of 41 and a number of translocations. The mitochondria displayed morphological alterations compared to the parental lines during the transition phase prior to the isolation of the ethidium bromide‐resistant cells. The mitochondria of the ethidium bromide‐resistant mutants appear somewhat enlarged with a normal morphology.The effect of ethidium bromide on selected respiratory enzymes in normal and virus‐transformed ethidium bromide‐resistant baby hamster kidney cells was determined. Ethidium bromide‐resistant cells exhibited a depressed level of cytochrome aa3. This depression could not be reversed by growth in ethidium bromide‐free media. Ethidium bromide‐resistant cells possessed the same cytochrome b, c, and c1 levels per cell as their corresponding parental lines.Purified mitochondria isolated from virus‐transformed ethidium bromide‐resistant cells exhibited a depression in cytochrome oxidase‐specific activity, while the ethidium bromide‐resistant control cells did not. All cell lines studied showed a depression in NADH‐ferricyanide and NADH‐cytochrome c reductase‐specific activities relative to parental BHK21/C13 cells. An increase in succinate dehydrogenase and succinate cytochrome c reductase‐specific activities was observed in ethidium bromide‐resistant control cells relative to their parental BHK21/C13 cells. No increase was observed in virus‐transformed ethidium bromide‐resistant cells.Ethidium bromide‐resistant control cells exhibited a two‐fold increase in oligomycin‐insensitive adenosine triphosphatase activity relative to their parental cells. All of the cell lines studied possessed equivalent oligomycin‐sensitive adenosine triphosphatase‐specific activity except for the virus‐transformed, dyeresistant mutant, whose activity was increased.
Sodium butyrate affects expression of fibronectin on CHO cells: Specific increase in antibody‐complement‐mediated cytotoxicity
Authors: Milhaud Pierre , Yamada Kenneth M. , Gottesman Michael M. 
Published: February 2005

AbstractCellular fibronectin is expressed only at very low levels on the surface of the spontaneously transformed Chinese Hamster Ovary (CHO) cell line, as detected by immunofluorescence studies and confirmed by resistance of CHO cells to complement‐mediated lysis in the presence of anti‐fibronectin antibody. Treatment of CHO cells with sodium butyrate results in a marked susceptibility to anti‐fibronectin complement‐mediated killing. Using selected complement‐containing sera from rabbits, it is possible to demonstrate that killing of butyrate‐treated CHO cells is absolutely and specifically dependent on the presence of antibody to fibronectin. The increased susceptibility of butyrate‐treated cells to complement‐mediated killing correlates with an increase in cell surface fibronectin detected by immunofluorescence studies. This increased antigen expression should allow the isolation of mutants with altered regulation of fibronectin expression in a cell line of proven value for somatic cell genetic studies.
Variant endothelial cells. Fibronectin as a transducer of signals for migration and neovascularisation
Authors: McAuslan B. R. , Hannan G. N. , Reilly W. , Stewart F. H. C. 
Published: February 2005

AbstractA morphologic and growth control variant of bovine aortal endothelial cells has been isolated and shown to synthesise factor VIII antigen (McAuslan and Reilly '79). The variant also possesses the endothelial surface markers angiotensin converting enzyme and α2‐macroglobulin. The normal cell synthesises fibronectin and deposits it underneath the cells; the variant also synthesises fibronectin. At least three times more fibronectin is distributed over the upper cell surface of variants. This correlates with the three‐fold increased binding of the replication inhibitor Con A and suggests a role of fibronectin in endothelial cell growth control. When stimulated to migrate by CuII ions, the variant leaves deposits of fibronectin in its trail; in contrast, migrating normal cells do not, but they do redistribute their surface fibronectin. As revealed by scanning electron microscopy, variant cells are unusual in that they grow over or under cultured normal endothelial cells. It is proposed that during the process of neovascularisation, variant cells have a special function as lead cells that lay down fibronectin on which an endothelium can become established.
Differences in pattern and level of plasminogen activator production between a cloned cell line from an ethylnitrosourea‐induced glioma and one from normal adult rat brain
Authors: Hince Trevor A. , Roscoe Joan P. 
Published: February 2005

AbstractThe production of plasminogen activator (PA) by two cloned cell lines, one from an ethylnitrosourea‐induced glioma (Al5A5) and the other from normal adult rat brain (ARBO C9), has been investigated. Three assays were used to detect and measure PA in harvest fluids, cells and cell lysates. Similar levels were detected in harvest fluids from both cell lines. However, the cell and lysate assays indicated much higher levels in the tumor line. When actively growing cells were compared Al5A5 cells had approximately 16 × more fibrinolytic activity than the control cells with a limit of detection in the order of 103 cells or 1 μg protein (cell lysate). In contrast for the control cells PA could only be detected when upwards of 104 cells or 5 ‐ 10 μg protein were assayed. Plaminogen activator in as few as 103 tumor cells could be detected in the presence of 104 non‐tumor cells. Plasmingoen activator in 26 μg protein of Al5A5 cell lysate could also be detected in the presence of 44 μg protein from ARBO C9 lysates indicating no inhibitory activity in the control cell lysates. Levels of PA in both harvest fluids and cell lysates wre determined as cultures progressed through the growth cycle. For cell lysates this showed a build‐up of PA in the normal cell line as the cells approached and attained confluence. A much higher level was measured in the tumor cells soon after seeding and maintained to confluence. No differences in growth cycle‐associated changes in secreted PA could be determined in harvest fluids: both cell lines showing similar levels at confluence.
Evidence for control of protein synthesis in HeLa cells via the elongation rate
Authors: Nielsen Peter J. , McConkey Edwin H. 
Published: February 2005

AbstractThe effects of fresh medium and serum on protein synthesis in suspension‐cultured HeLa cells after growth to high cell density (>5 × 105 cells/ml) were studied. Cells which were resuspended in fresh medium plus serum and grown for 24 hours (control) were compared with cells grown for 2 hours after resuspension (stimulated). The spectrum of proteins being synthesized by control and stimulated cells does not appear to be grossly different; that is, the weight and number average molecular weights of newly synthesized whole‐cell protein are about the same in both cultures. Also, no significant differences were observed in the number of ribosomes per polysome or in the fraction of total ribosomes in polysomes. However, the transit times (combined elongation and termination times) were found to differ significantly; the average transit time for control cells was 2.24 minutes, while the average transit time for stimulated cells was 1.26 minutes. (An appendex evaluating the methodology involved in measuring the transit time is included.) In agreement with the difference in transit time, the absolute rate of protein synthesis in stimulated cells was approximately 1.8 times the rate measured in control cells. These data are taken as evidence that under certain conditions, the rate of elongtion and/or termination of polypeptide chains limits the overall rate of translation, and that cells can respond to growth conditions by changing the elongation and/or termination rate of protein synthesis.
Coordinate secretion of mouse alphafetoprotein, mouse albumin and rat albumin by mouse hepatoma‐rat hepatoma hybrid cells
Authors: Cassio Doris , Hassoux René , Dupiers Michèle , Uriel José , Weiss Mary C. 
Published: February 2005

AbstractMouse hepatoma cells that secrete large amounts of α‐fetoprotein (AFP) and albumin have been crossed with rat hepatoma cells that secrete only albumin, and in relatively small amounts, to investigate the influence of each parental genome upon the expression of serum proteins. All of the ten independent hybrid clones examined produce mouse AFP and both mouse and rat albumin; none produces rat AFP. The absence of production of rat AFP by the hybrids suggests that different mechanisms are involved in the initiation and in the maintenance of expression of this function. The secretion of the three proteins by the hybrid cells is coordinate: Whatever the growth phase (exponential or stationary) and irrespective of the amounts produced over a wide range, the ratio secreted of mouse AFP to mouse albumin is near to one, and that of mouse albumin to rat albumin is near to five. In addition, even though the pattern of protein secretion during the growth cycle of hybrid cells is different from those of both parents, the products of both parental genomes conform to the new hybrid pattern. Finally, some hybrids secrete less of the proteins with increasing numbers of cell generations, yet all three continue to be secreted in coordinate fashion. Since the rates of secretion of serum proteins probably reflect their rates of synthesis, we conclude that coordinate secretion indicates coordinate synthesis, and may reflect coordinate transcription of the relevant genes.
Spreading of human fibroblasts in serum‐free medium: Inhibition by dithiothreitol and the effect of cold insoluble globulin (plasma fibronectin)
Authors: Grinneli Frederick , Feld Marian K. 
Published: February 2005

AbstractWe have tested the effect of dithiothreitol (DTT) treatment on the initial spreading of human fibroblasts in serum‐free medium in tissue culture dishes. Cell spreading was inhibited following treatment of these cells with 10 mM DTT. Inhibition occurred when the cells were treated at 37°C but not at 4° and was reversible metabolically but not by the addition of sulfhydryl oxidizing reagents. The inhibition was overcome when DTT‐treated human fibroblasts were plated on cold insoluble globulin (plasma fibronectin)—coated dishes. Under these conditions spreading appeared to be completely normal, including the formation of focal adhesions. Analysis of the fibronectin concentrations in the human fibroblasts following DTT treatment indicated that there was little decrease in the absolute level of activity as determined in a biological assay for BHK cell spreading on culture dishes. Analysis of the fibronectin distribution on the DTT‐treated human fibroblasts by indirect immunofluorescence using a specific anti‐CIG antiserum revealed that fibronectin was no longer deposited onto the culture dish surfaces. Even when the DTT‐treated human fibroblasts spread in the presence of fetal calf serum, the cell fibronectin remained for the most part in a perinuclear location. These results indicate that DTT treatment of human fibroblasts prevents the normal translocation of fibronectin from a perinuclear location to the surface of the culture dish. This study further supports our hypothesis that the initial spreading in serum‐free medium of fibroblasts from cell strains depends upon secretion of fibronectin onto the culture dish surface.
Regulation of prostaglandin synthesis during differentiation of cultured mouse myeloid leukemia cells
Authors: Honma Yoshio , Kasukabe Takashi , Hozumi Motoo , Koshihara Yasuko 
Published: February 2005

AbstractMouse myeloid leukemia cells (Ml) were induced to differentiate into mature macrophages and granulocytes by various inducers. The differentiated Ml cells synthesized and released prostaglandins, whereas untreated Ml cells did not. When the cells were prelabelled with [14C]arachidonate, the major prostaglandins released into the culture media were found to be prostaglandin E2, D2, and F2α in an early stage of differentiation, but the mature cells produced predominantly prostaglandin E2. The synthesis and release of prostaglandins were completely inhibited by indomethacin. Dexamethasone, a potent inducer of differentiation of Ml cells, did not induce production of prostaglandins in resistant Ml cells that could not differentiate even with a high concentration of dexamethasone. These results suggest that production of prostaglandins in Ml cells is closely associated with differentiation of the cells. Homogenates of dexamethasone‐treated Ml cells converted arachidonate to prostaglandins, but this conversion was scarcely observed with homogenates of untreated Ml cells. Dexamethasone and the other inducers stimulated the release of arachidonate from phospholipids. Therefore, induction of prostaglandin synthesis during differentiation of Ml cells may result from induction of prostaglandin synthesis activity and stimulation of the release of arachidonate from cellular lipids. Lysozyme activity, which is a typical biochemical marker of macrophages, was induced in Ml cells by prostaglandin E2 or D2 alone, as well as by inducers of differentiation of the cells, but it was not induced by arachidonate or prostaglandin F2α. These results suggest that prostaglandin synthesis is important in differentiation of myeloid leukemia cells.