bluuebunny/crossref_metadata_2025_split huggingface dataset:
Isolation and characterization of an erythroid cell line highly inducible to form erythroid burst‐like colonies
Authors: Kimura Nobuhiro , Mak Tak W. 
Published: February 2005

AbstractThe study of induction of Friend erythroleukemic cell lines during the last decade has enriched our understanding of late erythroid differentiation. In comparison, little information is available on early erythroid differentiation. We describe here the isolation and characterization of a highly inducible clone from a murine erythroid cell line, which is capable of forming colonies that possess properties of the early erythroid burst progenitor. We found that a combination of erythropoietin (Epo), spleen conditioned medium (SCM), and plasma from a patient with aplastic anemia (Apa) induces over 95% of cells from this clone (clone 12) to form colonies with the properties of burst or mixed burst blast‐like colonies. Examination of the culture conditions of these cells indicated that alpha medium was more efficient for colony induction than Iscove's medium, and that the addition of two‐mercaptoethanol did not improve the induction process. These factors (EPo, SCM, and Apa) must be present for 4 days in order for induction to take place. It is hoped that the isolation of this highly inducible cell clone will enrich our understanding of early erythroid differentiation.
Permeability of human blood platelets to glycerol
Authors: Armitage W. J. 
Published: February 2005

AbstractThe permeability of human platelets to glycerol was determined at 37°C, 25°C, and 0°C from the rate of change of cell volume after abrupt addition of 0.5 mol/liter glycerol in phosphate‐buffered saline. Intracellular water volume was measured employing both tritiated water and a photometric method. Intracellular glycerol was measured employing tritiated glycerol. The glycerol permeability coefficient derived from the tracer cell volume data was 4.0 ± 0.7 × 10 −7 cm/s at 37°C, and 1.1 ± 0.4 × 10−7 cm/s at 25°C, and the photometric data gave a permeability coefficient of 5.4 ± 0.4 × 10−7 cm/s at 37°C. The activation energy between 23°C and 37°C for glycerol permeation was 19.8 kcal/mol. The cells were virtually impermeable to glycerol at 0°C. The minimum intracellular water volume attained after the addition of 0.5 mol/liter glycerol at 37°C determined by the photometric method was 47.8% of normal water volume, whereas the minimum water volume calculated assuming that glycerol exerted its full osmotic effect (i.e., σ = 1) was 45.6%. The reflexion coefficient was therefore assumed to be unity. Neither method of cell volume determination could be used with 1 or 2 mol/liter glycerol: adequate separation of the cells from the labeled medium could not be achieved in the tracer method; in the photometric method, it was apparent that transmittance (660 nm) was influenced by one or more variables in addition to cell volume.
A fractionation procedure of mouse bone marrow cells yielding exclusively pluripotent stem cells and committed progenitors
Authors: Bauman J. G. J. , Wagemaker G. , Visser J. W. M. 
Published: February 2005

AbstractThe cell surface phenotype of pluripotent hemopoietic stem cells (CFU‐S) and committed progenitors (CFU‐C1, CFU‐C2, BFU‐E) of mouse bone marrow was analyzed with respect to their binding of wheat germ agglutinin (WGA) and two monoclonal antibodies, anti‐GM‐1.2 and anti‐PGP‐1. Stained cells were fractionated on the basis of differences in fluorescence and light scatter intensity using a light‐activated cell sorter. The 6% of the cells that bound most WGA and that also had a relatively high forward light scatter (FLS) and low perpendicular light scatter (PLS) contained nearly all stem cells (CFU‐S) and progenitors. Anti‐GM‐1.2 stained only mature myeloid cells, not CFU‐S or the in vitro colony‐forming cells. Anti‐PGP‐1 stained all bone marrow cells in varying intensities: lymphoid cells were dull, CFU‐S were intermediate, CFU‐C2 were brighter, and mature myeloid cells very bright. Enrichment of progenitor cells was performed by a two‐step sorting procedure. First, the 6% most WGA‐binding cells with high FLS and low PLS were sorted out. A 10–15‐fold enrichment of progenitors and CFU‐S was obtained. Next, these cells were restained with anti‐GM‐1.2 or anti‐PGP‐1 and again fractionated on the FACS. The GM‐1.2‐negative cells were then another four‐ to sevenfold more enriched for stem cells and progenitors. Of the cells in this fraction, 95% could be assigned to a colony‐forming unit. With anti‐PGP‐1, CFU‐C2 could be partly separated from more early cells such as CFU‐S and BFU‐E.
Inhibition of DNA synthesis by an inducer of differentiation of leukemic cells, 1 alpha, 25 dihydroxy vitamin D3, precedes down regulation of the c‐myc gene
Authors: Brelvi Zamir S. , Studzinski George P. 
Published: February 2005

AbstractTransient monocytic differntiation was induced in cultures of HL 60 cells by a four hour exposure to 1 alpha, 25‐dihydroxy vitamin D3 (1,25(OH)2D3). This treatment resulted in a rapid, selective, inhibition of DNA syntehsis, which was accompanied by reduced cellular levels of c‐myc mRNA, and a more gradual appearance of c‐fos mRNA. After removal of the inducer from the cultures, DNA syntehsis and c‐myc mRNA levels returned rapidly to near‐normal levels, but the expression of c‐fos gene continued to increase for 24 hr and then declined slowly. Studies with isolated nuclei showed that the inhibition of DNA synthesis can be detected earlier than the changes in transcriptional rates of the oncogenes studied, and that 1,25(OH)2D3 directly inhibits the DNA synthesis in isolated nuclei. Autoradiographic studies of [3H] thymidine incorporation showed that 1,25(OH)2D3 does not immediately block the progression of the cells into the S phase of the cell cycle, but that those cells which become differentiated as the result of a brief exposure to this inducer do have such a block. It is concluded that 1,25(OH)2D3 produces both an immediate and a delayed inhibition of DNA synthesis in HL 60 cells, that the immediate inhibition is not preceded by detectable changes in oncogene expression, and that the delayed inhibition is accompanied by an elevated expression of the c‐fos gene, and may be related to the monocytic differentiation of HL 60 cells.
Effect of alterations in the size of the vacuolar compartment on pinocytosis in J774.2 macrophages
Authors: Swanson Joel , Yirinec Brian , Burke Eugene , Bushnell Anne , Silverstein Samuel C. 
Published: February 2005

AbstractJ774.2 macrophages cultured in medium containing 10 mg/ml sucrose accumulate the sugar by pinocytosis and become highly vacuolated, due to the sugar's osmotic effect within the vacuolar compartment. When such cells are incubated in medium containing 0.5 mg/ml invertase, the enzyme reaches the sucrose vacuoles by pinocytosis, then cleaves the sugar to more permeant monosaccharides. Within 4 hours, the vacuoles shrink to smaller, phase‐dense organelles (Cohn and Ehrenreich, 1969, J. Exp. Med., 129:201). We have used this reversible expansion of the lysosomal compartment to address two questions: (1) Does the increased size of the lysosomal compartment affect pinocytic accumulation of solute, and (2) what is the fate of the vacuolar membrane and its soluble content during invertase‐induced vacuole shrinkage? Using lucifer yellow (LY) as a probe for pinocytic fluid influx and efflux, we found that vacuolated cells accumulated 30–50% less LY than controls and returned to higher rates of pinocytosis after invertase‐induced vacuole shrinkage. A similar reduction in LY accumulation was achieved after feeding cells latex beads to increase the size of the lysosomal compartment. Thus, treatments that increased the size of the lysosomal compartment reduced solute accumulation via pinocytosis. A dramatic shrinkage of LY‐containing sucrose vacuoles followed pinocytosis of invertase. Despite this reduction in size of the LY‐containing vacuoles, the overall rate of LY efflux did not increase significantly during invertase‐induced vacuole collapse. Electron microscopy revealed that during shrinkage, the excess vacuolar membrane was compressed into whorled membranous organelles (residual bodies), with fluid markers (colloidal gold and, by inference, LY) trapped inside. The trapping of LY inside lysosomes as J774.2 macrophages returned to their normal dimensions indicates that nearly all of the surplus membrane contents were removed from circulation as well.
Granulocyte‐macrophage colony formation in serum‐free culture: Effects of purified colony‐stimulating factors and modulation by hydrocortisone
Authors: Eliason James F. 
Published: February 2005

AbstractThe effects of three purified colony‐stimulating factors (CSFs) with different specificities for the granulocyte (G) and macrophage (M) lineages (G‐CSF, CSF‐1 and GM‐CSF) were studied in a serum‐free clonal assay system. The results were compared with those obtained in similar cultures containing fetal calf serum (FCS). Total clone (≥ 10 cells) and colony (≥ 50 cells) numbers were enhanced by FCS under most conditions. However, the extent of enhancement was highly dependent on the concentration and type of CSF. In some instances, FCS also altered the proportions of G, M, and mixed GM clones induced by the CSFs. In cultures stimulated with GM‐CSF, enhancement by FCS was significant only at low CSF concentrations, primarily due to increased numbers of M clones. In contrast, clonal growth was increased by FCS only at high concentrations of CSF‐1. Clone and colony numbers induced by G‐CSF were greatly increased in cultures with FCS at all CSF concentrations tested. Virtually all clones developing in serum‐free medium with G‐CSF were pure G, whereas, M and GM clones were usually present in serum‐containing cultures with high doses of G‐CSF. The effects of hydrocortisone (HC) were also examined in these experiments. Like modulation by FCS, modulation of clonal growth by HC depended on the CSF used as stimulus, having no effect in cultures with G‐CSF, inhibitory effects with CSF‐1, and variable effects with GM‐CSF related to CSF concentration.
Parathyroid hormone stimulation of alkaline phosphatase activity in cultured neonatal mouse calvarial bone cells: Involvement of cyclic AMP and calcium
Authors: Yee John A. , Sutton Julie K. , Shew Ronald L. , Olansky Leann 
Published: February 2005

AbstractThe involvement of cAMP and calcium in the rise in alkaline phosphatase (AP) activity observed when confluent, serum‐free primary cultures of neonatal mouse calvarial cells are treated with parathyroid hormone (PTH) has been studied. Synthetic bovine PTH [bPTH‐(1–34)] increased cellular cAMP at concentrations (10−9 to 10−7 M) previously found to elevate AP activity. Other substances that increase cAMP in these cells (forskolin, prostaglandin E2, 8‐bromoadenosine cAMP and 3‐isobutyl‐1‐methylxanthine) also increased enzyme activity. By comparison, increasing the concentration of calcium in the culture medium from 1.8 to 3.8 or 5.8 mM lowered the magnitude of the maximal AP response. In addition, treatment of cultures with the divalent cation ionophore A23187 caused a significant decrease in AP activity. These results suggest that: (1) cAMP mediates the rise in the specific activity of AP in cultured neonatal mouse calvarial cells treated with bPTH‐(1–34) and (2) the concentration of calcium in the environment significantly influences the responsivity of bone cells to the hormone.
Protein phosphorylation in cultured endothelial cells
Authors: Mackie Ken , Lai Yvonne , Nairn Angus C. , Greengard Paul , Pitt Bruce R. , Lazo John S. 
Published: February 2005

AbstractWe have investigated the protein phosphorylation systems present in cultured bovine aortic and pulmonary artery endothelial cells. The cells contain cyclic AMP‐dependent protein kinase, three calcium/calmodulin‐dependent protein kinases, protein kinase C, and at least one tyrosine kinase. No cyclic GMP‐dependent protein kinase activity was found. The cells also contained numerous substrates for cyclic AMP‐dependent protein kinase and protein kinase C. Fewer substrates were found for the calcium/calmodulin‐dependent protein kinases. There was little difference between either protein kinase activities or substrates when pulmonary artery endothelium was compared to aortic endothelium grown under similar culture conditions. It is likely that these various protein kinases and their respective substrate proteins are involved in mediating several of the actions of the hormones and drugs which affect the vascular endothelium.
Density‐dependent inhibition of expression of syncytiotrophoblastic markers by cultured human choriocarcinoma (BeWo) cells
Authors: Burres Neal S. , Cass Carol E. 
Published: February 2005

AbstractIn the presence of methotrexate, cultured human choriocarcinoma (BeWo) cells undergo a differentiative response that resembles normal trophoblastic development. In the current study, the effects of cell number and population density on drug‐induced conversion of BeWo cells from the cytotrophoblast‐like to the syncytiotrophoblastlike phenotype were investigated using as markers of differentiation formation of “giant” cells, a process shown to require exogenous purines, and expression of placental (heat‐stable) alkaline phosphatase. Giant cell formation, assessed by determination of cell volumes, was reduced in crowded cultures, and addition of hypoxanthine to growth media partially restored methotrexate‐induced cell enlargement. Cellular uptake of methotrexate, assessed by following the loss of methotrexate from cell culture fluids during drug exposures, was two‐threefold greater in sparsely populated than in densely populated cultures. Although the concentration of methotrexate in culture fluids of crowded cultures declined during exposures of 48 hr, the amount of extracellular drug remaining at 48 hr was well above the threshold for induction of the differentiative response. When culture population was held constant and population density was manipulated by varying the substratum available to cells, methotrexate‐induced cell enlargement was inversely related to population density. Expression of placental alkaline phosphatase, salvage of exogenous hypoxanthine, and synthesis of RNA were also reduced at high population densities. These results indicate that expression of markers of methotrexate‐induced differentiation of BeWo cells was inhibited in a density‐dependent manner that may have been related to reduced cellular uptake of the inducing agent and of exogenous nutrients (purines) from culture fluids.
Prominent glutamine oxidation activity in mitochondria of avian transplantable hepatoma induced by MC‐29 virus
Authors: Matsuno Tetsuya , Satoh Teikichi , Suzuki Haruo 
Published: February 2005

AbstractWell coupled mitochondria were isolated from transplantable chicken hepatoma induced by MC‐29 virus. The mitochondrial phosphate‐dependent and phosphate‐independent glutaminase activities were increased compared with those from normal chicken liver. Glutamate dehydrogenase was undetectable in the tumor mitochondria. Oxypolarographic tests showed the following: (1) glutamine oxidation was prominent in the tumor mitochondria and was mediated through an NAD‐linked reaction, while mitochondria from the liver showed a feeble glutamine oxidation; (2) glutamine oxidation by tumor mitochondria was inhibited either by aminooxyacetate, inhibitor of transaminases, or prior incubation of mitochondria with DON (6‐diazo‐5‐oxonorleucine), which inhibited mitochondrial glutaminases. Bromofuroate, inhibitor of glutamate dehydrogenase, had little or no effect; and (3) glutamate oxidation was also inhibited by aminooxyacetate, while it was not affected by DON. These findings clearly show a high glutamate oxidation activity in the hepatoma and indicate that the product of glutamine hydrolysis, glutamate, is catabolized via transamination in the mitochondria to supply ATP.
Down‐modulation of receptors for granulocyte colony‐stimulating factor on human neutrophils by granulocyte‐activating agents
Authors: Nicola N. A. , Vadas M. A. , Lopez A. F. 
Published: February 2005

AbstractPurified human blood neutrophils were able to bind radioiodinated murine granulocyte‐colony‐stimulating factor (G‐CSF) in a specific manner. This factor has previously been shown to stimulate functional activities of human and murine neutrophilic granulocytes and to be functionally analogous to human‐derived CSFβ. The binding of 125I G‐CSF to human neutrophils was competed for equally by unlabeled G‐CSF and CSFβ but not by other CSF's. Saturation analysis indicated that human neutrophils displayed about 700–1,500 receptors for G‐CSF/CSFβ per cell. Three other agents (N‐formyl‐methionine‐leucine phenylalanine, bacterial lipopolysaccharide, and human CSFα) known to activate neutrophils did not compete directly for G‐CSF binding sites but, in preincubation experiments at 37°C, were able to down‐modulate the expression of G‐CSF receptors on human neutrophils in a dose‐ and time‐dependent manner. This effect was specific since the same agents have been shown elsewhere to up‐regulate the expression of other granulocyte surface antigens and other agents were much less effective at down‐modulating G‐CSF receptors. Since the granulocyte‐activating agents increase the sensitivity of human neutrophils to G‐CSF/CSFβ and mimic some of the actions of G‐CSF on neutrophils, it is suggested that G‐CSF receptor down‐modulation might be a mechanism whereby these agents activate G‐CSF receptors and thereby exert some of their effects.
Effects of transformation on the expression of laminin and fibronectin by neural cells
Authors: Giotta Gregory J. , Newby Ron , Oliver Noelynn 
Published: February 2005

AbstractWe have studied laminin and fibronectin expression in a collection of rat cerebellar neural cell lines transformed with a mutant of Rous sarcoma virus which is temperature sensitive for transformation. We show that regardless of their neuronal or glial properties the cell lines produce both laminin and fibronectin. Laminin is expressed in similar amounts in cell lines grown at either the permissive or nonpermissive temperature for transformation, while fibronectin is generally expressed at higher levels in cells kept at the nonpermissive temperature. To provide further evidence that neural cells can produce laminin and fibronectin, double label immunofluorescence studies were conducted on primary cerebellar cultures. Both laminin and fibronectin were found to be present in the primary culture, and laminin was found to be associated with a subpopulation of astrocytes.
Reversible alterations in cultured pulmonary artery endothelial cell monolayer morphology and albumin permeability induced by ionizing radiation
Authors: Friedman Mitchell , Ryan Una S. , Davenport W. Clark , Chaney Edward L. , Strickland Debra L. , Kwock Lester 
Published: February 2005

AbstractThe effects of ionizing irradiation (0, 600, 1,500, or 3,000 rads) on the permeability of pulmonary endothelial monolayers to albumin were studied. Pulmonary endothelial cells were grown to confluence on gelatin‐coated polycarbonate filters, placed in serum‐free medium, and exposed to a 60Co source. The monolayers were placed in modified flux chambers 24 hours after irradiation; 125l‐albumin was added to the upper well, and both the upper and lower wells were serially sampled over 4 hours. The amount of albumin transferred from the upper well/hour over the period of steady‐state clearance (90–240 min after addition of 125l‐albumin) was 2.8 ± 0.2% in control monolayers and was increased in monolayers exposed to 1,500 or 3,000 rads (increase of 63 plusmn; 10% and 61 plusmn; 10%, respectively, P<0.01). No increase was found in monolayers exposed to 600 rads. The increases in endothelial albumin transfer rates were associated with morphologic evidence of monolayer disruption and endothelial injury which paralleled the changes in albumin permeability. Dose‐dependent alterations in endothelial actin filament organization were also found. Incubation of the monolayers exposed to 3,000 rads with medium supplemented with 10% fetal calf serum for 24 hours resulted in normalization of albumin permeability, improvement in morphologic appearance of the monolayers, and reorganization of the actin filament structure. These studies demonstrate that ionizing radiation is an active principle in the reversible disorganization of cultured pulmonary endothelial cell monolayers without the need of other cell types or serum components.
Regulation of gonadotropin receptors, gonadotropin responsiveness, and cell multiplication by somatomedin‐C and insulin in cultured pig Leydig cells
Authors: Bernier M. , Chatelain P. , Mather J. P. , Saez J. M. 
Published: February 2005

AbstractWe have investigated the effects of insulin and somatomedin‐C/insulinlike growth factor l(Sm‐C) in purified porcine Leydig cells in vitro on gonadotrophins (hCG) receptor number, hCG responsiveness (cAMP and testosterone production), and thymidine incorporation into DNA. Leydig cells cultured in a serum‐free medium containing transferrin, vitamin E, and insulin (5 μ/ml) maintained fairly constant both hCG receptors and hCG responsiveness. When they were cultured for 3 days in the same medium without insulin, there was a dramatic decline (more than 80%) in both hCG receptor number and hCG responsiveness. However the cAMP but not the testosterone response to forskolin was normal. Both insulin and Sm‐C at nanomolar concentrations prevent the decline of both hCG receptors and hCG‐induced cAMP production. This effect of both peptides was dose dependent with an ED50 of about 1 ng/ml and 5 ng/ml for SM‐C and insulin, respectively. Insulin and Sm‐C had no additive effect on these parameters. At nanomolar concentrations, Sm‐C and insulin enhanced hCG‐induced testosterone production but the effect of Sm‐C was significantly higher than that of insulin. However, the effect of insulin at higher concentrations (5 μg/ml) was significantly higher than that of Sm‐C at 50 ng/ml. In contrast, at nanomolar concentrations only Sm‐C stimulated [3H]‐thymidine incorporation into DNA and cell multiplication, the stimulatory effect of insulin on these parameters, was seen only at micromolar concentrations. These results indicate that both Sm‐C and insulin acting through their own receptors increase Leydig cell steroidogenic responsiveness to hCG by increasing hCG receptor number and improving some step beyond cAMP formation. ln contrast, the mitogenic effects of insulin are mediated only through Sm‐C receptors.
Caffeine contracture in the cultured chick myotube
Authors: Saito Koji , Ozawa Eijiro 
Published: February 2005

AbstractA possible function of Ca store site in cultured chick myotubes was examined by recording contraction of the myotube with special reference to the effect of caffeine. Caffeine at low concentrations (below 1 mM), applied focally on the myotube through a micropipette with a pressure pulse, elicited focal contraction without membrane potential changes. Procaine inhibited the caffeine contracture. Deuterium oxide also inhibited the caffeine contracture at low concentrations, but enhanced the maximal contracture. These observations are Similar to those in the mature frog muscle fiber in which the sarcoplasmic reticulum (SR) is a main site of caffeine action. On the basis of these similarities, it was considered that caffeine acts on SR to elicit contracture in the myotube. The ability of SR to accumulate and release Ca ion seemed to be low, because caffeine contracture decreased or disappeared in a Ca‐free solution in many myotubes.