bluuebunny/crossref_metadata_2025_split huggingface dataset:
Further studies of hormone‐sensitive sodium and potassium transport in red cells from developing chick embryos
Authors: Shanbaky Nabila M. , Wacholtz Mary C. , Sha'Afi Ramadan I. 
Published: February 2005

AbstractSodium and potassium transport in the definitive series of chick embryo red cells changes significantly, both qualitatively and quantitatively, during maturation. Sodium efflux and potassium influx consist of three parts: a ouabain‐sensitive, a furosemide‐sensitive, and a ouabain‐furosemide‐insensitive component. In chick red cells of most ages, the ouabain‐sensitive and furosemide‐sensitive parts of the cation fluxes do not overlap. Cation transport in the more mature red cells is increased significantly by epinephrine, whereas cation transport in red cells from younger embryos is stimulated much less. This is a β‐adrenergic effect of epinephrine and is mediated by cyclic AMP. The relative lack of response in younger embryos is not due to the absence of β‐adrenergic receptor or the lack of production of cyclic AMP. Ouabain has no effect on the hormone‐sensitive sodium or potassium transport. On the other hand, furosemide nearly completely abolishes the effect of epinephrine. In addition, there is a good correlation between furosemide‐sensitive components of both sodium and potassium transport and the epinephrine‐sensitive component. Furosemide has no effect on cyclic AMP levels in the presence or absence of epinephrine. This suggests that furosemide may act directly on the cation transport system. In the red cells from younger embryos, furosemide‐sensitive units are present but cannot be fully activated by epinephrine. Therefore, the lack of the hormone effect on cation movements in these cells is consistent with the view that the appropriate units are present, but do not respond fully to intracellular cyclic AMP levels.
Use of an encapsulated fluorescent probe to measure intracellular PO2
Authors: Podgorski Gary T. , Longmuir Ian S. , Knopp James A. , Benson Douglas M. 
Published: February 2005

AbstractThe objectives of this investigation were to produce a reliable, senstive probe to measure intracellular PO2 with a high degree of resolution and to apply this technique to biological systems. A fluorescent molecule, pyrene dissolved in paraffin oil, was encapsulated in polyacrylamide to form a probe of nanometer dimensions. The quantitative and microscopic oxygen values were determined by analyzing the quenching of the fluorescence of the probe by oxygen, as displayed on a television monitor by a silicon‐intensified‐target camera. The nanocapsules had a sensitivity of approximately 1 mm PO2, a spatial resolution of 0.5 μm, and a temporal resolution of milliseconds. Calibrated nanocapsules within nonrespiring Amoeba proteus responded to ambient partial pressures of oxygen. At two different ambient partial pressures, nanocapsules engulfed by respiring amoebas indicated an intracellular PO2 28 mm Hg less than extracellular PO2. The capsules retained their sensitivity to oxygen for at least 8 months.
The effect of phorbol esters on the proliferation of C3H‐10T1/2 mouse fibroblasts: Consideration of both stimulatory and inhibitory effects
Authors: Tomei L. David , Cheney John C. , Wenner Charles E. 
Published: February 2005

AbstractTPA stimulates cell cycle activation in both serum‐deprived and density‐inhibited cultures. The cells reestablish cycle arrest after no more than one generation, and addition of fresh drug produces no further response. However, cells freshly trypsinized can respond with a series of repetitive generations resulting in 3.5–4.0 population doublings over 72 hrs. In kinetic pulse experiments TPA enhanced 3H‐thymidine incorporation in densityinhibited cells stimulated by fresh serum but only after markedly suppressing incorporation 8–13 hrs after serum stimulation. When cells arrested by serum deprivation were pretreated with TPA, fresh serum stimulation led to initiation of 3H‐TdR incorporation 5 hrs earlier than untreated controls. However, TPA addition at the time of serum stimulation did not lead to a suppression at 8–13 hrs, whereas enhancement was observed during peak incorporation times regardless of whether the cells were pretreated with TPA during serum deprivation. The results support the concept that there can exist within G1 multiple states of responsiveness to phorbol esters. These pharmacologically induced states may be correlated with corresponding physiological states of the G1 phase of cell cycle.
Induction of polyamine limitation in Chinese hamster ovary cells by α‐methylornithine
Authors: Harada John J. , Porter Carl W. , Morris David R. 
Published: February 2005

AbstractChinese hamster ovary (CHO) cells in culture were limited for polyamines through the use of α‐methylornithine (αMO), a competitive inhibitor of ornithine decarboxylase. Initial exposure of the cells to the inhibitor caused growth rate and intracellular polyamine content to decline continuously. Reseeding the αMO‐treated cells into medium containing the inhibitor resulted in steady‐state (exponential) growth at cell densities below 5 × 103 cells/cm2, at a rate approximately twofold slower than untreated cells. Under these conditions, putrescine and spermidine were undetectable and spermine remained relatively constant at a level approximately half that found in untreated cells. Addition of exogenous putrescine elevated the polyamine content and stimulated the growth of αMO‐treated cultures. Thus, growth rate correlated with polyamine content in the αMO‐treated cells.The growth of reseeded. αMO‐treated cells became nonexponential at a density (5 × 103 cells/cm2) far below that at which untreated cells departed from exponential growth (1 × 105 cells/cm2). Medium obtained from high density, αMO‐treated cultures inhibited the growth of cells at low density in the presence of αMO. Doubling the concentration of the defined components of conditioned medium did not markedly affect its capacity to inhibit growth. However, dialysis completely removed the inhibitory activity from conditioned medium. The results imply that a low molecular weight inhibitor of growth is produced by polyamine‐limited cells. This is a variable that must be controlled in studies with polyamine‐limited animal cells.Morphological studies indicated that subcellular organelles, including mitochondria, were largely unaffected by treatment with αMO. The maintenance of mitochondrial integrity in the presence of αMO demonstrates that the swelling of mitochondria observed previously in cells treated with methylglyoxal bis(guanylhydrazone) was not due to polyamine limitation. αMO‐treated cells did, however, accumulate numerous cytoplasmic vacuoles. The identity of these vacuoles and their relationship to cellular physiology is not yet understood.
Suppression of the adipose conversion of 3T3 cells by acidified serum
Authors: Kuri‐Harcuch Walid , Green Howard 
Published: February 2005

AbstractThe adipose conversion of cultured 3T3‐F442A cells is strongly inhibited if the fetal bovine serum of the culture medium is briefly acidified before it is used. The inhibitory factor is a polypeptide with an apparent molecular weight of 24,000, and is inactivated by pronase or trypsin. Cells grown to confluence in the presence of this factor do not become spherical or accumulate triglyceride; they also do not increase the activity of their triglyceride‐synthesizing enzymes. The factor suppresses adipose conversion even in the presence of untreated serum. Once adipose conversion has begun in the absence of the inhibitory factor, subsequent addition of the factor does not arrest the conversion.
Coordinate regulation of gluconeogenesis by the glucocorticoids and glucagon: Evidence for acute and chronic regulation by glucagon
Authors: Kletzien Rolf F. , Weber Charles A. , Stumpo Deborah J. 
Published: February 2005

AbstractThe coordinate regulation of gluconeogenesis by the glucocorticoids and glucagon in primary cultures of adult rat liver parenchymal cells has been studied. The results suggest that glucagon stimulation of glucose production from 3‐carbon precursors is composed of at least two components which the glucocorticoids differentially affect. Glucagon treatment of hepatocytes results in an immediate increase in glucose production which is not blocked by cycloheximide and occurs in the absence of any detectable increase of phosphoenolpyruvate carboxykinase activity. This component appears to be regulated by a post‐translational mechanism and involves redirection of carbon flow from glycolysis to gluconeogenesis. The second component is characterized by the need for long‐term glucagon treatment. This increase in glucose production can be blocked by cycloheximide and is correlated with an increase in phosphoenolpyruvate carbooxykinase activity. The reaction that is accelerated by long‐term glucagon incubation is located prior to the triose‐phosphate level since long‐term incubation with glucagon fails to increase glucose production from dihydroxyacetone any more than does short‐term incubation. It is suggested that phosphoenolpyruvate carboxykinase rather than amino acid transport is the key pacemaker reaction in the long‐term incubation since the direction and magnitude of the response for glucocorticoid and glucagon stimulation of glucose production is the same whether alanine or lactate is used as the 3‐carbon precursor. The glucocorticoids exhibit an additive effect on glucagon‐stimulated glucose production for the first component whereas they amplify the second component.
Failure of hydrocortisone or growth factors to influence the senescence of fibroblasts in a new culture system for assessing replicative lifespan
Authors: Didinsky James B. , Rheinwald James G. 
Published: February 2005

AbstractIt has been reported that the replicative lifespan of human fibroblasts can be substantially extended by supplementing the growth medium with hydrocortisone or increased levels of serum proteins. These observations have been made only on cell populations transferred many times at high cell density, and cumulative population doublings have been recorded, rather than a more direct measure of cell division potential. We have measured the replicative potential of human fibroblasts cultured so as to avoid conditions of high cell density, medium depletion, and departure from exponential growth. Two fetal lung and two newborn foreskin fibroblast strains were serially passaged in the presence or absence of hydrocortisone (HC), epidermal growth factor (EGF), and fibroblast growth factor (FGF) until they senesced. At each passage cells were plated at densities sufficiently low that colony‐forming efficiency could be calculated. We determined cumulative population doublings and also estimated the number of cell generations attained under each condition. FGF caused small but possibly significant changes, while HC and EGF failed to substantially alter replicative lifespan. The reported effect of HC on the doubling potential of fetal lung fibroblasts is therefore not an inevitable action of this hormone on the senescence mechanism, but may instead depend for its apparent activity on the passage regimen used. The fibroblast's insensitivity to EGF as a modulator of replicative potential, as compared with the keratinocyte, whose lifespan can be tripled by EGF, implies that the mechanisms limiting the replicative potential of these two cell types are not identical.
Murine embryonal carcinoma hybrids: Decreased ability to spontaneously differentiate as a dominant trait
Authors: Oshima Robert G. , McKerrow James , Cox David 
Published: February 2005

AbstractThe F9 murine embryonal carcinoma (ec) cell line does not differentiate spontaneously at high frequency either in vitro or in vivo. A derivative of this variant ec cell line resistant to ouabain and thioguanine was fused with the PSA1 ec cell line, which retains the ability to differentiate to a variety of cell types, either in vitro or in vivo. Hybrids were selected in media containing ouabain, hypoxanthine, aminopterin, and thymidine at a frequency of approximately 10−3. Authentic hybrids were chosen by screening for the expected hybrid DNA content by flow microfluorometry and the lack of parental cell contamination in appropriate selective media. After injection into strain 129 mice, five of eight independently isolated hybrids formed tumors composed exclusively of embryonal carcinoma cells. The remaining three hybrids of this cross caused tumors composed of greater than 95% embryonal carcinoma cells but with small foci of differentiated tissues as well. Like the F9‐derived parental line, less than 10 percent of the clones, formed by the hybrids after six days of growth, contained cells that secreted plasminogen activator. Since control hybrids derived from the fusion of either two F9 derivatives or two pluripotent cell lines resembled their parental cell lines with regard to the types of tumors formed and their differentiation in vitro, it appears that the F9 phenotype is dominant to the pluripotent phenotype in early passage embryonal carcinoma hybrids.
The serum‐free growth of cultured cells in bovine colostrum and in milk obtained later in the lactation period
Authors: Steimer Kathelyn Sue , Packard Robert , Holden David , Klagsbrun Michael 
Published: February 2005

AbstractSeven established cell lines, including both epithelial cells and fibroblasts (MDCK, Vero, CV‐1, NRK, 3T3, F2408, and NIL8) and four early passage cell strains (bovine articular chondrocytes, bovine smooth muscle cells, human foreskin fibroblasts, and rat embryo cells) were cultured in serum‐free medium supplemented with milk obtained 1 day after birth (colostrum) or 80 days after birth (older milk). MDCK, Vero, CV‐1, NRK, and 3T3 grew readily in colostrum and attained saturation densities ranging from 22% to 63% of that in serum. There was no growth of F2408, NIL8, or the early passage strains in bovine colostrum. None of the 11 cell cultures grew in older milk. The temporal dependence of growth in milk was examined in detail using MDCK cells. Growth equivalent to that in serum occurred in 3% colostrum and in 15% milk obtained 2 days after birth. Milk obtained 3 days and 10 days after birth was not effective as a growth supplement for MDCK cells at any concentration. Those cells, unable to grow in colostrum or in older milk, could be induced to grow if culture dishes were precoated with fibronectin. In addition to fibronectin, it was necessary in some cultures to supplement colostrum or older milk with insulin and/or transferrin in order to achieve growth. In the presence of fibronectin and appropriate factors, the final saturation density attained in colostrum or older milk ranged from 25% to 100% of that in serum. The fibronectin contents of bovine colostrum and milk were determined. The fibronectin level of colostrum was found to be approximately 5% of bovine serum. There was no detectable fibronectin in the 80‐day‐old milk.
Biochemical properties of differentiation factors for murine myelomonocytic leukemic cells in organ conditioned media — separation from colony‐stimulating factors
Authors: Nicola Nicos A. , Metcalf Donald 
Published: February 2005

AbstractA number of different organs and tissues from untreated or endotoxin‐injected mice were surveyed for their ability to produce conditioned medium that could induce differentiation of the myelomonocytic leukemic cell line WEHI‐3B. All organs were able to do this but were much more effective from endotoxin‐injected animals. Production of the differentiation factor (DF) was dependent on protein synthesis and occurred throughout the 7‐day incubation period. DF produced by the different organs in vitro and found in vivo in endotoxin serum were indistinguishable from each other by several fractionation procedures but could be distinguished from the majority of granulocyte‐macrophage and macrophage colony‐stimulating factors (GM‐ and M‐CSFs). DF was precipitated by 55% ammonium sulfate whereas CSF required 85% saturation. DF did not bind to Concanavalin A‐Sepharose whereas CSF did. DF eluted later than CSF from Phenyl‐Sepharose columns. The dissociated molecular weight of DF was 21,000‐29,000 and was not greatly affected by neuraminidase treatment. There was no evidence that DF was glycoprotein in nature although this cannot yet be ruled out. Although the DF activity could be clearly separated from the bulk of GM‐CSF and M‐CSF activities, there was always residual CSF activity associated with the DF. This CSF was distinct from GM‐ and M‐CSF in its biochemical properties, in the fact that it only stimulated a subset of colony‐forming cells and that upon dilution it stimulated exclusively granulocytic colonies to develop. In two steps (ammonium sulfate precipitation and Phenyl‐Sepharose chromatography) it was possible to selectively enrich the differentiation activity over the CSF activity by 21‐fold.
Cellular mutations and drug resistance probed by herpes simplex virus
Authors: Campisi Judith , Pardee Arthur B. 
Published: February 2005

AbstractThe growth of herpes simplex virus type 1 (HSV‐1), monitored by plaque assay, was inhibited by the cellular antimetabolites thioguanine (TG), cytosine arabinoside (AraC), methotrexate (MTX), and 5‐fluorodeoxyuridine (5‐FUdR). These results suggested a rapid means for assaying cellular drug sensitivity, based on the ability of infected cells to support viral replication. We have explored the feasibility of a virus‐mediated assay for cellular metabolic function in two model systems.Using an immunofluorescence assay to assess viral growth, we found that all of the antimetabolites tested were effective in diminishing HSV‐1 specific fluorescence in human fibroblasts. However, a DNA‐damaging agent, bleomycin, was lethal to cells but was completely ineffective in reducing viral fluorescence. HSV‐1 growth was markedly decreased by TG in a normal human fibroblast strain, FS‐2. In contrast, a Lesch‐Nyhan strain (LNF), resistant to TG owing to its genetic defect, showed no suppression of viral growth in the presence of TG. The drug's effect on viral fluorescence closely paralleled its effect on cellular colony forming ability and rate of cellular DNA synthesis.Thymidine kinase‐deficient HSV‐1 (TK−HSV‐1) did not grow in a normal mouse fibroblast line (A31) in the presence of 5‐FUdR. However, a TK− derivative of the A31 line allowed full production of the TK−HSV‐1 antigens at low to moderate doses of 5‐FUdR.Two potential applications for this assay are the prenatal diagnosis of some genetic disorders and the rapid detection of drug resistant populations in tumor specimens. Toward these ends, we demonstrated that human fibroblasts from patients with the hereditary disorder Xeroderma pigmentosum (group A) were easily distinguished from normal human fibroblasts by their inability to support the growth of UV‐irradiated HSV‐1. We also investigated the effects of TG upon HSV‐1 fluorescence in two human tumor cell lines isolated from head and neck squamous cell carcinomas (SCC‐15) and (SCC‐25). Whereas TG was effective in reducing viral fluorescence in SCC‐15 cells, it was only marginally so in SCC‐25 cells. These latter cells showed the greater resistance to TG by growth and isotope incorporation experiments.
Effect of H+ on the kinetics of Na+‐dependent amino acid transport in ehrlich ascites tumor cells: Evidence for H+ as an alternative substrate
Authors: Smith Thomas C. , Robinson Susan C. 
Published: February 2005

AbstractThe effects of H+ on the kinetics of α‐aminoisobutyric acid (AIB) influx in Ehrlich ascites tumor cells have been investigated at different external Na+ concentrations. Elevation of [H+] in the presence of both high (154 mEq/l) and low (10 mEq/l) external Na+ leads to decreases in the maximum influx (J) and increases in the apparent Michaleis‐Menten constant (K) for influx of AIB. In the virtual absence of external Na+ (0.96 ± 0.04 mEq/l), alterations in [H+] are without measurable effect on AIB flux. Furthermore, addition of AIB (10 mM) to cell suspensions (pH 5.90) stimulates H+ uptake by the cells in either the presence or absence of Na+. The data are consistent with two kinetic models for Na+‐dependent amino acid transport: an order bireactant (Na+‐binding necessary before AIB binding) system or a random bireactant system. Both models require that H+ serve as an alternative substrate for Na+. The consistency of the models was tested by fit to data from the present study (not used to evaluate the kinetic parameters) and by prediction of the pH dependence of Na+‐dependent amino acid transport compared to earlier studies.
Decreased epidermal growth factor binding in cells growth arrested in G1 by nutrient deficiency
Authors: Robinson Robert A. , Volkenant Mary E. , Ryan Robert J. , Moses Harold L. 
Published: February 2005

AbstractPrevious studies have shown that the nontransformed AKR‐2B mouse embryo derived cell line may growth arrest by two separate mechanisms in the G1 phase of the cell cycle‐growth factor deficiency arrest (G0) and low molecular weight nutrient deficiency arrest. An examination of epidermal growth factor (EGF) receptors under the different resting or growth conditions has shown that rapidly growing cells or cells arrested due to growth factor deficiency have the expected amount of 125I‐EGF binding with approximately 105 receptors per cell being present in G0 arrested cells. In contrast, cells arrested due to nutrient deficiency show a reduction in 125I‐EGF binding to 10‐‐20% of that observed under the other conditions. This effect appears to be due to decreased receptor number and not to a change in the affinity of the receptor. Stimulation of DNA synthesis by nutrient replenishment causes a tenfold increase in EGF binding 20 hours later, with some increase in binding being detectable as early as six hours. The increase in binding is inhibited by cycloheximide and actinomycin D. This suggests that new mRNA synthesis as well as increased protein synthesis is required for the increase in EGF binding.
Membrane transport properties differ following return of serum‐deprived versus Ca++‐deprived human fibroblasts to a proliferative state
Authors: Tupper Joseph T. , Ryals William T. , Bodine Peter V. 
Published: February 2005

AbstractHuman lung fibroblasts (W138) can be brought to a quiescent state by removal of serum from the medium or by lowering of the extracellular Ca++. Upon return of Ca++ or serum, the cells enter the G1 phase and progress to S within 15–18 hours. Since multiple G1 phase blocks have been demonstrated, we wished to determine whether the Ca++ and serum block were equivalent since previous data suggested that these two medium components may act at a common point in the initiation of proliferation. We have evaluated the membrane transport of 86Rb, 3‐O‐methylglucose, AIB, and cycloleucine following stimulation of quiescent cells by Ca++ or serum. Serum stimulation results in large increases in the influx of all the substances tested. These increases are prevented if Ca++ is absent upon serum stimulation or they are rapidly diminished following Ca++ removal. In contrast, Ca++ stimulation of Ca++‐deprived cells causes little or no enhancement of any of the transport systems, yet the cells progress to S phase in a manner similar to serum‐stimulated cells. These results indicate that the Ca++ and serum G0 or G1 block are not equivalent and that the serum‐induced change in transport of these components does not appear necessary for successful G1 phase progression. Furthermore, the data suggest that the sequence in which Ca++ or serum are presented to the cells alters the ability of Ca++ to modulate the transport systems. Quiescent cells which are exposed to Ca++ prior to serum possess a Ca++ modulation of several transport systems. Cells which are exposed to Ca++ subsequent to serum do not appear to possess this Ca++ regulation.