bluuebunny/crossref_metadata_2025_split huggingface dataset:
Cytosolic calcium, calcium fluxes, and regulation of alveolar macrophage superoxide anion production
Authors: Stickle Douglas F. , Daniele Ronald P. , Holian Andrij 
Published: February 2005

AbstractThe recently available compound quin‐2, which acts as a high affinity fluorescent indicator for calcium in the cytosol, was used to examine the role of calcium mobilization in the alveolar macrophage during the stimulation of 0 production by the tripeptide N‐formyl norleucyl leucyl phenylalanine (FNLLP). After preloading with quin‐2, the production of 0 was measured in conjunction with the transfer of 45Ca+2 and changes in quin‐2 fluorescence upon stimultion with FNLLP. When cells were maintained in low (10 μM) extracellular calcium medium the presence of 1.5 mM quin‐2 in the cytosolic space partially inhibited the rate of 0 production upon stimulation by FNLLP. Addition of 1 mM Ca+2 to the medium prior to stimulation rapidly restored the cell's capability to produce 0 upon stimulation at rates equal to control and extended the duration of stimulated 0 production as well. Quin‐2 fluorescence measurements indicated an increase in cytosolic Ca+2 upon stimulation with FNLLP. This increase was lowest under conditions in which 0 production was inhibited. The addition of 1 mM Ca+2 to the medium caused by itself a rapid but transient increase in cytosolic Ca+2 as measured with quin‐2 without stimulating 0 production. This intracellularly redistributed calcium was determined to be the source of the greater increase in cytosolic calcium during stimulation in the presence of high extracellular calcium. Measurements of 45Ca+2 transfer demonstrated a buffering of cytosolic Ca+2 changes by quin‐2, which in low calcium medium could deplete calcium stores. It is suggested that this effect, prior to stimulation, was responsible for the mitigated 0 response for those cells maintained in low calcium medium, wherein calcium stores could not be replenished. These results suggested that the cell's mechanism for regulating cytosolic and bound calcium concentrations may also play an integral role in its normal mechanism for stimulated 0 production. They further support the postulate that the commonly observed rise in the concentration of calcium in the cytosol upon formyl peptide stimulation is a concomitant but nonregulatory event only.
Maturation involves suppression of voltage‐gated currents in the frog oocyte
Authors: Taglietti Vanni , Tanzi Franco , Romero Rossana , Simoncini Luciana 
Published: February 2005

AbstractVoltage‐ and time‐dependent currents having slow kinetics have been studied in plasma membranes of immature oocytes of the european frog, Rana esculenta. Ik, corresponding to an outward flow of K+, is activated at potentials more positive than about −40 mV, and subserves outward rectification; llr, corresponding to an outward flow of Cl−, is activated at potentials more negative than about −80 mV and subserves inward rectification. Such currents can act as negative feedback mechanisms in the control of membrane potential in the immature oocyte and limit to a somewhat restricted range its possible deviations from resting values. Besides lk, membrane depolarizations to potentials more positive than about + 30 mV are capable of activating lNa, corresponding to outflow of Na+. By contrast, the frog mature egg‐cell has a single voltage‐ and time‐dependent current, lM, activated at potentials more positive than +30 mV, with properties similar to lNa. The disappearance of lk and llr along with remarkable reduction in leakage lowers impedance in the egg membrane. It seems reasonable to suggest that the observed changes in membrane permeability reflect changes which have taken place along the maturation process and are of importance for successful fertilization.
The response of several murine embryonal carcinoma cell lines to stimulation of differentiation by α‐difluoromethylornithine
Authors: Schindler Joel , Kelly Michael , McCann Peter P. 
Published: February 2005

AbstractIn an attempt to better establish the relationship between polyamine levels and the differentiation of embryonal carcinoma cells, we have examined the ability of α‐difluoromethylornithine (DFMO), a known inducer of differentiation in one embryonal carcinoma cell line, to stimulate the differentiation of embryonal carcinoma cells from a variety of cell lines. Differentiation was monitored using a variety of criteria including morphological alterations and changes in biochemical and antigenic parameters. Depend ng on their response to difluoromethylornithine, three classes of cell lines could be identified, those which (1) differentiate extensively, (2) differentiate poorly, and (3) fail to differentiate. Three different classes of embryonal carcinoma cell lines reflect differential changes in polyamine levels resulting from inhibition of ornithine decarboxylase enzyme activity by DFMO. The specific cell lines which exhibit large decreases in both ornithine decarboxylase activity and polyamine levels also show extensive differentiation. The cell lines which show only moderate decreases in enzyme activity and polyamines differentiate poorly while the cell lines which fail to respond to DFMO in that polyamines do not drop below the threshold level necessary to induce differentiation fail to differentiate. These studies suggest that decreases in intracellular polyamines induce EC cell differentiation in vitro.
Reduced glutathione in chinese hamster ovary cells protects against inactivation of 3‐hydroxy‐3‐methylglutaryl Coenzyme A reductase by 2‐mercaptoethanol disulfide
Authors: Dotan Iris , Shechter Ishaiahu 
Published: February 2005

AbstractWhen the disulfide of 2‐mercaptoethanol (ESSE) is added to the medium of cultured Chinese hamster ovary (CHO) cells, a time and concentration dependent release of 2‐mercaptoethanol to the medium is observed. The reduction of ESSE to 2‐mercaptoethanol by cells is a saturable process, the rate being approximately 50 nmoles of 2‐mercaptoethanol per mg cell protein for an hour upon exposure to 250 μM ESSE. Reduction rate of ESSE by cells attached to a substratum is independent of glucose and insulin for periods up to 4 hours. However, in detached cells, swirled in suspension, addition of glucose and insulin is necessary in order to obtain a linear reduction rate of ESSE. The rate limiting enzyme in the sterol biosynthetic pathway, 3‐hydroxy‐3‐methyl‐glutaryl Coenzyme A reductase (E.C. 1.1.1.34), is inhibited by ESSE when isolated from CHO cells but total nonsaponifiable lipids synthesis from [2−14C]‐acetate in intact cells is not affected by ESSE at concentrations up to 500 μM. Cytosolic reduced glutathione can spontaneously exchange disulfide bonds with ESSE and thus prevent it from inhibiting the reductase. Cultured cells respond to ESSE administration by elevating their total and acid‐soluble glutathione levels. The use of ESSE as a perturbant of the GSH Status in cells is discussed.
Enterocytic differentiation of a subpopulation of the human colon tumor cell line HT‐29 selected for growth in sugar‐free medium and its inhibition by glucose
Authors: Zweibaum Alain , Pinto Moïse , Chevalier Guillemette , Dussaulx Elisabeth , Triadou Nicole , Lacroix Brigitte , Haffen Katy , Brun Jean‐Louis , Rousset Monique 
Published: February 2005

AbstractIn order to study the effect of glucose on the differentiation of cultured human colon cancer cells, a subpopulation of HT‐29 cells was selected for its capacity to grow in the total absence of sugar. These cells (GIc−cells) exhibit, after confluency, an enterocytic differentiation, in contrast to cells grown with glucose (Glc+ cells), which always remain undifferentiated. The differentiation is characterized by a polarization of the cell layer with apical brush borders and tight junctions, and by the presence of sucrase‐isomaltase. The differentiation of Glc−cells is reversible: the addition of glucose to postcon‐fluent cultures of Glc− cells results in an inhibiting effect on the expression of sucrase‐isomaltase; switching growing cultures of Glc−cells to the Glc+ medium for several passages results in a progressive reversion to the undifferentiated state, which is completed after seven passages. The dedifferentiation process is associated with a parallel, passage‐related, increase in the rates of glucose consumption and lactic acid production, and decreases of intracellular glycogen content, which return to the values of the undifferentiated original Glc+ cells. The values of these metabolic parameters are correlated, at each passage, with the degree of dedifferentiation of the cells. When these dedifferentiated cells, after having been cultured in Glc+ medium for 20 passages, are switched back to the Glc− medium, they readily grow without mortality, and reexpress the same enterocytic differentiation as the parent Glc− cells. These results show that the capacity of this subpopulation to grow and differentiate in the absence of sugar is a stable characteristic. They further suggest that glucose metabolism interferes with the program of differentiation of HT‐29 cells.
Forskolin induction of S‐100 protein in glioma and hybrid cells
Authors: Higashida Haruhiro , Sano Mamoru , Kato Kanefusa 
Published: February 2005

AbstractThe S‐100 protein level in mouse neuroblastoma (N18TG‐2 and NIE‐115), rat glioma (C6, C6BU‐1, and C6V‐1), and hybrid (NG108‐15, 140‐3, 141‐B, NBr10A, NBr20A, NCB20, and NX3IT) cells was determined with a sensitive enzyme immunoassay system that uses a rabbit antibody to bovine brain S‐100 protein. S‐100 protein was detected in glioma but not in neuroblastoma cells. All seven hybrid cells derived from neuroblastoma and glioma or other types of cells were found to possess a very little or undetectable S‐100 protein. The induction of S‐100 protein level in prestationary phase cultures of glioma C6BU‐1 cells was examined by forskolin, which was a highly specific activator of adenylate cyclase of the cells and produced morphological differentiation. After incubation with 10 μM forskolin for 48 hr, the S‐100 protein level increased 2–2.5‐fold in C6BU‐1 glioma cells whose mean control level was 60 ± 26 ng/mg protein (± SD). The forskolin induction of S‐100 protein in the cells was dose dependent, and the concentration of forskolin required for 50% activation of S‐100 protein was about 0.6 μM. The increase by forskolin was initiated from 10–15 hr after incubation with it and was inhibited with cycloheximide and actinomycin D. In NG108‐15 hybrid cells the induction of S‐100 protein was also observed by forskolin as well as prostaglandin (PG) E1 plus theophylline which are known to activate adenylate cyclase of the cells. The results indicate that S‐100 protein biosynthesis is genetically controlled in these clonal cells, and that S‐100 protein can be regulated in a cAMP‐dependent fashion in prestationary cultures.
Transient and sustained effects of hormones and calcium on membrane potential in a bone cell clone
Authors: Ferrier Jack , Illeman Alan , Zakshek Eva 
Published: February 2005

AbstractMeasurements were made of the electrophysiological and cAMP response to changes in extracellular [Ca2+] and to hormone application in a bone cell clone. Both transient and long‐term electrophysiological responses were studied. An increase in extracellular [Ca2+] usually resulted in a transient hyperpolarization of about 60‐sec duration. In addition, increases in extracelllular [Ca2+] from 0.9 to 1.8 mM and from 1.8 to 3.6 mM resulted in long‐term hyperpolarization and increased potential fluctuations. Increasing bathing [Ca2+] until the membrane potential reached the K+ equilibrium level resulted in a significant decrease in fluctuations. Addition to the bathing medium of quinine, a putative blocker of the Ca2+‐dependent K+ channel, resulted in long‐term depolarization of the mean membrane potential, and a long‐term decrease in potential fluctuations. Addition of Mg2+, a mild antagonist of Ca2+ entry into the cell, produced transient depolarization and reduction of potential fluctuations. These effects suggest that the potential fluctuations reflect cytoplasmic [Ca2+] fluctuations via Ca2+‐dependent K+membrane channels. Under an extracellular[Ca2+] of 1.8 mM, the application of prostaglandin E2 (PGE2), isoproterenol, and parathyroid hormone produced no significant effect on mean membrane potential or on the sustained potential fluctuations, but PGE2 did significantly raise intracellular cAMP. Under an increased bathing [Ca2+], significant changes in mean potential and fluctuations did occur in response to PGE2, but not in response to the other hormones, while the PGE2 effect on cAMP was not greatly changed. Hyperpolarizing transients of about 30‐sec duration occurred in response to all of the hormones, particularly at an extracellular [Ca2+] of 3.6 mM. Thus, there are both transient and long‐term electrophysiological responses to hormone application, with only the long‐term response correlated with the production of cAMP. These electrophysiological responses may represent separate transient and long‐term calcium transport responses to hormone application.
Dexamethasone and retinyl acetate similarly inhibit and stimulate EGF‐or insulin‐induced proliferation of prostatic epithelium
Authors: Chaproniere Donna M. , Webber Mukta M. 
Published: February 2005

AbstractProstatic epithelium proliferates in a defined medium consisting of basal medium RPMI1640 containing transferring (1 μg/ml), EGF (10 ng/ml), and insulin (3.7 μg/ml or 0.1 lU/ml). Although neither dexamethasone nor retinyl acetate affected the proliferation of prostatic epithelium in RPMI1640 containing trans‐ferrin alone, they modify the mitogenic effect of EGF and insulin. Dexamethasone at 10−10 M or retinyl acetate at about 3 × 10−9 M inhibits proliferation stimulated by EGF. Higher concentrations of dexamethasone (10−8−10−6 M) or retinyl acetate (3 × 10−8−10−7 M) enhance the mitogenic activity of EGF. Dexamethasone had a similar effect in the presence of insulin. However, retinyl acetate stimulated, but did not significantly inhibit, proliferation in the presence of insulin. These results suggest that both dexamethasone and retinyl acetate, and possibly other glucocorticoids and retinoids, may regulate the proliferation of prostate epithelium by a dose‐dependent modification of the activity of insulin and EGF.
Relationship of finite proliferative lifespan, senescence, and quiescence in human cells
Authors: Stein Gretchen H. , Namba Masayoshi , Corsaro Cheryl M. 
Published: February 2005

AbstractCell hybrids were formed between human diploid fibroblasts (HDF) and carcinogen‐transformed HDF to determine the relationship among: (1) finite proliferative lifespan, which we define as an age‐related failure of a population to achieve one population doubling in 4 weeks; (2) arrest in a senescent state, which we define as cessation of DNA synthesis in a viable culture that is at the end of its lifespan by the above definition; and (3) arrest in a quiescent state, which we define as cessation of DMA synthesis in a young culture that is crowded or mitogen‐deprived. HDF express all three of these phenotypes, which we have abbreviated FPL+, S+, and Q+, respectively. Carcinogen‐transformed HDF are transformed to immortality (FPL−) and inability to achieve quiescence (Q−). They have no S phenotype because, by definition, this phenotype only exists in FPL+ cells. Fusion of FPL+, Q+, S+ HDF × FPL−, Q− carcinogen‐transformed HDF produced hybrid clones that were FPL+, Q−, and S−, where the S− phenotype means that individual cells continued to synthesize DNA in cultures that had reached the end of their lifespan by our definition. These results are consistent with our hypothesis that senescent HDF and quiescent HDF may share a common mechanism for arrest in G1 phase. We have suggested that this could occur if the aging mechanism that is responsible for the FPL+ phenotype is a progressive decrease in the ability of cells to recognize or respond to mitogenic growth factors. If so, then cells would become physiologically mitogen‐deprived at the end of their lifespan, which would cause them to arrest in the senescent state by the same mechanism that causes young cells to arrest in the quiescent state when they are mitogen‐deprived. This hypothesis predicts that the FPL+ phenotype can be separated from the S+ phenotype–i.e., FPL+ cells can be S+ or S− –and that the Q and S phenotypes are linked‐i.e., FPL+ cells are either Q+ and S+ or Q− and S−. Both these predictions are supported by the present data.
Stimulation of hexose uptake by haemopoietic cell growth factor occurs in WEHI‐3B myelomonocytic leukaemia cells: A possible mechanism for loss of growth control
Authors: Whetton A. D. , Bazill G. W. , Dexter T. M. 
Published: February 2005

AbstractWEHI‐3B myelomonocytic leukaemia cells secrete a haemopoietic cell growth factor (HCGF) which facilitates the proliferation and development of multipotential stem cells and committed progenitor cells. Several cloned, nonleukaemic cell lines (FDC‐P cells) are absolutely dependent on HCGF and die in the absence of it. In these cell lines, factor dependence is associated with the ability of HCGF to increase glucose uptake, thereby controlling glycolytic flux and intracellular ATP levels. We have now investigated the effects of HCGF on glucose uptake in WEHI‐3B cells. At 20°C 2‐deoxyglucose uptake could be stimulated by the addition of HCGF to the extracellular medium. L‐glucose uptake was markedly lower than 2‐deoxyglucose uptake and did not respond to the addition of HCGF. At 37°C no HCGF stimulation of 2‐deoxyglucose uptake was found. However, at this temperature HCGF release from WEHI‐3B cells was markedly higher than at 20°C. Our experiments indicate that HCGF stimulates the glucose transport system in both WEHI‐3 cells and FDC‐P cells. The similarities between the WEHI‐3B cell and FDC‐P2 cell polypeptide phenotype were investigated using two‐dimensional isoelectric focussing/poly‐acrylamide gel electrophoresis. This revealed a high degree of correlation between the two cell types in their protein constituents, indicating a close relationship between the normal and leukaemic cells. These similarities between WEHI‐3B cells and FDC‐P2 cells are considered and their relevance to haemopoiesis and leukaemogenesis is discussed.
The deoxyglucose method adapted for studies of glucose metabolism in the early chick embryo
Authors: Baroffio Anne , Kucera Pavel 
Published: February 2005

Abstract14C‐2‐deoxyglucose (DG), currently employed in in vivo studies of brain glucose metabolism, has been used for determination of glucose consumption in the in vitro developing chick embryo. DG, presented in traces, accumulates in the embryo in proportion with incubation time. Analysis of tissue homogenates shows that the accumulated radioactivity is due to both phosphorylated (DGP) and nonphosphorylated DG. As it is only the radioactivity originating from the DGP that is proportional to glucose utilization, the nonphosphorylated DG must be washed out. The washout shows two distinct kinetics: a fast one corresponding to DG that has entered the cells but has not yet been phosphorylated and a slow one that is probably due to a dephosphorylated DGP coming from a different cellular compartment. On the basis of these results the optimal experimental conditions have been defined, allowing quantitative studies of glucose metabolism during the first day of development of the chicken embryo. From 18 to 24 hr of incubation (end of gastrulation), total glucose consumption increases from 50 nmol · h−1 at stage 3–4 to 90 nmol · h−1 at stage 6–7. This increase mainly reflects the growth of the blastodisc. Comparison with the values of O2 uptake measured at the same period of development suggests that only a fraction of the glucose consumed is oxidized, the major part being converted aerobically to lactate.
Regulation of cell proliferation inhibitory and stimulatory factors diffused by 3T3 cultured cells
Authors: Harel Louise , Blat Christiane , Chatelain Gilles 
Published: February 2005

AbstractThe growth rate of normal cells multiplied in vitro decreases as the cell density of the culture increases. Previous results suggested that this density‐dependent inhibition of growth in nontransformed cells was due to the diffusion of growth inhibitory substances in the medium of dense cultures. In this paper, we demonstrate that dense cultures of 3T3 cells secrete inhibitory and stimulatory factors. Macromolecules of conditioned medium were fractionated on Biogel P150 and the different fractions were tested on quiescent cultures of 3T3 cells stimulated or not to proliferate by addition of α globulin. When target cells were not stimulated to proliferate by addition of exocrine growth factors, we observed the inhibitory activity of a large molecular weight inhibitor (IDF45) and the stimulatory activity of autocrine growth factors (fraction about 35 and 10 K molecular weight), on the incorporation of 14C inosine into nucleotide pool and RNA. However, DNA synthesis was significantly stimulated with fraction 10 K only. This discrepancy between the stimulation of RNA and DNA synthesis may be explained by the presence, simultaneously, of inhibitory and stimulatory factors in fraction 35 and 10 K molecular weight. The presence of inhibitory factor was demonstrated when the fractions were tested on target cells stimulated to proliferate by α globulin addition and labeled with 14C thymidine. In these conditions, the stimulatory activity of autocrine growth factors was not observable, and only the inhibitory activity on DNA synthesis of fractions 35 and 10 K appeared. It is tempting to assume that the regulation of in vitro cell proliferation is determined by the balance between these antagonist stimulatory and inhibitory autocrine growth factors.
Estrogen receptor expression in serially cultivated rat endometrial cells: Stimulation by forskolin and cholera toxin
Authors: Heimann Ruth , Rice Robert H. , Gross Mary K. , Coe Eleanor L. 
Published: February 2005

AbstractSerially propagated with 3T3 feeder layer support, epithelial cells derived from normal rat endometrium expressed estrogen receptor activity. Specific binding of 17‐β‐estradiol was in the range of 30–60 fmol/mg of protein and was of high affinity (Kd = 0.3 nM). A survey of cell lines derived from several other normal epithelia showed that rat vaginal and human cervical cultures also had high‐affinity estrogen receptors (6–13 fmol/mg of protein), while rat epidermal and esophageal cells had no detectable activity. In the endometrial cultures, receptor levels were elevated nearly two‐ to fourfold by cholera toxin or forskolin in the medium. This effect was detectable after 4 hr but not 1 hr of treatment and did not occur in the presence of cycloheximide. We conclude that serially cultivated rat endometrial cells retain hormonal properties expressed in vivo while exhibiting some keratinocyte character. These cells may provide a useful model for study of receptor modulation.
An approach to a selection system for adenosine‐deaminase‐positive (ADA+) cells and detection of rat ADA+ “revertants”
Authors: Hischhorn Rochelle , Ellenbogen Amy , Martiniuk Frank 
Published: February 2005

AbstractWe have substituted deoxyadenosine or adenosine for hypoxanthine in the standard HAT selection system in an attempt to select for ADA‐normal (ADA+) cells. ADA− human lymphoid line cells could not utilize deoxyadenosine as an alternative to hypoxanthine as a purine source (DAT) and failed to grow but were only somewhat inhibited in growth when adenosine was substituted for hypoxanthine (AAT). In contrast, ADA+ cells utilized adenosine or deoxy‐adenosine as efficiently as hypoxanthine as a purine source. Growth in DAT, but not in HAT, of an artificial mixture of one ADA+ human lymphoid cells in 1,000 ADA− cells resulted in enrichment of ADA+ cells to 25–86% of total cells. When we grew a rat ADA− cell line in two variations of the DAT system, we detected at least three electrophoretically different ADA+ patterns, one of which corresponded to normal rat ADA. These could represent “revertants.”