bluuebunny/crossref_metadata_2025_split huggingface dataset:
Glucose metabolism in murine fetal cortical brain cells: Lack of insulin effects
Authors: Gorus Frans K. , Hooghe‐Peters Elisabeth L. , Pipeleers Daniël G. 
Published: February 2005

AbstractGlucose uptake and oxidation were markedly higher in cultured than in freshly isolated neural cells, prepared from murine fetal brain cortices. The hexose transport process–measured as 3‐O‐methyl‐D‐glucose uptake–appeared comparable in both conditions, and proceeded proportionally to the extracellular sugar concentration up to 6 mM. In contrast, glucose oxidation occurred independently of the prevailing glucose concentration from 1.4 mM on. Acute or chronic exposure to insulin exerted no effect upon cellular glucose uptake or oxidation. These results suggest that glucose handling by maturing fetal cortical cells is mainly determined by the rate of cellular glucose breakdown rather than by the rate of glucose transport into the cell; the marked rise in cellular glucose metabolism during culture might result from the synthesis and/or activation of a key enzyme in glucose catabolism. Our observations also indicate that the previously described neurotrophic effects of insulin are not mediated via enhanced glucose handling.
Regulation of the rate of cell cycle progression in quiescent cytolytic T cells by T cell growth factor: Analysis by flow microfluorometry
Authors: Sekaly Rafick P. , MacDonald H. Robson , Nabholz Markus , Smith Kendall A. , Cerottini Jean‐Charles 
Published: February 2005

AbstractWe have previously shown that >90% of B6.1 cells, a murine cytolytic T lymphocyte (CTL) cloned line which is solely dependent on T cell growth factor (TCGF) for continuous growth in vitro, accumulates in the G1 phase of the cell cycle after transfer into culture medium containing no TCGF. Moreover, when such quiescent cells are exposed again to TCGF, >85% reenter the S phase and subsequently divide in a relatively synchronous fashion. In this study, the regulation of the rate of cell cycle progression of quiescent B6.1 cells after exposure to TCGF was analyzed using two complementary DNA staining techniques, namely, the propodium iodide method (to enumerate cells entering the S phase) and the Hoechst 33342‐bromodeoxyuridine substitution technique (to enumerate cells which have gone through mitosis). After TCGF addition, quiescent B6.1 cells resumed DNA synthesis and divided after a lag phase of 10 and 20 h, respectively. The duration of the lag phase was found to be dependent on the length of time during which quiescent B6.1 cells had been deprived of TCGF, but was independent of the concentration of TCGF used for restimulation. In contrast, the proportion of cells responding to TCGF as well as the rate of their first passage through mitosis was dependent on TCGF concentration. The presence of TCGF for at least 6 h was required for a maximal response. Moreover, direct evidence was obtained that TCGF by itself was able to stimulate proliferation of quiescent B6.1 cells in the absence of other growth factors and serum constituents other than bovine serum albumin, transferrin, and lipids.
Effects of temperature on aggregation and the mitogen‐induced exit of lymphocytes from the resting state
Authors: Hall David J. , O'Leary James J. , Rosenberg Andreas 
Published: February 2005

AbstractWe have determined the temperature dependence of the kinetics of entry into the first S phase of phytohemagglutinin‐stimulated lymphocytes under conditions varying the stability of substrata over which the cells have settled. An exponential model was used to characterize entry into S phase. This model yields as parameters duration of lag period, t0, apparent first order rate constant for entry, k, and the number of cells committed to enter the first S phase, NA(t0). Values of t0 and NA(t0) show a 1.5‐fold and 2.0‐fold decrease and increase, respectively, over a 4°C temperature range and are independent of variation in substrate stability. The temperature dependence of the apparent first‐order rate constant, k, however, is strongly influenced by stability. The observed activation energy increases from 3.0 kcal to 37 kcal when the substratum is agitated. This correlates well with reduced adherence of multicellular aggregates in agitated samples. The temperature dependencies for these three parameters are all numerically different, indicating that these parameters are determined by different rate‐limiting processes. We propose that the mechanism mirrored by k is linked to the adherence of multicellular aggregates to the substratum.
Desensitization of the serum effect on Na+ influx in cultured human fibroblasts
Authors: Villereal Mitchel L. , Owen Nancy E. 
Published: February 2005

AbstractStimulation of an amiloride‐sensitive Na+ influx pathway, which mediates Na+/H+ exchange, has been postulated to be an important step in the initiation of DNA synthesis in quiescent human fibroblasts. If the elevation of intracellular Na+ or the alkalinization of intracellular pH resulting from the activation of this system is a trigger for subsequent mitogenic events, then its inactivation may also be important to cellular functions. We investigated the duration of the activation of Na+ influx by serum in human foreskin fibroblasts (HSWP). It was found that activation of Na+ influx by 10% serum was transient, declining with a t1/2 = 15 min. Similarly, the Na+ content of the cells rose rapidly following serum addition and decreased with a t1/2 = 15 min. In addition, both the lys‐bradykinin‐ and the vasopressin‐stimulated Na+ influx and Na+ content declined with a t1/2 of approximately 15 min. Similar results were obtained using both Tris‐buffered and Hepes‐buffered, amino‐acid‐free EMEM. Finally, the above experiments were repeated under conditions normally used to assess the mitogenic response of cells. It was found that in cells arrested in G0 by serum deprivation in CO2‐buffered EMEM, the serum activated Na+ flux was also transient with a t1/2 of approximately 20 min. The desensitization of cells to serum could be readily (t1/2 = 20′) reversed by a subsequent incubation of cells in serum‐free medium. Stimulation of Na+ influx by both the divalent cation ionophore A23187 and the phospholipase activator melittin in also desensitized rapidly, suggesting the process is independent of receptor downregulation. The desensitization during serum preincubation occurred in both low Na+ and low pH medium suggesting that the process is not due to negative feedback on the transport system via a rise in cellular Na+ concentration or a rise in intracellular pH. Although the mechanism of desensitization is at present not known, it is likely to be a physiologically important event.
Sodium‐potassium exchange in sea urchin egg. II. Ionic events stimulating the Na+‐K+ pump activity at fertilization
Authors: Ciapa B. , Allemand D. , Payan P. , Girard J. P. 
Published: February 2005

AbstractThe preceding paper (Ciapa et al., 1984) provided biochemical and kinetic characterization of the Na+‐K+ exchange in Paracentrotus lividus eggs. The present work is a study of the ionic events involved in the stimulation of the Na+‐K+ transporter after fertilization. Fertilization in low Na+‐external medium containing amiloride (0.1 mM) suppresses the stimulation of the net efflux of H+ and 86Rb uptake. Activation of eggs with the ionophore A23187 leads to stimulation of both Na+‐H+ exchange and ouabain‐sensitive 86Rb influx. When eggs were activated with A23187 in artificial seawater, 86Rb uptake and 24Na influx showed similar saturable kinetics with respect to the external Na+. A23187 treatment of eggs in Na+ ‐free artifical seawater did not stimulate the Na+‐K+ exchange until 10 mEq Na+ was added. Activation of eggs by NH4Cl (5 mM) stimulated 86Rb influx and Na+ exit; both fluxes were ouabain sensitive. Monensin increased cell Na+ of unfertilized eggs without any significant increase in intracellular pH: a condition in which 86Rb influx was not markedly stimulated. Addition of 10 mEq Na+ to unfertilized eggs in Na+‐free artificial seawater stimulated 86Rb uptake but to a lower extent that did 10 mEq Na+ plus sperm. It is concluded that (1) the stimulation of the Na+‐K+ pump at fertilization has an absolute requirement for the Na+‐H+ exchange; (2) the alkalinization of eggs resulting from the acid efflux is a prerequisite for the enhancement of the Na+‐K+ pump; (3) the amount of Na+ entering eggs at fertilization determines the intensity of the Na+‐K+ exchange; (4) early events of fertilization such as exocytosis and calcium release which may be involved in the stimulation of the Na+‐K+ pump must necessarily be coupled to cell alkalinization.
Effects of inhibitors of protein degradation on the rate of protein synthesis in Chinese Hamster Ovary cells
Authors: Scornik Oscar A. 
Published: February 2005

AbstractIn the absence of serum and amino acids, cultured Chinese Hamster Ovary cells released to the medium two thirds of the leucine produced by protein degradation. Because protein synthesis requires all the amino acids, the loss of leucine implies incomplete reincorporation of the other amino acids as well. Leupeptin (0.45 mg/ml) and chloroquine (up to 40 μM) inhibited protein breakdown by 21 and up to 41%, respectively, and resulted in proportional decreases in protein synthesis. Chloroquine abolished the stimulation of protein breakdown by amino acid deprivation. From the values of protein synthesis and leucine output with and without chloroquine, it is estimated that the stimulation of protein degradation not only permitted continuing protein synthesis but also increased amino acid output. In the presence of serum or amino acids protein breakdown was slower than in their absence and less sensitive to inhibition by chloroquine, but proportional effects on synthesis and degradation were still observed. It is suggested that protein degradation may be necessary for the maintenance of optimum intracellular concentrations of amino acids even in the presence of extracellular amino acids.
An in vitro clonal assay of adherent stem cells (ASC) in mouse marrow
Authors: Reincke Ursula , Rosenblatt Margery , Hellman Samuel 
Published: February 2005

AbstractHematopoietic stem cells with high proliferative capacity can be assayed when stromal bone marrow cultures are overlaid with limiting dilutions of marrow samples. This leads to hematopoietic growth after 4 weeks in a fraction of cultures, consistent with expectations based on Poisson statistics. It will be shown that monoclonal cultures are obtained that last from 2 to 15 weeks and that can generate up to several million mature granulocytes. The originating clone‐forming cell is named adherent stem cell (ASC) because of its adherence to plastic or stromal surfaces. The ASC is comparable to the CFU‐S in frequency, proliferative capacity and in its ability to give rise to CFU‐S. As an unexpected additional finding we report that a mode of “clonal succession” was apparent in cultures which expressed more than one clone.
Flexible association of hemopoietic differentiation programs in multilineage colonies
Authors: Lim B. , Jamal N. , Messner H. A. 
Published: February 2005

AbstractPluripotent hemopoietic progenitors lose potentialities during the process of differentiation. We have examined events that lead to lineage restriction by determining the cellular composition of 785 multilineage colonies grown from peripheral blood samples of glucose‐6‐phosphate‐dehydrogenase (G‐6‐PD) heterozygous volunteers. Of these colonies, 762 contained only one isoenzyme type and were considered to be of clonal origin. A considerable heterogenity was observed. Some colonies were composed of cells belonging to two different lineages, while other colonies contained three or more different cell types. A small number of colonies consisted—in addition to myeloid cells—of T‐lymphocytes. The variable association within individual colonies of members belonging to different hemopoietic lineages suggests a flexible determination and expression of differentiation programs by early progenitors.
Interaction of circulating cell‐derived and plasma growth factors in stimulating cultured smooth muscle cell replication
Authors: Clemmons David R. 
Published: February 2005

AbstractMultiple growth factors that circulate in plasma have been shown to stimulate cellular growth in vitro. The plasma growth factors appear to stimulate DNA synthesis in cultured fibroblasts only after prior exposure of cell growth factors derived from circulating cell types, such as platelets and macrophages. The purpose of these studies was to investigate the role of the plasma growth factors in stimulating smooth muscle cell replication following exposure to platelet‐derived growth factor (PDGF). Following transient exposure to PDGF, insulin stimulated smooth muscle cell replication but only when supraphysiologic concentrations were used (i.e., greater than 1.0 μg/ml). Somatomedin‐C (Sm‐C), in contrast, was found to stimulate a 320% increase in [3H]thymidine incorporation when concentrations that are present in extracellular fluids were used (i.e., 0.5–10 ng/ml). Epidermal growth factor (EGF), an important mitogen for multiple cell types, caused a 70% increase in [3H]thymidine incorporation when added to quiescent cells following PDGF exposure, and EGF caused a substantial increase in the absolute level of [3H]thymidine incorporation when coincubated with Sm‐C. When EGF (1 ng/ml) was incubated simultaneously with concentrations of Sm‐C between 1 and 10 ng/ml plus Sm‐C‐deficient plasma, maximal [3H]thymidine incorporation was 2.1‐fold greater in the presence of EGF. In contrast, insulin (20 ng/ml), when coincubated with Sm‐C under similar conditions, had no enhancing effect on the cellular response to Sm‐C. None of the plasma factors tested was an effective stimultant of replication when incubated either in serum‐free medium or in the presence of Sm‐C‐deficient plasma without prior PDGF exposure. Hydrocortisone was shown to inhibit smooth muscle cell replication in concentrations between 10−7and 10−5M. In summary, multiple plasma growth factors can stimulate the smooth muscle cell replication, and Sm‐C appears to be most effective of those tested. Insulin and EGF appear to work by different mechanisms; that is, EGF can facilitate the cellular response to Sm‐C, whereas insulin is effective only at supraphysiologic concentrations at which it will directly bind to Sm‐C receptors.
Interferon inhibition of thymidine incorporation into DNA through effects on thymidine transport and uptake
Authors: Pfeffer Lawrence M. , Tamm Igor 
Published: February 2005

AbstractReplenishment of medium after 72 hr of growth of HeLa‐S3 cells in dense suspension cultures increased [3H]‐thymidine uptake into cells and incorporation into DNA, with the levels reaching a peak ∼ 12 hr following medium change; β interferon inhibits the enhanced uptake of [3H]‐thymidine and labeling of DNA in a dose‐dependent manner. Some reduction in these processes is observed at a concentration as low as 1 u/ml, and ∼ 75% inhibition at 640 u/ml. Kinetic analysis has revealed that the rate of labeling of the acid‐soluble pool with [3H]‐thymidine, measured either at 22°C, or 37°C, is reduced in interferon‐treated (640 u/ml, 24 hr) HeLa‐S3 cells. At 22°C, the initial rate of thymidine transport at a high (500 μM) thymidine concentration, determined within the first 30 sec of [3H]‐thymidine addition was depressed by 44% in interferon‐treated HeLa cells. At 37°C, labeled precursors accumulate in acid‐soluble material for ∼ 8 min after the addition of [3H]‐thymidine, after which an apparent equilibrium level is attained. At this temperature, the rate of thymidine uptake and the apparent equilibrium level attained were depressed by 70% in interferon‐treated HeLa cells. The reduced incorporation of [3H]‐thymidine into DNA in interferon‐treated HeLa‐S3 cells can be largely explained by interferon inhibition of thymidine transport and phosphorylation.
Treatment of lymphoblastoid cells with interferon decreases insulin binding
Authors: Faltynek Connie R. , McCandless Sarah , Baglioni Corrado 
Published: February 2005

AbstractLymphoblastoid Daudi cells, which are highly sensitive to growth inhibition by interferon (IFN), can be grown in a defined serum‐free medium containing insulin, transferrin, and albumin as the only proteins. We examined whether the growth inhibition by IFN could be in part due to a change in receptors for insulin or transferrin. Cells treated for at least 2 days with 100 units/ml of IFN‐α2 bound less 125I‐insulin and after 3 days of treatment this binding was reduced by more than 50%. No change in the binding of 125I‐transferrin was observed. Treatment with IFN of Raji cells, which are resistant to growth inhibition by IFN, resulted in a similar decrease in 125I‐insulin binding. Growth inhibition of Daudi cells by serum deprivation had no effect on 125I‐insulin binding. Therefore, the IFN‐induced loss of insulin binding sites is not a consequence of growth inhibition.
Self‐inhibition of chloride transport in ehrlich ascites tumor cells
Authors: Levinson Charles 
Published: February 2005

AbstractPrevious studies have shown that mediated Cl− transport which occurs by at least two processes (Cl−‐dependent cation cotransport and Cl− self‐exchange) becomes progressively inhibited when extracellular Cl− exceeds about 60 mM (Hoffmann et al., 1979). To account for this type of kinetic behavior, that is, self‐inhibition, an anion transport system possessing two sites, a high affinity transport site and a lower affinity modifier site is suggested (Dalmark, 1976). In the present experiments we have attempted to determine which of the mediated transport pathways is susceptible to self‐inhibition by studying the dependence of the steady state Cl− flux on the extracellular Cl− concentration and how DIDS, an inhibitor of Cl− self‐exchange, and H+ affect this relationship. Addition of DIDS to Ehrlich cells results in inhibition of Cl− transport at every Cl− concentration tested (40–150 mM). Moreover, the Cl− flux/Cl− concentration relationship no longer exhibits self‐inhibition, suggesting that this phenomenon is a characteristic of the Cl− self‐exchanger rather than of the Cl−‐dependent cation cotransport system. Lowering the extracellular pH (pHo) from 7.35 to 5.30 stimulates Cl− transport by a process that saturates with respect to [H+]. Half‐maximal stimulation occurs at pHo 6.34. A comparison of the kinetic parameters, Ks and Jmax, calculated from the ascending limb of the Cl− flux/Cl− concentration curve at pHo 7.30 to those at pHo 5.50 show that the values for Ks are almost identical (23.6 mM and 21.3 mM, respectively), while the values for Jmax [(22.2 mEq/Kg dry wt). min] differ by only 15%. This finding along with the observation that DIDS completely blocks H+ stimulation of Cl− transport is compatible with the suggestion that H+ interact at the modifer site of the Cl− self‐exchanger and thereby prevents self‐inhibition.
Apparent desensitization of swiss 3T3 cells to the mitogens FGF and vasopressin
Authors: Richmond K. M. V. , Riddle P. N. , Brooks R. F. 
Published: February 2005

AbstractThe growth factors FGF and vasopressin were found to have only a transient effect on confluent quiescent monolayers of Swiss 3T3 cells. Whether measured as cumulative entry into S‐phase by autoradiography, or as cell division by time‐lapse filming, the elevated rate of cell proliferation was maintained only over 10‐15 hr. Several trivial or artifactual explanations for this transience were ruled out, including toxicity of 3H‐thymidine; exhaustion or degradation of medium components, nutrients or growth factors (although some medium depletion was observed); and the generation during quiescence of cells incapable of division. We have also eliminated heritable variation in the capacity to respond to individual growth factors. However, unstable phenotypic heterogeneity in growth factor requirements between cells may play some part, as found elsewhere for the response to low concentrations of serum (Brooks et al, 1984). Cell populations that had ceased to respond to vasopressin recovered their sensitivity after 2‐3 days' incubation in conditioned medium lacking vasopressin. The phenomenon thus resembles the mitogen‐induced desensitization described by Collins and Rosengurt (1982, 1983). However, in our case, the loss of sensitivity was not selective for vasopressin but applied also to epidermal growth factor (EGF) and to prostaglandin F2α. Furthermore, changes in responsiveness to vasopressin with time were associated with changes in cell density. Although some element of selective desensitization has not been ruled out, the transient response in our experiments can be accounted for in terms of unstable heterogeneity in growth factor requirements and/or in terms of density‐dependent regulation of growth.
Variant (MDCK) kidney epithelial cells altered in response to inducers of dome formation and differentiation
Authors: Lever Julia E. 
Published: February 2005

AbstractConfluent cultures of the MDCK kidney epithelial cell line exhibit dome formation, a result of transepithelial fluid transport influenced by cell‐cell and cell‐substratum interaction. Dome formation was inducible by hexamethylene bisacetamide (HMBA) or dimmethylformamide (DMF), compounds known as inducers of cell differentiation (Lever, 1979b). Analysis of the incidence of the dome‐forming phenotype in colonies derived nonselectively from the MDCK cell line suggested that inducers recruit an increased fraction of the cell population to express dome formation. Variant MDCK cell lines were isolated which differed from the parental line in response to inducers while retaining cuboidal epithelial morphology. In five independently isolated and cloned MDCK variants, dome formation was not inducible by DMF and only marginally increased by HMBA. This phenotype was also associated with increased cell adhesiveness to a plastic substratum. Results from cocultivation experiments suggested that the DMF‐unresponsive phenotype of variant cells may be partially overcome by cell‐cell contact with wild‐type cells. Sodium pump transport activity assessed by ouabain‐sensitive Rb+ uptake was partially inhibited by HMBA and by DMF in a “wild‐type” inducer‐responsive clone. By contrast, DMF did not inhibit ouabain‐sensitive Rb+ uptake in DMF‐unresponsive variant clones, and sodium pump inhibition by HMBA was greatly diminished. This close correspondence between altered sodium pump modulation by inducers in variant clones and their altered dome‐forming response reinforces our previous conclusions (Kennedy and Lever, 1984) that sodium pump modulation is closely associated with mechanisms of inducer action. Taken together, these findings implicate cell‐cell interaction, cell‐substratum interaction and sodium pump modulation in regulation of the differentiated phenotype of this cell line.