bluuebunny/crossref_metadata_2025_split huggingface dataset:
The potentiation of adenine toxicity to Chinese hamster cells by coformycin: Suppression in mutants with altered regulation of purine biosynthesis or increased adenylate‐deaminase activity
Authors: Debatisse M. , Berry M. , Buttin G. 
Published: February 2005

AbstractWhen added to medium containing coformycin (2 μM or above), adenine is lethal to Chinese hamster fibroblasts at the concentration inhibiting de novo purine biosynthesis (Debatisse and Buttin, '77b). Rescue by hypoxanthine suggested that cells die of IMP starvation when the analog can turn off deamination of both adenosine and adenylate. As predicted from this hypothesis, two classes of variants resistant to the mixture of coformycin + adenine have been isolated: Class 1 variants have altered control of de novo IMP biosynthesis; they fall into two subclasses on the basis of their resistance to adenosine. Class 2 variants have a 6–10‐fold increased level of AMP‐deaminase (E.C.: 3.5.4.6); their growth in the selective medium is temperature‐dependent, a property accounted for by the observation that cell growth in the presence of coformycin imposes a gradual thermodependent decay of specific AMP‐deaminase activity in both wild‐type and variant lines. This control by coformycin of AMP‐deaminase activity isunaltered in mutants deficient in the four activities of adenosine‐kinase, APRT, HGPRT and deoxycytidine‐kinase. Most of the resistant variants are unstable and exhibit either increased or reduced resistance, depending on prolonged growth in selective or normal medium.
DNA synthesis and proliferation of human lymphocytes in vitro: III. Fate of cycling cells in aging cultures of phytohemagglutinin stimulated human lymphocytes
Authors: Mendelsohn John , Hudig Dorothy , Castagnola Janice 
Published: February 2005

AbstractThere are few data available on cell cycle events that occur when proliferation of normal cells in culture is curtailed due to “natural aging” of the culture conditions. Stathmokinetic and cytofluorometry studies were performed on PHA‐stimulated human lymphocyte cultures for eight consecutive days. Cell proliferation peaked on day 5 and then gradually decreased. Percent labeled mitosis curves performed each day demonstrated that, for those cells which progressed to mitosis, the cell cycle time remained constant at 18 ± 1 hour throughout the entire period of culture. However when the fate of all cells pulse‐labeled with 3H‐thymidine (S phase cells) was followed daily, only 64 ± 5% of labeled cells reached mitosis on day 3 and <20% on day 6. When the growth fraction was estimated by standard methods (with the labeling index) and used to predict future cell counts in the culture, proliferation was greatly overestimated; but after correcting the growth fraction for labeled cells not reaching mitosis, proliferation was accurately predicted by a newly derived “dividing fraction.” Flow cytofluorometry confirmed accumulation of cells in S and G2 + M phases, and mitotic indices ruled out accumulation in M phase. Assessment of non‐viable cells with cytofluorometry demonstrated that death occurred in all phases of the cell cycle. We conclude that with increasing age of culture, an increased fraction of cycling PHA‐stimulated lymphocytes fail to progress all the way to mitosis and are arrested in S or G2 phases. These observations provide evidence against the existence of a specific “restriction point” in G1 or at the G1/S interface in aging proliferating human lymphocyte cultures, but it remains to be determined whether cells arrested in S or G2 phases retain the capacity to complete the cell cycle in more favorable culture environments.
Passive membrane permeability to small molecules and ions in transformed mammalian cells: Probable role of surface phosphorylation
Authors: Makan Nizar R. 
Published: February 2005

AbstractAddition of ATP to medium surrounding intact, transformed 3T3 cells causes the formation of aqueous channels in the plasma membrane. This effect of extracellular ATP is sharply dependent on the pH and temperature of the incubation medium, and is inhibited by low levels of La3+ or ruthernium red; inhibition is also obtained with concentrations of Mg2+ ions that exceed a ratio of Mg/ATP of one. The effect of ATP on membrane channel formation is unaffected by chelators of metal ions or by prior modification of the cell surface with various surface‐active enzymes or sulfhydryl reagents. Under conditions which favor aqueous channel formation, incubation of intact 3T6 cells with ATP (γ‐32P) leads to phosphorylation of two membrane components with apparent molecular weight of 40,000 (40K) and 110,000 (110K) daltons; the 110K component which is unaffected by trypsin under normal conditions is rendered trypsin‐sensitive by the phosphorylation reaction, probably as a result of a conformational change. Conditions which inhibit aqueous channel formation also inhibit phosphorylation of the 110K protein and decrease the labeling of the 40K component. These results indicate the probable role of cell surface phosphorylation, involving one or both of these components, in the formation of aqueous channels in transformed 3T3 cells. Aqueous channel formation by extracellular ATP is not associated with gross unfolding of the cell surface as revealed by lactoperoxidase‐catalyzed iodination of the 3T6 cell surface.
Transmembrane potential changes during phagocytosis in rat alveolar macrophages
Authors: Miles P. R. , Bowman L. , Castranova V. 
Published: February 2005

AbstractStudies were carried out to measure changes in the transmembrane potential of rat alveolar macrophages during exposure of the cells to zymosan particles or to the membrane perturbant, phorbol‐12‐myristate‐13‐acetate (PMA), and to determine if changes in membrane potential are related to superoxide anion release. Exposure of the cells to either zymosan or PMA leads to membrane depolarization, which precedes superoxide anion release. Furthermore, the magnitude of the depolarization is dependent upon the concentration of either zymosan or PMA. During exposure of the alveolar macrophages to increasing levels of zymosan, there is an increase in the amount of superoxide released as well as an increase in the magnitude of the depolarization. Incubation of the cells in medium containing 150 mMK+, a medium which causes membrane depolarization, leads to superoxide release from resting cells and a decrease in the amount of superoxide released from cells exposed to zymosan. These results indicate that release of superoxide anion from rat alveolar macrophages is related to membrane depolarization and suggest that the transmembrane potential change may act as a signal to initiate the phagocytotic responses of the cells.
Studies of the copper‐binding proteins in Menkes and normal cultured skin fibroblast lysates
Authors: LaBadie Gundula U. , Beratis Nicholas G. , Price Peter M. , Hirschhorn Kurt 
Published: February 2005

AbstractProteins of approximately 10,000 daltons (presumably metallothionein) and greater than 75,000 daltons bound 64Cu when this metal was added to fibroblast lysates. Treatment with either 2‐mercaptoethanol or the disodium salt of ethylenediamine tetraacetic acid demonstrated that the high molecular weight copper‐binding proteins in lysates prepared from both normal and Menkes fibroblasts exhibited a relatively low affinity for copper compared to the 10,000 dalton protein(s). No difference was detected in the affinity of the low molecular weight protein(s) of normal and Menkes fibroblast lysates for copper. The amount of 64Cu bound to the 10,000 dalton protein(s), however, was approximately two to three times greater in lysates prepared from Menkes fibroblasts than from normal fibroblasts. Mixing experiments indicated that the increased binding of 64Cu to the 10,000 dalton protein(s) in lysates of Menkes fibroblasts did not result from the deficiency of a factor that effects the cleavage of copper from this protein(s), from the presence of a soluble inhibitor, or from the lack of an activator. In addition, the use of lysates, rather than whole cells, demonstrated that the observed differences in copper binding between the normal and the Menkes fibroblasts were not caused by an abnormality in the membrane transport of copper in the mutant cells. Thus the findings suggest that the increased accumulation and the reduced efflux of copper previously observed in cultured Menkes fibroblasts result either from an increased amount of the 10,000 dalton copper‐binding protein(s) or from an increased capacity of this molecule(s) for copper.
Amiloride‐resistant Madin‐Darby Canine kidney (MDCK) cells exhibit decreased cation transport
Authors: Taub Mary , Saier Milton H. 
Published: February 2005

AbstractCation transport systems were investigated in mutant Madin‐Darby Canine Kidney (MDCK) cells resistant to the diuretic drug amiloride. The mutants were isolated previously as clones resistant to the cytotoxic effects of 3 × 10−4 M amiloride. Decreased amiloride transport by the Na+ channel was implicated as the basis of the resistance (Taub, '78). Consistent with this hypothesis, Na+ accumulation was lower in amiloride resistant cells than in normal sensitive MDCK cells. Kinetic studies indicated that Na+ uptake in MDCK cells occurs by a single ATP independent transport system—the Na+ channel.In several amiloride‐resistant clones, including clone Amr2, the decreased Na+ uptake was associated with a decrease in both the Km and Vmaxfor Na+ uptake by the Na+ channel. In Amr2 cells no significant alteration in the inhibitory effect of amiloride on Na+ uptake was observed. As the Na+ channel may actually be a general uptake system for monovalent cations (a number of cations inhibit Na+ uptake), the uptake of these inhibitory cations was examined in Amr2 cells. Both Ca++ and ouabain‐insensitive Rb+ uptake occurred at decreased rates in Amr2 cells as compared with normal MDCK cells. However, further uptake studies suggested that Na+, Ca++ and ouabain‐insensitive Rb+ uptake all occur by different systems. Thus several transport systems may be defective in Amr2 cells. Amr2 cells were also resistant to the inhibitory effects of amiloride on CO2 evolution from pyruvate. These observations indicate that alterations at a number of molecular sites may be associated with defective Na+ transport via the Na+ channel in amiloride‐resistant cells. Thus the amiloride‐resistant cells are potentially valuable in examining the interrelationships between Na+ transport and other cellular functions.
Sodium and rubidium uptake in cells transformed by Rous sarcoma virus
Authors: Bader John P. , Okazaki Takumi , Brown Nancy R. 
Published: February 2005

AbstractRates of uptake and intracellular concentrations of monovalent cations were measured in virus‐transformed and nontransformed chick embryo (CE) cells. Uptake of 22Na+ into cells transformed by the BH strain of Rous sarcoma virus (RSV‐BH) (CE‐BH) was about double the rate of uptake into CE cells, or cells transformed by the Schmidt‐Ruppin strain (RSV‐SR): CE‐SR. Likewise, the rate of efflux of 22Na+ was greater in CE‐BH cells than in CE or CE‐SR cells. The greater permeability of CE‐BH cells to Na+ was apparent in higher intracellular Na+ concentrations. Experiments with cells exhibiting temperature‐dependent transformation showed that new RNA and protein synthesis was a requirement for the acquisition of increased Na+ permeability, suggesting that the change is an indirect effect of the virus‐coded transformation‐inducing protein. Rates of 86Rb+ uptake, used as a measure of K+ influx, were indistinguishable in CE, CE‐BH, and CE‐SR cells. Also, equilibrium intracellular levels of 86Rb+ were similar in transformed and nontransformed cells, as were observed concentrations of K+. Also, no differences in ATPase activity, as indicated by ouabain binding or temperature sensitivity, were observed. We conclude that monovalent cations play no direct role in RSV‐induced transformation, although the higher levels of Na+ in CE‐BH cells may be responsible for other distinguishing biochemical features of these cells.
Sulfhydyl group involvement in the modulation of neutral amino acid transport in thymocyte membrane vesicles
Authors: Kwock Lester 
Published: February 2005

AbstractMembrane vesicles from rat thymocytes accumulate 2‐aminoisobutyric acid in the presence of 0.1 M NaCl. Uptake is half maximal between 15 and 30 seconds after addition of the amino acid and reaches a plateau value after about 2 minutes. The uptake of 2‐aminoisobutyric acid can be modulated by various sulfhydryl reagents. Reduced glutathione leads to an inhibition of uptake whereas oxidized glutathione increases uptake. Agents such as insulin and diamide which can induce disulfide formation lead to an activation of transport. These data indicate that uptake of the Na+‐dependent amino acid, 2‐aminoisobutyric acid, in thymocytes is modulated by a putative plasma membrane, sulfhydryl‐containing protein.
Regulation of cell division and of tyrosinase in B16 melanoma cells by imidazole: A possible role for the concept of metabolite Gene regulation in mammalian cells
Authors: Montefiori David C. , Kline Ellis L. 
Published: February 2005

AbstractResults of hemacytometer cell counts and of tyrosinase measurements made by the Pomerantz method demonstrate that imidazole added to the medium of cultured B 16 mouse melanoma cells can stimulate tyrosinase specific activity and inhibit cell division. These effects are greater than with adenosine 3′,5′ cyclic monophosphate (cAMP) or the cAMP‐phosphodiesterase inhibitor theophylline. The effects of imidazole on cell division and tyrosinase are enhanced by theophylline and antagonized by cAMP. Cyclic AMP‐phosphodiesterase activity in cell‐free extracts can be inhibited by theophyllne and stimulated by imidazole. However, imidazole does not affect cAMP‐phosphodiesterase specific activity in vivo, nor does it affect intracellular cAMP concentrations as determined by competitive protein‐binding assays. In contrast, the specific activity of cAMP‐phosphodiesterase in vivo is stimulated by cAMP and theophylline, supporting the hypothesis that cAMP and agents which increase intracellular cAMP concentrations induce the synthesis of cAMP‐phosphodiesterase. Studies with actinomycin‐D and cycloheximide support the hypothesis that cAMP can also mediate posttranslational activation of tyrosinase. Similar experiments suggest that imidazole, or a derivative therof, can induce the synthesis of tyrosinase at the pretranslational level of control. We hypothesize that this type of regulation (pretranslational) by imidazole may define a role for the concept of “Metabolite Gene Regulation” (MGR), in mammalian cells.
Fluorimetric measurements and chromatin condensation patterns of nuclei from 3T3 cells throughout G1
Authors: Moser Gertrude C. , Fallon Robert J. , Meiss Harriet K. 
Published: February 2005

AbstractUsing two cytological methods based on nuclear morphology, quinacrine dihydrochloride (QDH) staining and premature chromosome condensation (PCC), it has been possible to identify cell cyle positions within G1 of growing and arrested 3T3 cells. The fluorescent intensity of QDH‐stained interphase cells appears to decrease as the cells pass from mitosis to S phase. Likewise, the length and thickness of prematurely condensed chromatids can be related to the cells' position within the G1 period. Data are presented that deal with three interrelated topics: (1) We determined by fluorometric measurements of nuclei from 3T3 cells that the visual observation of the decrease in QDH fluorescence during G1 reflects an actual decrease in total fluorescence and not a dispersion of the fluorescent chromatin in a larger nuclear area. (2) We correlated the results obtained by QDH staining with those of PCC on the same cell samples blocked in G1 by different conditions. Serum‐starved and contact‐inhibited cell nuclei had the highest intensity, hydroxyurea‐treated ones had the lowest intensity, while that of isoleucine‐deprived cells was in between. The same relative order of G1 positions was obtained based on PCC morphology. Thus, both methods monitor the state of chromatin condensation and can be used to identify cell cycle position within G1.(3) We showed with both methods that the states of chromatin resulting from the various G1 blocking conditions differ from each other.
Synthesis of a metallothionein‐like protein in cultured human skin fibroblasts: Relation to abnormal copper distribution in Menkes' disease
Authors: Bonewitz Roland F. , Howell R. Rodney 
Published: February 2005

AbstractA metallothionein‐like protein (MTP) is synthesized in normal diploid human skin fibroblasts cultured in Zn‐ or Cu‐supplemented medium. Synthesis of MTP is not detected in cells cultured without metal supplementation of complete tissue‐culture medium. Cultured fibroblasts from patients with Menkes' disease accumulate excess Cu which chromatographs both with highmolecular‐weight protein(s) and with a Cu‐MTP. Under normal culture conditions, the Menkes' MTP incorporates [35S]‐cystine, but not appreciable amounts of 65Zn. However, Menkes fibroblasts retain the ability to incorporate 65Zn into MTP in response to Zn supplementation of the medium. The results do not support the idea that Menkes' disease results from a failure of Cu to bind to MTP, but rather that an elevated intracellular Cu concentration in Menkes' disease fibroblasts leads to association of excess Cu with high‐molecular‐weight protein, stimulating synthesis of a Cu‐binding MTP.
Modulation of functional and optimal (Na+‐K+)ATPase activity during the cell cycle of neuroblastoma cells
Authors: Mummery Christine L. , Boonstra Johannes , Van Der Saag Paul T. , De Laat Siegfried W. 
Published: February 2005

AbstractFunctional and optimal activities of the (Na+‐K+)ATPase, as determined by ouabain‐sensitive K+ influx in intact cells and ATP hydrolysis in cell homogenates respectively, have been measured during the cell cycle of neuroblastoma (clone Neuro‐2A) cells. The cells were synchronized by selective detachment of mitotic cells.The ouabain‐sensitive K+ influx decreased more than fourfold from 1.62 ± 0.11 nmoles/min/106 cells to 0.36 ± 0.25 nmoles/min/106 cells on passing from mitosis to early G1 phase. On entry into S phase a transient sixfold increase to 2.07 ± 0.30 nmoles/min/106 cells was observed, followed by a rapid decline, after which the active K+ influx rose again steadily from 1.03 ± 0.25 nmole/min/106 cells in early S phase to 2.10 ± 0.92 nmoles/min/106 cells just prior to the next mitosis. The ouabain‐insensitive component rose linearly through the cycle in the same manner as the protein content/cell.Combining total K+ influx values with efflux data obtained previously showed that net loss of K+ occurred with transition from mitosis to G1 phase while net accumulation occurred with entry into S. Throughout mid‐S phase net K+ flux was virtually zero, but a large net influx occurred again just before the next mitosis.The (Na+‐K+)ATPase activity measured in cell homogenates decreased rapidly from mitosis to G1 phase and increased steadily throughout S phase, but the transient activation on entry into S phase was not observed.Complete inhibition of the (Na+‐K+)ATPase mediated K+ influx by ouabain (5 mM) prevents the cells from entering S phase, while partial inhibition by lower concentrations of ouabain (0.2 and 0.5 mM; km = 0.17 mM) causes partial blockage in G1 and, to a lesser extent, a reduced rate of progression through the rest of the cell cycle. We conclude that the transient increase in (Na+‐K+)ATPase mediated K+ influx at the G1/S transition is a prerequisite for entry into S phase, while maintenance of adequate levels of K+ influx is necessary for normal rate of progression through the rest of the cell cycle.
The reversible inability of isoproterenol‐activated parotid gland cells to initiate DNA synthesis in the hypocalcemic thyroparathyroidectomized rat
Authors: Tsang B. K. , Whitfield J. F. , Rixon R. H. 
Published: February 2005

AbstractAn intraperitoneal injection of the beta‐adrenergic drug dl‐isoproterenol hydrochloride (100 mg/kg body weight) into male (190–210 g) albino rats caused two cyclic AMP surges (peaking at 10 minutes and again between 8 and 12 hours) and the initiation of DNA synthesis (between 16 and 20 hours) in the parotid glands. The parotid cells in hypocalcemic thyroparathyroidectomized rats still responded to isoproterenol injection by generating the two cyclic AMP surges, but they did not initiate DNA synthesis unless a blood calcium‐elevating combination of parathyroid hormone (50 USP units/100 g of body weight) and 1α,25 (OH)2 vitamin D3(200 pmoles/100 g of body weight) was injected along with the isoproterenol.
Unbalanced growth and cell death in HeLa S3 cultures treated with DNA synthesis inhibitors
Authors: Frankfurt Oskar S. 
Published: February 2005

AbstractFlow cytometry indicated that significant amounts of dsRNA were accumulated in HeLa S3 cells blocked at or near G1/S boundary by hydroxyurea (HU) or excess thymidine (TdR). The dsRNA/DNA ratio increased in these cells in a manner characteristic of unbalanced cell growth. In HU‐treated cells, dsRNA content was maximal 16 hours after addition of the drug and did not change significantly during the next 24 hours. The DNA content in blocked cells increased by 10%. Cell viability assessed by colony formation in soft agar decreased exponentially in HU‐treated cultures after 16 hours of incubation. Correlation between loss of cell viability and rate of cell proliferation after removal of HU was observed, as determined by cell count and analysis of cell cycle progression. In TdR‐treated cultures cells slowly progressed into mid S‐phase during 40 hours and dsRNA accumulation continued during this period. Cell viability was not significantly affected by treatment with excess TdR, indicating that unbalanced growth per se, as measured by dsRNA accumulation, is not lethal for the cells. After reversal of DNA synthesis inhibition by removal of the drug, cells treated with HU for 16 hours or TdR for 16–24 hours promptly progressed through the cell cycle. This progression was accompanied by accumulation of significant amounts of dsRNA. As a result, cells in G2 phase had a very high dsRNA content leading to retention of the unbalanced condition (increased dsRNA/DNA ratio) in the daughter cells. It is suggested that dsRNA accumulation in the cell is controlled to a certain degree by cell progression through the S phase. This type of control, evidently, was reflected in limited dsRNA accumulation in the cells blocked at or near G1/S border, in continuous dsRNA accumulation in the cells slowly progressing through S phase, and in accumulation of large amounts of dsRNA after renewal of progression through the S phase.
Isolation and preliminary characterization of bleomycin‐resistant mutants from Chinese hamster ovary cells
Authors: Akiyama Shin‐Ichi , Kuwano Michihiko 
Published: February 2005

AbstractStable mutants resistant to an anticancer antibiotic, bleomycin‐A2, were selected in Chinese hamster ovary (CHO) cell either spontaneously or after ethylmethane sulfonate mutagenesis. Fluctuation analysis showed that bleomycin resistance occurs in CHO at a rate of 6.50–6.58 × 10−7 mutations per cell per generation. Bleomycin‐A2‐resistant cell lines exhibited increased resistance to bleomycin analogs—bleomycin‐A5, ‐B2, ‐B4, and pepleomycin. Colchicine, mitomycin C, and ultraviolet light irradiation inhibited colony formation equally in CHO cells and in bleomycin‐resistant mutants. Cell‐cell hybridization tests showed that bleomycin‐resistance behaves as a dominant trait. Bleomycin‐in‐activating activity in the mutant cell extracts was three to fourfold higher than that in extracts of the parental CHO cell.