bluuebunny/crossref_metadata_2025_split huggingface dataset:
Cellular responses to external ATP which precede an increase in nucleotide permeability in transformed cells
Authors: Weisman Gary A. , De Barun K. , Friedberg Ilan , Pritchard Robert S. , Heppel Leon A. 
Published: February 2005

AbstractTransformed mouse fibroblasts, such as 3T6, exhibit an increase in plasma membrane permeability to nucleotides and other normally impermeant molecules when incubated with external ATP in an alkaline medium low in divalent cations. Increased nucleotide permeability, induced by external ATP, occurs after a 3‐ to 5‐min lag period. Prior to this event, there is a dramatic Na+ influx and K+ efflux, a significant reduction in the levels of intracellular ATP and organic phosphates, and a reduction in the plasma membrane potential. Accordingly, we postulate that these cellular responses to external ATP play a role in the efflux of nucleotides.Ouabain, a specific inhibitor of the plasma membrane (Na+, K+)‐ATPase, acts together with low concentrations of external ATP to increase nucleotide permeability in 3T6 cells. This effect occurs at concentrations of ouabain and ATP which alone do not increase nucleotide permeability. In addition, ouabain and low concentrations of ATP alone have little effect on the level of intracellular ATP. This is in contrast to energy inhibitors and uncouplers which appear to enhance nuclectide permeability by lowering the intracellular ATP concentration. Ouabain alone causes a threefold increase in intracellular Na+ levels and a similar reduction in intracellular K+ levels under our experimental conditions, supporting the idea that ion fluxes are involved in the mechanism of permeabilization.
Regulation of mitotic activity and the cell cycle in primary chick muscle cells by neurotransferrin
Authors: Popiela Heinz , Taylor Daniel , Ellis Stanley , Beach Robert , Festoff Barry 
Published: February 2005

AbstractWe previously demonstrated that neurotransferrin (NTF), a transferrin extracted from adult chicken peripheral nerves, promotes growth of primary chick muscle cells in the absence of embryo extract. NTF was shown to stimulate DNA synthesis and cell proliferation. In the present study, we demonstrate that NTF is a mitogen using two independent methods; counts of orcein‐stained mitotic figures and analysis of cell cycle kinetics with a fluorescence‐activated cell sorter. In low‐density cultures mitotic activity increases with increasing doses of NTF followed by a plateau at concentrations greater than 6 μg/ml. Residual, embryonic mitotic activity progressively declines with time after plating muscle cells in the absence of NTF. Absence of NTF for 2 days causes cells to lose irreversibly their myogenic potential. In the presence of NTF, mitotic activity increases for 2 days followed by a decline concurrent with myoblast fusion and formation of myotubes. Cell cycle analysis showed that NTF addition causes cell populations to shift from Gt to S and G2 + M within 18.5 hr. Muscle cells, plated at high densities in the absence of NTF, show mitotic activities similar to those plated at low densities in the presence of NTF. Addition of NTF to high‐density cultures is ineffective in stimulating mitosis. These studies show that at typical cell plating densities, NTF is a required mitogen for primary chick muscle cell cultures.
Different growth factors stimulate cell division of rat mammary epithelial, myoepithelial, and stromal cell lines in culture
Authors: Smith J. A. , Winslow D. P. , Rudland P. S. 
Published: February 2005

AbstractA number of single‐cell‐cloned cell lines have been used to examine the growth‐promoting effects of putative mammotrophic agents on the various cell types in normal and neoplastic rat mammary glands. A partially purified novel pituitary‐derived growth factor stimulates only cuboidal epithelial cells to divide whereas fibroblast growth factor (FGF) stimulates the growth of stromal and myoepithelial‐like cells. Epidermal growth factor (EGF) has a widespread but variable growth‐stimulating action, but prolactin and growth hormone are essentially inactive when added alone at a concentration of 5 μg/ml. Phosphoethanolamine stimulates the growth of one epithelial cell line and a derivative myoepithelial‐like cell line, but is inactive on the other cell lines tested. The use of defined cloned cell lines provides a direct and reproducible assay for the identification and purification of inducers of mammary growth.
A reduction in the activity of the NA+, K+ ‐pump in dimethylsulfoxide‐treated friend erythroleukemia cells is not due to partial inactivation of the NA+, K+ ‐ATPase
Authors: Schaefer Andras , Munter Karl‐Heinz , Geck Peter , Koch Gebhard 
Published: February 2005

AbstractTreatment of Friend‐erythroleukemia cells with 1.5% dimethylsulfoxide (DMSO) caused a decrease in ouabain sensitive 86Rb+‐uptake beginning six to seven hours after DMSO addition indicating a reduced function of the Na+, K+‐pump. However, analysis of the ouabain sensitive 86Rb+‐uptake after Na+‐preloading of the cells as well as measurements on the Na+, K+‐ATPase activity in isolated membrane fragments revealed that no inhibition of the Na+, K+‐ATPase occurred during the first 12 hours. On the contrary the Na+, K+‐ATPase activity was initially enhanced and then returned to control levels during the early phase of induction by DMSO. On the other hand, 22Na+‐transport into DMSO‐treated cells was reduced similar to the ouabain sensitive 86Rb+ uptake in cells without Na+ preloading. The piretanide sensitive 86Rb+‐uptake, due to the Na+, K+, 2Cl− cotransport system was inhibited after seven hours exposure to DMSO. Some three hours after DMSO addition the incorporation of 35S‐methionine into proteins began to decrease, which was accompanied with or followed by a reduction in the methionine uptake of DMSO treated cells. Membrane‐potential‐dependent tetraphenylphos‐phonium cation uptake was not altered relative to the controls in the first 12 hours following DMSO addition. These results suggest that the reduced activity of the Na+, K+‐pump in Friend cells after DMSO exposure is not due to inhibition of the Na+, K+‐ATPase, but most probably due to a smaller Na+‐influx, which results from inhibition of Na+‐cotransport processes (amino acid uptake, Na+, K+, 2Cl− cotransport system).
In vitro stimulation of articular chondrocyte differentiated function by 1,25‐dihydroxycholecalciferol or 24R,25‐dihydroxycholecalciferol
Authors: Harmand M. F. , Thomasset M. , Rouais F. , Ducassou D. 
Published: February 2005

AbstractThe effects of 1,25‐dihydroxycholecalciferol (1,25‐(OH)2D3) (10−13M‐10−8 M) and 24R,25‐dihydroxycholecalciferol (24R,25‐(OH)2D3) (10−12M‐10−7M) on cell proliferation and proteoglycan deposition were examined in our newly developed multilayer culture system for rabbit and human articular chondrocytes. The cells are embedded in an extracellular matrix similar to that seen in vivo and maintain their in vivo phenotype. We extracted and purified native proteoglycans and degraded material from three culture compartments: the medium, intercellular matrix, and cells. Proteoglycan synthesis and deposition were analyzed by measuring 35SO4 incorporation, hexuronic acid, and galactose contents. In both rabbit and human chondrocyte cultures, chronic 1,25‐(OH)2D3 treatment inhibited chondrocyte proliferation and stimulated proteoglycan synthesis and accumulation in the three compartments at 10−12‐10−8M. maximal effect was at 10−10M. Cell proliferation was reduced by 55% and the content of hexuronic acid (or galactose) was increased to about three times that of controls in all compartments. 1,25‐(OH)2D3 did not alter the proteoglycan composition. Chronic 24R,25‐(OH)2D3 treatment induced comparable effects with a maximum at 10−8M. When human dermal fibroblasts were treated as above both vitamin D metabolites increase mitosis. 1,25‐(OH)2D3 mainly reduced the pericellular deposition of proteoglycans, while 24R,25‐(OH)2D3 appeared to reduce their synthesis and deposition in both medium and pericellular compartments. These results suggest that both 1,25‐(OH)2D3 and 24R,25‐(OH)2D3 act specifically on articular chondrocytes to promote phenotype expression.
The effects of epidermal growth factor on cell proliferation and prolactin production by GH3 rat pituitary cells
Authors: Yajima Yukiko , Saito Toshikazu 
Published: February 2005

AbstractThe effects of epidermal growth factor (EGF) were studied in rat pituitary tumor cells, GH3, grown in serum‐supplemented and serum‐free chemically defined media. EGF (1 nM) increased the cell number to 132% of the control cultured in the defined medium during a 6‐day incubation period, while it decreased the cell number to 60% of the control in the serum‐supplemented medium. EGF altered the morphology of the cells grown in the defined medium more markedly to an elongated conformation than that of cells grown in the serum‐supplemented medium. EGF also stimulated prolactin (PRL) production by culture in the presence or absence of serum. The effects of the cell density of GH3 on the action of EGF were shown to appear in two ways. The mitogenic influence of EGF was more effective on, and more responsive to, high‐density cells, whereas the stimulatory action on PRL production was less effective on high‐density cells. However, the inhibitory effects on cellular growth appeared independently of cell densities. The results obtained with 125I‐EGF binding experiments indicated that the number of binding sites, affinity, and internalization of EGF receptors were similar in either serum‐supplemented or serum‐free culture. At low cell density, the number of available 125I‐EGF binding sites per cell was larger than at high cell density. These results suggested that there was no apparent correlation between EGF binding and its differing effects on the growth of GH3 cultured in the serum‐supplemented and the defined medium.
Rat mammary preadipocytes in culture produce a trophic agent for mammary epithelia—prostaglandin E2
Authors: Rudland Philip S. , Twiston Davies Anna C. , Tsao Sai‐Wah 
Published: February 2005

AbstractCell strains and cell lines rat mammary (Rama) 350–353 have been isolated from the slowly adherent stromal fraction of enzymatically digested rat mammary glands. Primary cultures of this fraction yield fat cells on extended culture. Their proportion can be increased with horse serum or growth hormone in the medium, and this increase is associated with a 100‐fold or more increase in the release of radioimmunoassayable prostaglandins of the E type (PGE). The stromal cell strains and lines that are capable of yielding fat cells also secrete elevated levels (>100 ng/mg/24 hr) of PGE; the fast‐sticking epithelial fraction in primary cultures and the epithelial cell lines derived from it secrete 10–100 times less. Chromatography and radioisotopic labeling of the culture media from Rama 352 cells identify the PG as PGE2. PGE2 with insulin and hydrocortisone maximally stimulates [3H]DNA synthesis of epithelial cell lines and primary cultures from normal and tumorous glands by 2–4‐fold at concentrations (10–20 ng/ml) well below those released by the preadi‐pocytic stromal cells (20–100 ng/ml). Medium exposed to most cultured cells stimulates [3H]DNA synthesis of one epithelial cell line, Rama 25, by 2–4‐fold. Prevention of the synthesis of PGE2 in Rama 352 cultures with indomethacin or flurbiprofen abolishes the mitogenic activity present in the culture medium, and the PG receptor antagonist polyphloretin phosphate inhibits completely the mitogenic activity for Rama 25 cells. Myoepithelial‐like cell lines normally secrete moderate levels of PGE (10–100 ng/mg/24 hr) but the mitogenic activity for Rama 25 cells released from one such line, Rama 29, is not abolished by preventing the synthesis of PC's nor by PG‐receptor antagonists.
Evolution of cAMP phosphodiesterase activity in cultured myometrial cells: Effects of steroids and of successive subcultures
Authors: Vallet‐Strouve C. , Ferre F. , Breuiller M. 
Published: February 2005

AbstractCyclic AMP phosphodiesterase (PDE) activity was characterized in culture of ewe myometrial cells and its sensitivity to steroid hormones was tested. Cultured myometrial cells were maintained from the first to the 20th subculture in the presence of 2% of serum in a medium supplemented with 1 μM of insulin. It was found that myometrial cells possess a PDE activity with atypical kinetics. The nonlinear responses in Lineweaver‐Burke plots suggest the presence of high‐ and low‐affinity PDE activities. In cell culture, apparent Km values were similar to those obtained from the original myometrium. Vmax values increased with successive subcultures, revealing an increase in the capacity of the cells to degrade cAMP; in parallel, the growth rate decreased. The PDE specific activity in cultured myometrial cells was inhibited by estra‐diol or progesterone. When added together, no synergistic effect was obtained. The rate of inhibition for both steroids was constant during successive passages for both low‐ and high‐affinity conditions. Results obtained in myometrial cell long‐term culture were compatible with reports in other species in vivo. Considering the role of cAMP in the regulation of uterine functions, subcultured myometrial cells provided us a useful experimental system with which to study the cAMP metabolism process.
The intracellular pH of quiescent and proliferating human and rat thymic lymphocytes
Authors: Grinstein S. , Cohen S. , Lederman H. M. , Gelfand E. W. 
Published: February 2005

AbstractWe compared the cytoplasmic pH (pHi) of quiescent and actively cycling thymic lymphocytes. Human and rat thymocyte suspensions were fractionated by centrifugation on one step albumin density gradients. The pellet was composed of small, quiescent cells and the interphase contained mostly larger, actively cycling cells with a high proliferation index. When measured using [14C]‐dimethyloxazolidinedione (DMO), pHi in the large cells of both species was approximately 0.15 units more alkaline than in the small cells. However, these differences were not detectable when pHi was measured with carboxylated fluorescein derivatives generated in situ by cytoplasmic enzymes. This apparent discrepancy can be explained by compartmentation of DMO, which accumulates in the alkaline mitochondrial matrix. Comparison of the mitochondrial content of quiescent and cycling thymocytes by several methods showed that the latter contained over 2.5‐fold more mitochondria per unit cell volume. Assuming a constant intramitochondrial pH, this difference can account for the observed accumulation of DMO (i.e., apparent cytoplasmic alkalinity) in the actively proliferating cells. Therefore, no evidence was found for the existence of differences in pHi between quiescent and proliferating lymphocytes. Moreover, caution must be exercised when comparing DMO partition data in cells with varying relative mitochondrial content.
Amino acid activation of amino acid transport system N early in primary cultures of rat hepatocytes
Authors: Weissbach Lawrence , Kilberg Michael S. 
Published: February 2005

AbstractSystem N, a neutral amino acid transport system characterized in rat hepatocytes, shows significant changes in activity during the first 24 hr of primary culture (Weissbach, L., Handlogten, M.E., Christensen, H.N., and Kilberg, M.S. [1982] J. Biol. Chem. 257:12006–12011). Experiments presented in the present report demonstrate that during the first 12 hr of primary culture System N can be stimulated by individual amino acids in the culture medium by a cycloheximide‐insensitive mechanism. This enhanced activity results from an elevation in the Vmax of the transport system, and the magnitude of the increase is related to the concentration of the amino acid in the culture medium. Nonsubstrates as well as substrates of System N are effective, and trans‐stimulation does not appear to play a role in this phenomena. Transport by Systems ASC, Gly, and L is enhanced by the presence of amino acids in the culture medium, but these systems are significantly less sensitive than System N. The results suggest that amino acids act at a posttranslational step to activate System N activity.
2,3,7,8‐Tetrachlorodibenzo‐p‐dioxin: Examination of biochemical effects involved in the proliferation and differentiation of XB cells
Authors: Knutson Joyce C. , Poland Alan 
Published: February 2005

AbstractXB, a cell line derived from a mouse teratoma, differentiates into stratified squamous epithelium when incubated with 2,3,7,8‐tetrachlorodibenzo‐p‐dioxin (TCDD). To examine the possible biochemical mediators of this response, we compared the effects produced by TCDD to those elicited by other compounds which stimulate epidermal proliferation and/or differentiation in mice. XB/3T3 cultures keratinize when incubated with cholera toxin, epidermal growth factor (EGF), or TCDD, but not 12‐0‐tetradecanoylphorbol‐13‐acetate (TPA). Incubation of XB cells with TCDD (10−9M) for 48 hours produces a 20% increase in thymidine incorporation, a response which is neither as large nor as rapid as that produced by cholera toxin, TPA, or EGF. Although both cholera toxin and TCDD stimulate differentiation and thymidine incorporation in XB/3T3 cultures, cholera toxin increases cAMP 30‐fold in these cells, while TCDD does not affect cAMP accumulation at any of the times studies (15 min to 120 hours). Inhibitors of arachidonic acid metabolism, which block epidermal proliferative responses to TPA in vivo, do not prevent the differentiation of XB cell in response to TCDD. In XB/3T3 cultures, TPA stimulates arachidonic acid release at all times tested (1, 6, and 24 hours) and increases the incorporation of 32Pi into total phospholipids and phosphatidylcholine after 3 hours. In contrast, TCDD affects neither arachidonic acid release nor the turnover of phosphatidylinositol or phosphatidylcholine at any of the times tested. Although we examined biochemical effects which have been suggested as part of the mechanism of TCDD and which are produced by other epidermal proliferative compounds in XB cells, no mediator of the TCDD‐produced differentiation of XB/3T3 cultures was observed.
Pig reticulocytes. V. Development of Rb+ influx during in vitro maturation
Authors: Lauf P. K. , Zeidler R. B. , Kim H. D. 
Published: February 2005

AbstractInflux of the K+ analogue Rb+ was measured through the ouabain‐sensitive Na+/K+ pump and the ouabain‐insensitive “leak” pathways in Cl− or NO in mature red cells from adult pigs and in reticulocytes naturally occurring in 7‐day‐old piglets. In reticulocytes, Rb+ influxes by the two pathways were of about equal magnitude in Cl− (13 and 10 mmoles/liter cells × hr) and at least 25‐fold larger than in mature red cells (0.5 and 0.4 mmoles/liter cells × hr). In Na + media, a portion of the ouabain‐insensitive “leak” flux of Rb+ was Cl− dependent (Rb+Cl− transport) as NO replacement reduced Rb+ influx by 90% in reticulocytes and by 40% in mature red cells. The sulfhydryl reagent N‐ethylmaleimide (NEM) stimulated Rb+Cl− transport about twofold in reticulocytes and up to 13‐fold in mature red cells. When reticulocytes matured to erythrocytes during in vitro incubation, about 90% of both ouabain‐sensitive Rb+ pump and ouabain‐insensitive Rb+Cl− influx were lost. In contrast, the NEM‐stimulated Rb+Cl− transport changed much less throughout this period, suggesting an entity operationally but not necessarily structrually distinct from the basal Rb+Cl− transport. Although the experimental variability precluded a full assessment of significant changes in the small Na+/K+(Rb+) pump and Rb+Cl− fluxes in mature pig red cells kept for the same time period in vitro, Rb+ flux changes in reticulocytes appear to be maturational in nature, reflecting parallel activity transitions of Na+/K+ pump and Cl−‐dependent K+ fluxes in vivo.
Comparison of intracellular and extracellular mitogenic activity
Authors: Gajdusek Corinne M. , Schwartz Stephen M. 
Published: February 2005

AbstractEndothelial cell‐derived growth factor (ECDGF) is a soluble mitogen secreted in vitro by bovine aortic endothelium. ECDGF is a mixture of at least two distinct heat‐stable and trypsin‐sensitive mitogens. Large amounts of mitogenic activity were found in lysates prepared from cultured endothelial cells. Other nonmitogen‐secreting cells in culture, including bovine dermal fibroblasts and vascular smooth muscle cells, also contained a similar activity. In contrast to ECDGF, the lysate mitogenic activities were sensitive to heat (56°C) and were not inactivated by trypsin. Similar to platelet‐derived growth factor (PDGF), ECDGF and cell lysate mitogens promoted cell proliferation in the absence of other defined mitogens when added to culture medium and after exposure to plastic. The cytoplasmic mitogens, however, were distinct from PDGF by receptor competition assays and other criteria.
Maturation of human promyelocytic leukemia cells induced by nicotinamide: Evidence of a regulatory role for ADP‐ribosylation of chromosomal proteins
Authors: Lucas Diane L. , Tanuma Sei‐Ichi , Davies Peter J. A. , Wright Daniel G. , Johnson George S. 
Published: February 2005

AbstractWe have studied the role of ADP‐ribosylation of chromosomal proteins in the regulation of myeloid cell maturation using the HL‐60 cell line as a model. Nuclei isolated from this human promyelocytic leukemia cell line contained (ADP‐ribose)n synthetase activity, whereas little or no enzymatic activity was detectable in normal human blood neutrophils. Furthermore, the activity of (ADP‐ribose)n synthetase was decreased in HL‐60 cells when they were induced to mature with retinoic acid (RA). To determine whether reduced (ADP‐ribose)n synthetase activity is simply a result of induced maturation or whether it is a necessary precedent event for the maturation process, we evaluated the effects of nicotinamide (NAm) and its methyl derivative, N′‐methylnicotinamide (N′‐Met‐NAm), agents which decrease ADP‐ribosylation. Treatment of HL‐60 cells with these drugs caused the cells to undergo maturation and to acquire certain of the morphologic, functional, and biochemical characteristics of normal neutrophils. N′‐Met‐NAm was more potent than NAm in inducting maturation; at a concentration of 0.8 mM, it caused greater than 80% of the cells to mature, whereas a tenfold greater concentration of NAm was required to induce a similar degree of maturation. NAm and N′‐Met‐NAm also potentiated the maturation of HL‐60 cells induced by RA. Exposure of cells to noninducing concentrations of these compounds caused a leftward shift in the dose‐response curve for RA; maturation was observed at 10−11 M RA in the presence of either 2 mM NAm or 0.2 mM N′‐Met‐NAm while 10−9 M RA was required to induce maturation in their absence. A leftward shift in the dose response curve for maturation in the presence of low doses of NAm or N′‐Met‐NAm did not occur with another inducer, dimethyl formamide (DMF). Two enzymes, NAD glycohydrolase and tissue transglutaminase, that are abundant in macrophages, were induced by RA but not by NAm. N′‐Met‐NAm decreased by about 75% the amount of endogenous (ADP‐ribose)n in a selected fraction of chromosomal proteins which included histone H1 and the nonhistone high mobility group proteins. The results of this study support the concept that ADP‐ribosylation of chromosomal proteins influences the regulation of human myeloid cell maturation.