bluuebunny/crossref_metadata_2025_split huggingface dataset:
Proliferation and differentiation of abelson virus‐infected murine myeloid leukemia cell lines does not involve an autocrine mechanism
Authors: Hines David L. 
Published: February 2005

AbstractCulture in agar of cloned promonocytic leukemia cell lines derived from Abelson virus‐infected mice produced colonies of both a compact and diffuse morphology. Diffuse colonies contained fewer cells capable of forming colonies when recultured in agar than did compact colonies. Serial subcloning of cells from diffuse, but not compact, colonies ultimately led to the complete loss of colony‐forming cells, i.e., to clonal extinction. The production of both compact and diffuse agar colonies was independent of the cell density of either the static liquid culture from which cells were taken for culture in agar, or the number of cells per agar culture. Furthermore, bioassays of culture supernatants indicated the leukemia cells did not secrete hemopoietic growth factors active on normal hemopoietic cells, transforming growth factors active on adherent cell lines, or factors that influenced the growth of the leukemic cells themselves. Collectively, these data suggest neither growth‐factor independent replication nor the spontaneous differentiation of Abelson virusinfected myeloid cells involves autocrine secretion of growth regulators.
Delayed processing/export of messenger RNA under conditions of reduced protein synthesis
Authors: Muralidhar M. G. , Johnson L. F. 
Published: February 2005

AbstractThe rates of processing and export of a variety of nuclear RNA species into the cytoplasmic compartment were studied by determining the rates of incorportion of tritiated uridine into nuclear and cytoplasmic RNA species. In exponentially growing cells, the rates of nuclear processing/export varied by more than a factor of ten for the six different mRNA species that were examined. Differences in the rates did not appear to be correlated with either the number or the sizes of introns in the genes for the RNA species. When cells were maintained under conditions of reduced protein synthesis (starvation for isoleucine and glutamine or exposure to cycloheximide), the processing rates for each species decreased by a factor of about 3. The decrease was not caused by the inability of hnRNA to associate with proteins, since the nuclear RNP distribution appeared normal in amino acidstarved cells.
Induction of system A amino acid transport through long‐term treatment with ouabain: Correlation with increased (Na+/K+)‐ATpase activity
Authors: Schenerman Mark A. , Leister Kirk J. , Trachtenberg David K. , Racker Efraim 
Published: February 2005

AbstractMouse embryo fibroblast cells (C3H‐10T½) and the methylcholanthrenetransformed derivative (MCA‐10T½) were treated with basal modified Eagle's medium (BME) containing 10% fetal bovine serum and varying concentrations of ouabain ranging from 0.05 mM to 0.7 mM for 16 h in culture. After replacing the ouabain‐containing medium with Earl's balanced salts solution, System A amino acid transport activity increased from approximately 40 to 500 pmol AIB accumulated · mg protein−1 · min−1 in the C3H‐10T½ cells and from approximately 300 to 700 pmol AIB accumulated · mg protein−1 · min−1 in the MCA‐10T½ cells. The (Na+/K+)‐ATPase pump activity also increased form approximately 12 to 46 nmol Rb+ accumulated · mg protein−1 · min−1 in the normal cells and from approximately 20 to 42 nmol Rb+ accumulated · mg protein−1 · min−1 in the transformed cells. System A and the (Na+/K+) ATPase activity were maximally increased at approximately0.4–0.6 mM ouabain in the normal cells in contrast to the transformed cells which were maximally stimulated at a concentration of approximately 0.2 mM ouabain. This treatment with ouabain increased the [Na+]i/[K+]i as measured by atomic absorption spectroscopy, and thereby decreased the Na+ and K+ electrochemical gradients. Our data show that the internal ion gradients inverted at a lower concentration of ouabain in the transformed cells compared to the normal cells. The ouabain‐induced increased in pump and System A activity shown here was used as a tool to further investigate the coordinated ion transport regulation in the control of cell growth.
Studies on the increase in cytosolic free calcium induced by epidermal growth factor, serum, and nucleotides in individual A431 cells
Authors: Gonzalez Fernando A. , Gross David J. , Heppel Leon A. , Webb Watt W. 
Published: February 2005

AbstractThe response of cytosolic calcium [Ca2+]i to epidermal growth factor (EGF), fetal calf serum, and nucleotides was determined in individual A431 cells, using the fluorescent probe fura‐2 and quantitative digital video fluorescence microscopy. In the presence of 1 mM external Ca2+, EGF caused a rapid rise in [Ca2+]i, followed by a slower and variable decrease. The cells responded after a lag that varied from 10 to 30 seconds, and there was considerable cell‐to‐cell variation in extent of the rise in [Ca2+]i. A second challenge with EGF gave negative results. No response was obtained in nominally Ca2+‐free medium supplemented with 100 μM EGTA. Somewhat similar results were obtained with fetal calf serum except that a rise in [Ca2+]i was observed both in the presence and absence of external Ca2+. The A431 cells responded to external ATP with a rise in [Ca2+]i in less than 10 seconds, both in Ca2+‐containing and Ca2+‐free media. A coverslip with attached cells was mounted on a small chamber, allowing complete change of medium in 2 seconds. A nearly full response was obtained with only 10 seconds of contact of cells with ATP‐containing medium. After washing out ATP, there was little or no response to a second addition given 100 seconds after the first. However, a second response was obtained when the concentration of agonist was increased 10–20‐fold. These data favor the idea of receptor desensitization. Both homologous and heterologous receptor desensitization was observed. A transient rise in [Ca2+]i was also noted with UTP, while ITP and CTP were inactive.
Methotrexate‐poly(lysine) as a selective agent for mutants of chinese hamster ovary cells defective in endocytosis
Authors: Ryser Hugues J.‐P. , Mandel Richard , Hacobian Asteghik , Shen Wei‐Chiang 
Published: February 2005

AbstractMethotrexate (MTX) covalently linked to poly(L‐lysine) [poly(Lys)] enters cells by endocytosis, is degraded in lysosomes and, upon liberation of small molecular methotrexate, is cytocidal to Chinese hamster cells in culture. This drug conjugate was used to select mutants resistant to MTX‐poly(Lys), which were examined for defects in endocytosis. Two mutants resistant to MYX‐poly(Lys) and sensitive to free MTX, MPL 3–4 and MPL 2–5, internalized the conjugate in normal fashion, but had a decreased ability to degrade it to small molecular drug. The magnitude of this defect in the two mutants correlated with their level of resistance. In addition, both mutants were cross resistant to diphtheria toxin and modeccin and hypersensitive to ricin. While MPL 3–4 internalized MTX‐poly(Lys) and inulin normally, it showed decreased endocytosis via the mannose‐6‐phosphate receptor and decreased uptake of 125I‐alpha‐2 macroglobulin. Acidification of subcellular fractions was measured using the partitioning of acridine orange. In MPL 3–4, the ATP‐driven acidification of the endosome‐containing cell fractions was slightly decreased (80% of controls), while acidification of the heavy lysosome‐containing fraction was normal. Complementation analysis using hybrids of MPL 3–4 × MPL 2–5 indicated that the mutations occurred at the same gene, but were expressed with different severity. This genotype is identical to that of the End 2 mutants described by Roff et al. (1986). Thus, surprisingly, mutants with identical genotypes were isolated independently by totally different selection procedures.
Inhibition of hexose transport by adenosine derivatives in human erythrocytes
Authors: May James M. 
Published: February 2005

AbstractThe human erythrocyte membrane carriers for hexoses and nucleosides have several structural features in common. In order to assess functional similarities, the effects of adenosine derivatives on hexose transport and cytochalasin B binding sites were studied. Adenosine inhibited zero‐trans uptake of 3‐0‐methylglucose half‐maximally at 5 mM, while more hydrophobic adenosine deaminase‐resistant derivatives were ten‐ to 20‐fold more potent transport inhibitors. However, degradation of adenosine accounted for very little of this difference in potency. Hexose transport was rapidly inhibited by N6‐(L‐2‐phenylisopropyl)adenosine at 5°C in a dose‐dependent fashion (EC50 = 240 μM), to lower the transport Vmax without affecting the Km. A direct interaction with the carrier protein was further indicated by the finding that N6‐(L‐2‐phenylisopropyl)adenosine competitively inhibited [3H]cytochalasin B binding to erythrocytes (Ki = 143 μM) and decreased [3H]cytochalasin B photolabeling of hexose carriers in erythocyte ghosts. The cross‐reactivity of adenosine and several of its derivatives with the hexose carrier suggests further homologies between the carriers for hexoses and nucleosides, possibly related to their ability to transport hydrophilic molecules through the lipid core of the plasma membrane.
Differential expression of c‐myc and the transferrin receptor in G1 synchronized M1 myeloid leukemia cells
Authors: Neckers Leonard M. , Tsuda Hiroyuki , Weiss Ervin , Pluznik Dov H. 
Published: February 2005

AbstractWe have studied the transcriptional activity, steady‐state mRNA levels, and steady‐state protein levels of the c‐myc and transferrin receptor (TfR) genes in murine M1 myeloid leukemia cells arrested in G1 phase of the cell cycle by different methods. When cells are growth‐arrested by density inhibition, a technique that places the majority of cells in early G1, c‐myc protein, as detected by Western analysis, is expressed at 80% of the level seen in proliferating cells. Steady‐state mRNA levels and, to a lesser extent, transcriptional activity of the c‐myc gene, parallel the protein findings. Under these conditions, TfR gene expression is much lower than in normally cycling cells. We have previously demonstrated that density‐inhibited M1 cells, released from density inhibition and treated with the DNA polymerase alpha inhibitor aphidicolin, remain in G1, but at a point temporally closer to S phase. Cells treated in this manner demonstrate reduced transcriptional activity and expression of the c‐myc gene, but TfR gene expression approximates the level found in proliferating cells. These data suggest that neither c‐myc nor TfR gene expression is constant throughout the G1 phase of the cell cycle in M1 cells. c‐myc gene expression is highest in early G1 and falls to low levels by late G1, while the reverse is true for TfR gene expression.
Effects of human neutrophil chemotaxis across human endothelial cell monolayers on the permeability of these monolayers to ions and macromolecules
Authors: Huang Ada J. , Furie Martha B. , Nicholson Susan C. , Fischbarg Jorge , Liebovitch Larry S. , Silverstein Samuel C. 
Published: February 2005

AbstractWe have developed a method for studying the permeability properties of human endothelia in vitro. Human umbilical vein endothelial cells (HUVEC) were cultured on a substrate of human amnion. Confluent monolayers of these cells demonstrated 6–12 ωcm2 of electrical resistance (a measure of their permeability to ions) and restricted the transendothelial passage of albumin from their apical to their basal surface. To determine whether leukocyte emigration alters endothelial permeability in this model, we examined the effects of migrating human polymorphonuclear leukocytes (PMN) on these two parameters. Few PMN migrated across the HUVEC monolayers in the absence of chemoattractants. In response to chemoattractants, PMN migration through HUVEC monolayers was virtually complete within 10 minutes and occurred at random locations throughout the monolayer. PMN migrated across the monolayer via the paracellular pathway. Although one PMN migrated across the monolayer for each HUVEC, PMN migration induced no change in electrical resistance or albumin perme‐ability of these monolayers. At this PMN:HUVEC ratio, these permeability findings were correlated morphologically to measurements that HUVEC paracellular pathway size increases by less than 0.22% with PMN migration. This increase is insufficient to effect a measurable change in the electrical resistance of the endothelial cell monolayer. These findings demonstrate that increased permeability of cultured endothelial cell monolayers is not a necessary consequence of PMN emigration.
Acetyl‐LDL stimulates macrophage‐dependent plasminogen activation and degradation of extracellular matrix
Authors: Falcone Domenick J. , Ferenc Michael J. 
Published: February 2005

AbstractThe ability of acetyl‐LDL to stimulate macrophage‐dependent plasminogen activation and degradation of extracellular matrix was examined. We have found that expression of plasminogen activator activity in response to the scavenger receptor ligand varied among cell populations. Exposure to acetyl‐LDL stimulated plasminogen activator expression by cells which constitutively released low levels of activator. These include a virally transformed macrophage‐like cell line (RAW246.7), concanavalin A and C. parvum‐activated macrophages. The stimulation of plasminogen activator activity was independent of cellular lipid accumulation since nonlipoprotein inhibitors of acetyl‐LDL binding to the scavenger receptor stimulated activator expression in great excess to that observed with acetyl‐LDL. In contrast, acetyl‐LDL was unable to induce soluble plasminogen activator activity in cells which normally do not express it. These include a macrophage‐like cell line (J774A.1) and resident peritoneal macrophages. Furthermore, acetyl‐LDL was unable to modulate the copious secretion of activator by inflammatory macrophages elicited with thioglycolate. When macrophages were tested for their ability to degrade smooth muscle cell derived matrix, solubilization by resident, elicited, and activated cells was variously increased in the presence of plasminogen. Furthermore, exposure to acetyl‐LDL enhanced plasmin‐dependent degradation by resident cells and activated cells, whereas matrix degradation by elicited cells was unaffected.
Binding of diphtheria toxin to CHO‐K1 and vero cells is dependent on cell density
Authors: Schaefer Erik M. , Moehring Joan M. , Moehring Thomas J. 
Published: February 2005

AbstractWe studied the binding of 125I‐labeled diphtheria toxin (DTX) to receptors on monolayer cultures of Chinese hamster ovary cells (CHO‐K1) and Vero cells. The number of DTX receptors detected on the cell surface was shown to be dependent on the cell density (number of cells per unit area). Cells at low density (≤23,000 cells per cm2 for CHO‐K1 cells; ≤80,000 cells per cm2 for Vero cells) had more receptors for DTX than cells at higher densities. The difference in receptor number between low‐ and high‐density cells was 33‐fold for CHO‐K1 cells and 19‐fold for Vero cells. We estimated the maximum number of DTX receptors on low‐density CHO‐K1 and Vero cells to be 50,000 and 370,000 per cell, respectively. The cell density at which the binding of DTX was reduced to 50% of maximum was considerably lower for CHO‐K1 cells than for Vero cells (33,000 vs. 220,000 cells per cm2, respectively). Vero cells grown on a surface that had been conditioned by high‐density cells bound less DTX, suggesting that interaction of these cells with the underlying extracellular matrix might regulate the number of cell surface receptors for DTX. Low‐density cells were more sensitive to DTX than high‐density cells, suggesting that low‐density cells possessed an increased number of functional receptors that actively transported DTX to the cytosol. CHO‐K1 and Vero cells were equally protected by SITS (4‐Acetamido‐4prime;‐Isothiocyano‐Stilbene‐2,2′‐disulfonic Acid), a compound that has been shown to inhibit the binding and entry of DTX in Vero cells, suggesting that intoxication of CHO‐K1 and Vero cells is mediated by a similar mechanism. The data illustrate the importance of taking into account the cell density when measuring the number of DTX receptors on adherent cells.
Plasminogen activation and lysis of blood clots induced by cells in vitro
Authors: Knox Peter , Crooks Sheila 
Published: February 2005

AbstractWhen human diploid fibroblasts were seeded onto the surface of blood clots, lysis of the clot occurred as a result of the release of cellular plasminogen activator. A number of aspects of this lysis were studied. 1. There was no significant difference in rates of lysis of whole blood clots, platelet‐rich plasma clots, and platelet‐poor plasma clots brought about by the same number of fibroblasts. 2. Clot lysis was promoted by nondividing cells and by proliferating cells. 3. Using cycloheximide to block protein synthesis it was found that the plasminogen activator released by fibroblasts had an active half‐life of less than an hour. 4. When clots were washed prior to the addition of cells then lysis occurred at an increased rate. This was probably due to the removal of α2‐antiplasmin from the clots, since when antisera to α2‐antiplasmin was added to clots, lysis also proceeded at an increased rate. 5. Medium conditioned by fibroblasts did not promote clot lysis even when antiplasmin was removed by washing or by addition of antisera. 6. Cells had to be in direct contact with the clot in order to bring about lysis; when cells were separated from clots by permeable membranes there was no lysis. 7. When cross‐linking of fibrin was reduced by the inhibition of transglutaminase, the rate of clot lysis was increased.
DNA synthesis in U‐2 OS human osteosarcoma cells is independent of PDGF binding to functional cell surface receptors
Authors: Richter Michael R. , Graves Dana T. 
Published: February 2005

AbstractPrevious studies have shown that suramin reveals specific PDGF binding sites on U‐2 OS human osteosarcoma cells. Stud es presented here indicate that U‐2 OS cells pretreated with suramin internalize and degrade 125l‐PDGF and respond to PDGF by increased tyrosine kinase activity and amino acid transport. However, DNA synthesis in these cells is not reduced by incubation with the PDGF blocking agent suramin and is not stimulated by exogenous PDGF. These data indicate that U‐2 OS cells possess functional PDGF receptors but that high levels of DNA synthesis in these cells is unrelated to the binding of secreted PDGF to these cell surface receptors. Thus, it is unlikely that the PDGF mitogen produced by U‐2 OS cells stimulates proliferation through an autocrine mechanism involving secretion and subsequent binding to PDGF receptors.
Regulation of murine plasmacytoma transferrin receptor expression and G1 traversal by plasmacytoma cell growthfactor
Authors: Neckers L. M. , Nordan R. P. 
Published: February 2005

AbstractMany plasmacytomas arising in BALB/c mice require a specific, macrophage‐derived growth factor in order to proliferate in vitro. Since transferin receptor expression is normally regulated by tissue‐specific growth factors and because expression of these receptors is required for cell proliferation, we examined the interaction of plasmacytoma growth factor (PCT‐GF) on transferrin receptor expression and cell cycle progression in several PCT‐GF‐dependent and independent plasmacytoma cell lines maintained in vitro. We found that removal of PCT‐GF results in a rapid and specific loss of transferrin receptor expression with concomitant G1 arrest in early G1. The time required for G1 arrest to become maximal correlates closely to the initial level of surface transferrin receptor expression and the rate of decay following removal of PCT‐GF. The calcium channel blocker diltiazem interferes with the ability of PCT‐GF to maintain transferrin receptor expression in PCT‐GF‐dependent cell lines and causes a G1 arrest of the cell population. When added to a PCT‐GF‐independent cell line, diltiazem also inhibited transferrin receptor expression and caused G1 arrest. Thus, both PCT‐GF‐dependent and ‐independent plasmacytoma cell lines require transferrin receptor expression for growth. In factor dependent cell lines, transferrin receptor expression requires exogenous PCT‐GF, while in factor‐independent cells, transferrin receptor expression is constitutive. In both cell types, intracellular calcium levels may play a role in receptor expression.
Epidermal growth factor‐dependent growth of human KB cells in a defined medium and altered growth factor requirements of KB mutants resistant to EGF‐Pseudomonas exotoxin conjugates
Authors: Amano Fumio , Gottesman Michael M. , Pastan Ira 
Published: February 2005

AbstractA serum‐free culture system was established for human KB carcinoma (HeLa) cells that consisted of a chemically defined medium and several growth factors including epidermal growth factor (EGF), insulin, transferrin, hydrocortisone, and ethanolamine. EGF and insulin showed the greatest effects on the growth rate of KB cells. Insulin‐like growth factor l (IGF‐l) at the same concentration as insulin stimulated cell growth less than insulin. Transferrin, hydrocortisone, or ethanolamine had no growth‐stimulatory effects alone but were stimulatory when combined with EGF and/or insulin. Transforming growth factor‐beta inhibited growth and triiodothyronine stimulated growth. The growth factor requirements were established for several KB mutants with low EGF receptor levels that had been selected for resistance to a conjugate of EGF with Pseudomonas exotoxin (EGF‐PE). Three of five KB mutants did not respond to EGF; two other mutants responded to a lesser extent than the parental KB cells. Four mutants had a reduced response to insulin and responded to T3; one mutant (ET‐30) responded to neither. These results indicate that KB cells selected for EGF‐PE resistance have lost their growth response to EGF and illustrate the usefulness of serum‐free medium for studying the growth factor requirements of mutants with altered receptor levels.
Desensitization of adenylate cyclase and cyclic AMP flux during the early stages of liver regeneration
Authors: Mahler Stephen M. , Wilce Peter A. 
Published: February 2005

AbstractLiver regeneration is controlled by a complex network of interactions between hormones, growth factors, and a variety of hepatotrophic factors. Transient increases in cAMP in the early stages of liver regeneration that are necessary for DNA synthesis and subsequent mitosis have been reported; however, studies on the mechanisms that control cellular cAMP levels during liver regeneration, namely adenylate cyclase activity, cAMP‐dependent phosphodiesterase activity, and cAMP efflux from the cell, have been generally incomplete. In this study we have shown that although there are three peaks in intracellular cAMP levels in the first 24 hours after partial hepatectomy, the adenylate cyclase activity stimulated by glucagon, prostaglandin E2, adrenaline, and fluoride in vitro decreases with time. Kd and BMAX of hepatocyte glucagon and β receptors were similar to the sham controls. Our results are consistent with a mixed homologous/heterologous desensitization of the adenylate cyclase system. There was also a loss of cAMP‐dependent phosphodiesterase activity after partial hepatectomy. We speculate that even though the hormone‐stimulated adenylate cyclase system has been desensitized, the system retains the ability to respond to the transient pulses of the variety of hormones secreted after partial hepatectomy and thus raise the intracellular concentration of cAMP. The decrease in cAMP‐dependent phosphodiesterase may be necessary to prevent rapid breakdown of cAMP.