bluuebunny/crossref_metadata_2025_split huggingface dataset:
GM‐CSF triggers a rapid, glucose dependent extracellular acidification by TF‐1 cells: Evidence for sodium/proton antiporter and PKC mediated activation of acid production
Authors: Wada H. Garrett , Indelicato Stephen R. , Meyer Lorraine , Kitamura Toshio , Miyajima Atsushi , Kirk Gregory , Muir Victoria C. , Parce J. Wallace 
Published: February 2005

AbstractThe extracellular acidification rate of the human bone marrow cell line, TF‐1, increases rapidly in response to a bolus of recombinant granulocyte‐macrophage colony stimulating factor (GM‐CSF). Extracellular acidification rates were measured using a silicon microphysiometer. This instrument contains micro‐flow chambers equipped with potentiometric sensors to monitor pH. The cells are immobilized in a fibrin clot sandwiched between two porous polycarbonate membranes. The membranes are part of a disposable plastic “cell capsule” that fits into the microphysiometer flow chamber. The GM‐CSF activated acidification burst is dose dependent and can be neutralized by pretreating the cytokine with anti‐GM‐CSF antibody. The acidification burst can be resolved kinetically into at least two components. A rapid component of the burst is due to activation of the sodium/proton antiporter as evidenced by its elimination in sodium‐free medium and in the presence of amiloride. A slower component of the GM‐CSF response is a consequence of increased glycolytic metabolism as demonstrated by its dependence on D‐glucose as a medium nutrient. Okadaic acid (a phospho‐serine/threonine phosphatase inhibitor), phorbol 12‐myristate 13‐acetate (PMA, a protein kinase C (PKC) activator), and ionmycin (a calcium ionophore) all produce metabolic bursts in TF‐1 cells similar to the GM‐CSF response. Pretreatment of TF‐1 cells with PMA for 18 h resulted in loss of the GM‐CSF acidification response. Although this treatment is reported to destroy protein kinase activity, we demonstrate here that it also down‐regulates expression of high‐affinity GM‐CSF receptors on the surface of TF‐1 cells. In addition, GM‐CSF driven TF‐1 cell proliferation was decreased after the 18 h PMA treatment. Short‐term treatment with PMA (1–2h) again resulted in loss of the GM‐CSF acidification response, but without a decrease in expression of high‐affinity GM‐CSF receptors. Evidence for involvement of PKC in GM‐CSF signal transduction was obtained using calphostin C, a specific inhibitor of PKC, which inhibited the GM‐CSF metabolic burst at a subtoxic concentration. Genistein and herbimycin A, tyrosine kinase inhibitors, both inhibited the GM‐CSF response of TF‐1 cells, but only at levels high enough to also inhibit stimulation by PMA. These results indicate that GM‐CSF activated extracellular acidification of TF‐1 cells is caused by increases in sodium/proton antiporter activity and glycolysis, through protein kinase signalling pathways which can be both activated and down‐regulated by PMA. © 1993 Wiley‐Liss, Inc.
Polarized secretion of IGF‐I and IGF‐I binding protein activity by cultured aortic endothelial cells
Authors: Taylor W. Robert , Nerem Robert M. , Alexander R. Wayne 
Published: February 2005

AbstractInsulin‐like growth factor‐I (IGF‐I) secretion by the vascular endothelium has been proposed to play a role in the regulation of vascular smooth muscle cell proliferation. Because vascular smooth muscle cells are adjacent to the abluminal surface of the endothelium, we tested the hypothesis that secretion of IGF‐I by endothelial cells is polarized. Porcine aortic endothelial cells were cultured on permeable membranes and IGF‐I measured by radioimmunoassay. Basal secretion exceeded apical secretion by a ratio of 2.3 ± 0.2:1.0 (P < 0.05). We also identified 35 kDa IGF‐I binding protein activity that is preferentially secreted on the basal surface of endothelial cells. We conclude that both IGF‐I and IGF‐I binding protein activity secretion by endothelial cells is polarized towards the basal surface of the endothelium. A polarized secretion mechanism for IGF‐I may be of importance in the normal growth and differentiation of the vasculature as well as in the development of vascular pathology. © 1993 Wiley‐Liss, Inc.
Cell cycle‐dependent gene expression in V point‐arrested BALB/c‐3T3 cells
Authors: Olson James E. , Winston Jeffrey T. , Whitlock James A. , Pledger W. J. 
Published: February 2005

AbstractDensity‐arrested BALB/c‐3T3 cells stimulated to proliferate in an amino acid‐deficient medium arrest in mid‐G1 at a point termed the V point. Cells released from V point arrest require 6 hr to traverse late G1 and enter S phase. As data presented here show that mRNA synthesis is needed for 2–3 hr after release of cells from the V point, after which inhibition of mRNA synthesis does not prevent entry into S phase, we used this mid‐G1 arrest protocol to analyze gene expression in late G1. We found that although stimulation of cells in amino acid‐deficient medium did not inhibit the induction of genes expressed in early G1, genes normally expressed in late G1 were expressed only after release from the V point. The expression of late G1 genes in cells released from the V point was temporally similar, in respect to G1 location, as was seen in stimulation of quiescent Go cells. As this protocol effectively divides gene expression into early (pre‐V point) and late (post‐V point) categories, it should be useful in studies of growth factor‐modulated events that regulate traverse of late G1 and commitment to DNA synthesis. In addition, we used c‐myb antisense oligonucleotides to show that c‐myb expression, which occurs in late G1, is required for BALB/c‐3T3 fibroblasts to traverse late G1 and initiate DNA synthesis. © 1993 Wiley‐Liss, Inc.
Orthovanadate both mimics and antagonizes the transforming growth factor β action on normal rat kidney cells
Authors: Rijksen Gert , Völler Maureen C. W. , Van Zoelen Everardus J. J. 
Published: February 2005

AbstractNormal rat kidney [NRK] cells grown in the presence of epidermal growth factor (EGF) or platelet‐derived growth factor (PDGF) have a normal phenotype and undergo density‐dependent growth inhibition, whereas in the presence of multiple growth factors, density arrest is lost and the cells become phenotypically transformed. We studied the influence of the protein tyrosine phosphatease (PTPase) inhibitor sodium orthovanadate on the mitogenic stimulation of NRK cells by growth factors and on transformation‐linked properties as loss of density‐dependent growth inhibition and anchorage‐independent growth. The fraction of cells in serum‐deprived monolayer cultures that is induced to proliferate upon mitogenic stimulation by EGF or PDGF is only slightly enhanced upon addition of low concentrations (25–50 μM) of vanadate. Addition of vanadate per se induces proliferation of only a very limited amount of cells, but results in a shift of the dose‐response curves for other growth factors to lower concentrations. Vanadate added in combination with EGF or PDGF is able to mimic the effect of transforming growth factor β (TGFβ) in inducing phenotypic transformation. In monolayer cultures density‐dependent growth inhibition is lost and anchorage‐independent proliferation is observed on dishes coated with poly(2‐hydroxy‐ethyl methacrylate) (polyHEMA). The extent of these changes is similar to that induced by TGFβ. However, the morphology of the obtained colonies in polyHEMA‐coated dishes is quite different. Cells transformed by TGFβ in the presence of EGF form rather amorphous colonies, whereas in the presence of orthovanadate colonies are formed that tend to fall apart in loose cells. The effect of vanadate on cell transformation is dependent on the growth factor conditions in a bimodal way. When a suboptimal dose of growth factor(s) is used, 25 μM vanadate is very effective in preventing density‐induced growth inhibition and stimulating anchorage‐independent proliferation. However, the same concentration of vandate is inhibitory when cells are maximally stimulated and antagonizes the transforming effect of TGFβ added in combination with other growth factors. It is hypothesized that vanadate acts on a set of different protein tyrosine phosphatases. Some of these are positive and others negative regulators of growth. © 1993 Wiley‐Liss, Inc.
Intracellular free sodium concentrations in GH4C1 cells
Authors: Törnquist Kid , Ekokoski Elina 
Published: February 2005

AbstractIn the present investigation, intracellular sodium ([Na+]i) levels were determined in GH4C1 cells using the fluorescent probe SBFI. Fluorescence was determined by excitation at 340 nm and 385 nm, and emission was measured at 500 nm. Intracellular free sodium ([Na+]i) was determined by comparing the ratio 340/385 to a calibration curve. The ratio was linear between 10 and 60 mM Na+. Resting [Na+]i in GH4C1 cells was 26 ± 6.2 mM (mean ± SD). In cells incubated in Na+‐buffer [Na+]i decreased to 3 ± 3.6 mM. If Na+/K+ ATPase was inhibited by incubating the cells with 1 mM ouabain, [Na+]i increased to 47 ± 12.8 mM in 15 min. Stimulating the cells with TRH, phorbol myristyl acetete, or thapsigargin had no effect on [Na+]i. Incubating the cells in Ca2+‐buffer rapidly increased [Na+]i. The increase was not inhibited by tetrodotoxin. Addition of extracellular Ca2+, nimodipine, or Ni2+ to these cells immediately decreased [Na+]i, whereas Bay K 8644 enhanced the influx of Na+. In cells where [Na+]i was increased the TRH‐induced increase in intracellular free calcium ([Ca2+]i) was decreased compared with control cells. Our results suggest that Na+ enters the cells via Ca2+ channels, and [Na+]i may attenuate TRH‐induced changes in [Ca2+]i in GH4C1 cells. © 1993 Wiley‐Liss, Inc.
Parathyroid hormone‐induced changes in alkaline phosphatase expression in fetal calvarial osteoblasts: Differences between rat and mouse
Authors: Jongen J. W. J. M. , Bos M. P. , Van Der Meer J. M. , Herrmann‐Erlee M. P. M. 
Published: February 2005

AbstractWe studied the effects of parathyroid hormone (PTH) on two markers of the osteoblast phenotype: alkaline phosphatase (AP) (activity and mRNA) and cyclic adenosine monophosphate (cAMP) accumulation. Osteoblast‐like cells derived from fetal rat (ROB) and mouse (MOB) calvariae were isolated by collagenase treatment. Cells were cultured in α‐Minimal Essential Medium (MEM) with 2% fetal calf serum (FCS) for 4 days. In ROB and MOB bPTH(1‐34) induced a fast increase (up to 5 minutes) in cAMP accumulation. When equal amounts of cells were seeded, the cAMP accumulation was higher in MOB than in ROB. No difference in basal AP activity was observed between ROB and MOB. When bPTH(1‐34) was added to ROB for the last 24 or 48 hr, AP activity decreased dose dependently. However, MOB treated with bPTH(1‐34) for the last 24 or 48 hours showed an increase of AP activity. Basal AP activity was positively correlated with the seeding density of ROB and MOB cultures. Basal AP activity influenced the degree of inhibition (ROB) or stimulation (MOB) after incubation with bPTH(1‐34). © 1993 Wiley‐Liss, Inc.
Effects of growth factors on cell cycle arrest in dolichyl phosphate‐depleted cultures
Authors: Doyle J. W. , Ward‐Bailey P. F. , Kandutsch A. A. 
Published: February 2005

AbstractPreviously we showed that CHO cell growth is arrested in the G1 or G0 phase within 24 h after the biosynthesis of mevalonic acid is blocked. The growth‐limiting factor under these conditions appeared to be dolichyl phosphate or one of its glycosylated derivatives with consequent decrease in the synthesis of N‐linked glycoproteins (Doyle, J.W., and A.A. Kandutsch, 1988, J. Cell Physiol. 137:133–140; Kabakoff, B., J.W. Doyle, and A.A. Kandutsch, 1990, Arch. Biochem. Biophys. 276:382–389). We show herein that cell surface glycoproteins are depleted in the inhibited cultures and that growth arrest is delayed when supraphysiological concentrations of insulin, insulin‐like growth factor‐1 (IGF‐1) and bFGF are added to the culture medium. Apparently an elevated level of a growth factor increases the length of time during which a threshold level of occupied receptor is maintained as the number of glycosylated receptor molecules declines. The results support the idea that cellular levels of dolichyl phosphate and its derivatives may limit cell division by controlling the numbers of functional receptors for growth factors and of other glycoproteins on the cell surface. © 1993 Wiley‐Liss, Inc.
Mutant cell line demonstrating a block in insulin and insulin‐like growth factor type 1 (IGF‐1) induced mitogenesis
Authors: Leckett Blaine , Spurmanis Aleksandrs , Allen Cynthia , Germinario Ralph J. 
Published: February 2005

AbstractRecently, we have isolated a Chinese hamster cell variant (IV‐A1‐j) resistant to an insulin‐diphtheria‐A chain toxic conjugate (Leckett and Germinario: Cytotechnology [in press]. This cell line exhibited a decreased level of insulin binding, but normal growth in serum‐containing medium when compared to the parental cell line (V‐79). In this paper we further demonstrate that while IV‐A1‐j cells are capable of growing in serum‐containing medium, they are insensitive to the mitogenic actions of either insulin or IGF‐1. In contrast, epidermal growth factor (EGF) and/or α‐thrombin (THR) generate a mitogenic effect in IV‐A1‐j cells comparable to that observed in the parental V‐79 cells. The combination of EGF and/or THR with either insulin or IGF‐1 results in an increase in V‐79 cell growth above EGF and/or THR alone. On the other hand, insulin or IGF‐1 in the presence of other mitogens did not stimulate further growth in IV‐A1‐j cells. While insulin binding was lower in IV‐A1‐j cells, internalization of 125I‐insulin was not different in the two cell types. Additionally, insulin‐stimulated glycogen synthesis and protein synthesis were not different in the two cell types. These observations are consistent with insulin and IGF‐1 sharing a mitogenic signalling pathway in Chinese hamster fibroblasts and that this pathway is distinct from other growth factor signalling pathways. The fact that this pathway is defective in the IV‐A1‐j cell line indicates the potential usefulness of these cells in identifying a key step(s) in the insulin (IGF‐1) mitogenic pathway. © 1993 Wiley‐Liss, Inc.
Position‐dependent nuclear accumulation of the retinoblastoma (RB) protein during in vitro myogenesis
Authors: Szekely Laszlo , Jin Pei , Jiang Wei‐Qin , Rosén Anders , Wiman Klas G. , Klein George , Ringertz Nils 
Published: February 2005

AbstractThe expression of the retinoblastoma (RB) protein has been studied during in vitro muscle differentiation by immunofluorescence staining with three different antibodies against RB protein. Proliferating mononucleate L6 rat myoblasts showed a low level of expression. As cells began to enter a nonreplicating G0 state, the cell population became heterogeneous. Some nonreplicating cells showed a high level of expression. Nuclei at the two ends of myotubes were strongly positive, whereas centrally located nuclei showed low RB expression. Overexpression of the human RB protein in rat L6 myotubes from a Semliki forest virus (SFV)‐based, transient expression vector produced a similar picture. Terminally located nuclei expressed human RB at a much higher level than did the centrally located nuclei. The results suggest that individual nuclei with a multinucleated syncytium may undergo position‐dependent specialization. © 1993 Wiley‐Liss, Inc.
Marrow stroma‐derived osteogenic clonal cell lines: Putative stages in osteoblastic differentiation
Authors: Fried A. , Benayahu D. , Wientroub S. 
Published: February 2005

AbstractThis report documents characterization of five osteogenic cell subpopulations of bone marrow stroma. The clonally derived cell lines were isolated from the parental line MBA‐15 known to express osteoblastic‐associated features in vitro and to form bone in vivo. The latter, presumably “arrested” at a particular stage along the osteogenic lineage, are useful models to study the processes involved in the differentiation of bone forming cells. The clones differ in their morphology, proliferation rate, quantities and distribution of extracellular matrix proteins, levels of alkaline phosphatase activity and activation of adenylate cyclase by parathyroid hormone and/or prostaglandin E. These properties have been retained during prolonged growth and subculturing through many passages. MBA‐15.4 is a presumptive preosteoblast with a fibroblast‐like appearance; it proliferates rapidly, synthesizes equal amounts of collagen and noncollagenous proteins, and produces constitutively low levels of alkaline phosphatase. This clone has PGE2‐stimulated adenylate cyclase activity and a very low constitutive response to PTH. On the other hand, MBA‐15.6 has a large polygonal morphology with limited proliferative potential, synthesizes twice as much noncollagenous proteins as collagen, has high alkaline phosphatase activity, and responds strongly to PTH. The characteristics of the other clones place them between these two categories. The effects of 10−7 M dexamethasone or 10−12–10−8 M 1,25 dihydroxyvitamin D3 on growth and differentiation further strengthen the variance between these clones. The different in vitro characteristics of the various clones were directly reflected in their bone formation ability in vivo. When transplanted under the renal capsule, MBA‐15.33 formed a thick fibrous tissue, MBA‐15.4 formed small foci of bone, and MBA‐15.6 formed massive woven bone at the same period of time. © 1993 Wiley‐Liss, Inc.
Interactions of sertoli cells with laminin are essential to maintain integrity of the cytoskeleton and barrier functions of cells in culture in the two‐chambered assembly
Authors: Tung Pierre S. , Fritz Irving B. 
Published: February 2005

AbstractThe addition of anti‐laminin IgG to the basal surfaces of rat Sertoli cells in culture in a two‐chambered assembly results in a perturbation of F‐actin arrangements, including disruption of the pericellular circumferal rings, impairments of the Sertoli cell permeability barrier, and subsequently focal defoliation, followed by cell reaggregation. The pentapeptide YIGSR, which competes with the laminin receptor for laminin (Kleinman and Weeks: Curr. Oph. Cell Biol., 1:964–967, 1989; Graf et al.: Biochemistry, 26:6896–6900, 1987) also elicited focal defoliation of Sertoli cells from the extracellular matrix‐coated filter in the two‐chambered assembly. Addition of YIGSR to Sertoli cell cultures resulted in cell detachment within 2 to 3 h. In contrast, the irrelevant peptide YIGSE had no detectable effects. The anti‐laminin IgG was effective only when added to the chamber in which access was readily available to the basal surfaces of Sertoli cells, but YIGSR was effective when added either to the outer chamber or to the inner chamber. These data were interpreted to indicate that the Sertoli cell barrier generated in the two‐chambered assembly allowed a relatively rapid diffusion of YIGSR between chambers, but prevented the rapid equilibration of anti‐laminin IgG between compartments. Addition of anti‐laminin IgG to the basal, but not to the apical surfaces of Sertoli cells, resulted in more rapid rates of equilibration of [3H]‐methoxyinulin and [86Rb]Cl across the Sertoli cell monolayer. This evidence of impairment to the integrity of the barrier was detected prior to the disruption of stress fibers and focal defoliation, but after evidence of dissolution of the circumferal F‐actin ring, which occurred within 1 h after addition of anti‐laminin IgG. We consider the possibility that a transmembrane link exists between extracellular laminin and cytoskeletal elements which modulates the circumferal F‐actin ring. We further postulate that this linkage can influence the nature of tight junctional complexes, and thereby the integrity of the Sertoli cell barrier. © 1993 Wiley‐Liss, Inc.
Induction of GTP‐cyclohydrolase I mRNA expression by lectin activation and interferon‐γ treatment in human cells associated with the immune response
Authors: Schott Karin , Gütlich Markus , Ziegler Irmgard 
Published: February 2005

AbstractThe development of tetrahydrobiopterin synthesis during lectin stimulation of resting human T lymphocytes (Kerler et al. [1989] FEBS Lett., 250:622–624), the interferon‐γ induced neopterin production by human monocytes/macrophages (Huber et al. [1984] J. Exp. Med., 160:310–16), and the control of tetrahydrobiopterin synthesis in activated T cells by the synergistic action of interferon‐γ and interleukin 2 (Ziegler et al. [1990] J. Biol. Chem., 265:17026–17030) were previously explained by modulation of the apparent GTP‐cyclohydrolase I activation. In this study we demonstrate that increases in GTP‐cyclohydrolase I activity which occur after lectin induction and after cytokine treatment correlate with increased steady state mRNA levels specific for this enzyme. The enhancement of interferon‐γ induced enzyme activity in primed T cells by interleukin 2 also corresponds to further increases in mRNA expression. The steady state GTP‐cyclohydrolase I mRNA levels in primed T cells, however, do not correlate with the steep decline which follows the culmination of enzyme activity 44 hours after treatment. This indicates that the down‐regulation of apparent GTP‐cyclohydrolase I activity is caused by posttranslational modification of the protein. © 1993 Wiley‐Liss, Inc.
Core proteins of soluble chondroitin sulfate proteoglycans purified from the rat brain block the cell cycle of PC12D cells
Authors: Katoh‐Semba Ritsuko , Oohira Atsuhiko 
Published: February 2005

AbstractThe effects of soluble chondroitin sulfate proteoglycans (CSPGs) purified from the rat brain on proliferation of and neurite outgrowth from PC12D cells (Katoh‐Semba et al., J Neurosci Res 17:36, 1987) were investigated. When PC12D cells are cultured under standard conditions, they proliferate with a doubling time of about 2 days, irrespective of the presence or absence of NGF. However, the addition of a mixture of several types of purified soluble brain CSPG (50 nmol uronic acid/ml) to the culture medium prevented the increase in the number of PC12D cells as well as the nerve growth factor (NGF)‐induced neurite extension. The dose for 50% inhibition (ID50) was 1.6 nmol/ml for cell proliferation and 2.7 nmol/ml for neurite elongation. The increase in cell number seemed to stop around 6 h after exposure to culture medium supplemented with brain‐derived CSPGs, and even substratum‐attached CSPGs were able to exert such inhibitory effects. Only brain‐type CSPGs, not a cartilage‐derived CSPG (PGH) or a hyaluronate‐binding PGH, had such inhibitory effects. Furthermore, these inhibitory activities were associated only with the core proteins of brain‐derived CSPGs, and not with polysaccharide chains from brain‐derived CSPGs. Incorporation of [3H]thymidine into DNA did not decrease for at least the first 12 h. Consequently, the amount of DNA per cell after 48 h of culture was about twofold higher in cells treated with brain CSPGs than in nontreated cells after exposure to the medium with CSPGs. Microspectrophotometry revealed that the population of cells with a high DNA content was greater in the culture treated with brain‐derived CSPGs than in the control culture. These findings indicate that purified soluble brain CSPGs block the cell cycle of PC12D cells at the G2 phase with resultant cessation of cell proliferation and the inhibition of neurite outgrowth. © 1993 Wiley‐Liss, Inc.