bluuebunny/crossref_metadata_2025_split huggingface dataset:
Arabinosylcytosine‐induced accumulation of DNA nicks in myotube nuclei detected by in situ nick translation
Authors: Masuck Tony M. , Taylor Andrew R. , Lough John 
Published: February 2005

AbstractThis laboratory has recently reported the occurrence of DNA nicking at the onset of terminal skeletal myogenesis by using the technique of in situ nick translation (Dawon and Lough: Dev. Biol., 727:362–367, 1988). Because 1‐beta‐D‐arabi‐nofuranosylcytosine (araC), a cytocidal agent that is routinely used to remove dividingfibroblasts from myogenic cultures, inhibits DNA repair, it was of interest to determine whether araC treatment resulted in an accumulation of the endogeously created nicks. Thus, we have assessed the accumulation of DNA nicks in myotube cells during a 20 hour araC treatment period at the onset of terminal myogenesis (44–64 hours in vitro) by using three techniques: alkaline sucrose gradient density centrifugation, kinetic in situ nick translation, and cellular in situ nick translation. Although alkaline sucrose gradient centrifugation revealed no detectable nicking after 20 hours, kinetic in situ nick translation analysis revealed subtle but significant increases in DNA nicks caused by araC within 7 hours of drug application, and a 1.5‐fold increase in DNA repair sites after 20 hours of drugtreatment. That theseo bservationsreflectednicking specifically in myotube nuclei was determined by immunocytochemical localization of nicked sites after repair with a biotinyafed nucleotide analog (biotin‐11 ‐dUTP). The effects of araC wereonly incompletely reversible, whetheror not thedrugwas removedfrom the cultures, within 2 days of the treatment period. These results indicate that araC inhibits the repair of endogenously created nicks in the DNA of non‐dividing, terminally differentiating myotube cells, and provide further evidence for DNA nicking and repair as a feature of terminal myogenic differentiation.
Butyrate induces an EctoMg2+‐ATPase activity in Li‐7A human hepatoma cells
Authors: Murray Sandra L. , Knowles Aileen F. 
Published: February 2005

AbstractThe human hepatoma cell line (Li‐7A) possesses a high concentration of epider‐mal growth factor (EGF) receptors and exhibits ectoATPase activity in the presence of either MgATP or CaATP (Knowles: J. Cell. Physiol., 734:109–116, 1988). Growth for 96 hours in the presence of both EGF and cholera toxin or another cyclic AMP elevating agent induced an ectoATPase activity which was more active with CaATP and resistant to inhibition by the sulfhydryl reagent, p‐chloromercuriphenylsulfonate (pCMPS) (Knowles: Arch. Biochem. Biophys., 263: 264–271, 1988). In contrast, treatment of cells with butyrate, a short chain organic acid which can be derived from the analogue, dibutyryl cyclic AMP, resulted in a 4‐7‐fold increase of an ectoATPase which was more active with MgATP and highly sensitive to pCMPS inhibition. Maximal induction by butyrate required 48 hours and was dependent on butyrate concentration, but was independent of EGF and cyclic AMP elevating agents. Of six organic acids tested, butyrate was most effective in the induction of the ectoMg2+‐ATPase. The increase in the ectoMg2+‐ATPase activity could be prevented with actinomycin D and cycloheximide, indicating that both transcription and translation were necessary for induction. In addition to the induction of the ectoMg2+‐ATPase, bu‐tyrate induced alkaline phosphatase activity, but had no effect on a third ectoen‐zyme, 5′‐nucleotidase. These data further support our proposal that two distinct ectoATPases exist in the plasma membranes of Li‐7A hepatoma cells.
Nuclear and cytoplasmic localization of different basic fibroblast growth factor species
Authors: Renko M. , Quarto N. , Morimoto T. , Rifkin D. B. 
Published: February 2005

AbstractThe subcellular distribution of basic fibroblastic growth factor (bFGF) was analyzed by subcellular fractionation and immunofluorescence to gain insight into potential mechanisms for its release from cells. Subcellular fractionation of either SK‐Hep‐1 cells or NIH 3T3 cells transfected with a bFGF cDNA revealed that the 18 kd form of bFGF was found primarily in the cytosolic fraction, whereas the 22 and 24 kd forms of bFGF were found preferentially in ribosomal and nuclear fractions. Analysis of bFGF distribution by immunofluorescence using an antibody that recognized all forms of bFGF indicated both cytoplasmic and nuclear localization but failed to reveal any growth factor in structures representing secretory vesicles. Therefore, bFGF has a distribution inconsistent with that of a secretory protein.
Human bone cell enzyme expression and cellular heterogeneity: Correlation of alkaline phosphatase enzyme activity with cell cycle
Authors: Fedarko N. S. , Bianco P. , Vetter U. , Robey P. Gehron 
Published: February 2005

AbstractAlkaline phosphatase, long implicated in biomineralization, is a feature of the osteoblast phenotype. Yet in cultured bone cells, only a fraction stain positive histochemically. To determine whether osteoblast enzyme expression reflects cellular heterogeneity with respect to cell cycle distribution or length of time in culture, the activities of alkaline phosphatase, tartrate‐resistant and ‐sensitive acid phosphatases, and non‐specific esterases were assayed kinetically and histo‐chemically. In asynchronous subconfluent cultures, < 15% of the cells stained positive and assayed activity was 0.04 IU/106 cells/cm2. After 1 week, the percent of alkaline phosphatase positive‐staining cells increased 5‐fold, while activity increased 10‐fold. Non‐specific esterases and tartrate‐sensitive acid phosphatase were constitutive throughout time in culture, whereas tartrate‐resistant acid phos‐phatase activity appeared after 2 weeks. Cell cycle analysis of human bone cells revealed a growth fraction of 80%, an S phase of 8.5 h, G2 + 1/2 M of 4 h, and a G1 of 25–30 h. In synchronous cultures induced by a thymidine‐aphidicolin protocol, alkaline phosphatase activity dropped precipitously at M phase and returned during G1. A majority of the alkaline phosphatase activity lost from the cell surface at mitosis was recovered in the medium. Tartrate‐sensitive acid phos‐phatase and non‐specific esterase levels were relatively stable throughout the cell cycle, while tartrate‐resistant acid phosphatase activity was not assavable at the density used in synchronous cultures. From these data, variations in alkaline phosphatase activity appear to reflect the distribution of cells throughout the cell cycle.
Colony stimulating factor‐1 Is a negative regulator of the macrophage respiratory burst
Authors: Phillips Wayne A. , Hamilton John A. 
Published: February 2005

AbstractSeveral cytokines have previously been shown to prime macrophages for enhanced release of oxygen radicals in response to subsequent stimulation. We now demonstrate that the presence of the macrophage‐specific colony stimulating factor‐1 (CSF‐1) inhibits the priming of murine macrophages by a variety of agents including tumor necrosis factor α, granulocyte/macrophage colony stimulating factor, interferon‐γ, and bacterial lipopolysaccharide. CSF‐1 is also able to reduce the respiratory burst in the absence of priming. Our results indicate that CSF‐1 is a potent negative regulator of the macrophage respiratory burst which acts to oppose the priming (enhancing) action of macrophage activating agents. We propose that CSF‐1 may have a potentially important and previously unrecognized, role as a Physiological regulator which restricts or terminates the activation of macrophages in order to prevent an uncontrolled inflammatory reaction.
Plasminogen activators and their inhibitors in the neuromuscular system: I. Developmental regulation of plasminogen activator isoforms during in vitro myogenesis in two cell lines
Authors: Festoff Barry W. , Rao Jasti S. , Maben Cecilia , Hantaï Daniel 
Published: February 2005

AbstractPlasminogen activators (PAs), were estimated qualitatively and quantitatively in two different clonal murine skeletal muscle cell lines. Both cell lines produced the two major types of PAs found in mammalian cells, urokinase‐type (uPA) and tissue type (tPA). These two lines are models for the study of myogenesis in vitro, but differ in several growth and differentiation characteristics. Because of their possible involvement in these characteristics we assayed the expression of PAs in both cell systems during development in culture. Utilizing fibrin zymography two isoforms of tPA were detected. One co‐migrated with human tPA at 75 kd and another may represent a tPA:inhibitor complex at 105 Kd. Several isoenzymes of uPA were detected and these changed depending on whether cell homogenates or conditioned medium was analyzed and whether myogenic cells were at single‐cell myoblast or multi‐nucleated myotube stage. Species‐specific antisera to mouse uPA identified 4 uPA bands in muscle cell medium and 5 in cell layers. Antigenic uPA bands also varied depending on stage of myogenesis. Quantitative amidolytic studies using chromogenic substrates showed that maximal PA activity, both uPA and tPA, occurred at the time of myoblast fusion. Furthermore, uPA activity in membranes increased during myogenesis, while both uPA and tPA in medium decreased after fusion. These studies indicate that muscle PA expression is developmentally regulated and may correlate with growth and differentiation in skeletal muscle.
Differences in inositol phosphate production in rat tail artery and thoracic aorta
Authors: Labelle Edward F. , Murray Bonnie M. 
Published: February 2005

AbstractPharmacomechanical coupling of vascular smooth muscle is believed to be mediated by inositol trisphosphate (IP3). Numerous studies have demonstrated an increase in inositol phosphates following tissue stimulation using either intact aortic strips or cultured cells from aorta. However, little information is available concerning inositol phosphates in vascular tissue other than in the large conduit vessel, the aorta. This present study was designed to examine the role of inositol phosphate metabolism following adrenergic stimulation of the muscular rat tail artery as compared to the aorta. Segments of thoracic aorta and tail artery from male Sprague Dawley rats were labe ed with [3H]inositol and stimulated with norepinephrine. The norepinephrine concentration that resulted in a half‐maximal stimulation of inositol phosphates was ⌣︁−6 M in both the aorta and tail artery. Although the sensitivity of the two vessels to norepinephrine stimulation were similar, the stimulated levels of IP, IP2, and IP3 were from 1 to 2 orders of magnitude greater in the tail artery than in aorta. IP production in aorta and tail artery was a linear function of time (from 0 to 30 min). Significant levels of IP3 (the 1, 4, 5‐IP3 isomer as determined by HPLC) could only be detected in the tail artery and appeared to be produced optima ly after 5 min of stimulation. The several order of magnitude increase in adrenergic stimulated inositol phosphate production in the tail artery was not due to either an increased magnitude of [3H]inositol incorporated into PI, PIP, and PIP2 or to a greater percentage of smooth muscle cells per unit tissue of the rat tail artery. We believe the resul s of this study demonstrate that the increased inositol phosphate metabolism n the vascular smooth muscle cells of the tail artery is an intrinsic property of the cell. Moreover, due to the significant levels of all inositol phosphates produced in the tail artery, this muscular artery may be a better model, as compared to the aorta, for future studies investigating pharmacomechanical coupling of vascular smooth muscle.
Caffeine, an inhibitor of endocytosis in dictyostelium discoideum amoebae
Authors: Gonzalez Carlos , Klein GéRard , Satre Michel 
Published: February 2005

AbstractThe effect of the trimethylxanthine, caffeine, was examined on the growth and endocytosis pathways of the vegetative amoebae of the cellular slime mold Dic‐tyostelium discoideum. Caffeine at concentrations of 1.5–3 mM was found to inhibit axenic growth, fluid‐phase pinocytosis, and secretion of lysosomal enzymes. Cell viability was unaffected by incubation for 16 hours with 5 mM caffeine but decreased markedly thereafter. Phagocytosis of the bacterium Esch‐erichia coli by Dictyostelium amoebae was also inhibited by caffeine, although at concentrations twofold to threefold higher. Caffeine rapidly entered into amoebae to reach an equilibrium between extracellular and intracellular concentrations, and it was not appreciably metabolized by Dictyostelium. Inhibition of growth and endocytosis was reversible upon removal of the drug and was partially counteracted by 10 mM adenosine. As caffeine discharged intracellular calcium stores in Dictyostelium (Abe et al., 1988), its inhibitory effect on endocytosis could result from the perturbation of calcium homeostasis. In agreement with this hypothesis, the cation La3+ (10 μM), a Ca2+‐transport inhibitor, also strongly reduced fluid‐phase pinocy'osis.
Hybrids from fusion of normal human T lymphocytes with immortal human cells exhibit limited life span
Authors: Pereira‐Smith Olivia M. , Robetorye Susan , Ning Yi , Orson Frank M. 
Published: February 2005

AbstractA number of normal human cell types have been shown to exhibit cellular senescence in vitro. We and others had found that fusion of normal human fibroblasts with immortal human cells yielded hybrids having limited lifespan. This indicated that the phenotype of cellular senescence is dominant and that immortality results from recessive changes in genes involved in growth control. They also supported the hypothesis that senescence results from genetic mechanisms rather than random damage. Since T lymphocytes are a highly differentiated cell type, in contrast to fibroblasts, it was of interest to determine whether similar mechanisms caused senescence in the T cells. We therefore fused normal human T lymphocytes with an immortal human cell line to determine whether they could restore the senescent, nondividing phenotype in hybrids, as do normal human fibroblasts. Eleven of fifteen hybrid clones studied exhibited limited proliferative potential after achieving a range of population doubling similar to that observed in the cell fusion studies involving normal fibroblasts. These results provide evidence that cellular senescence in T lymphocytes occurs via genetic mechanisms.
Role of the Na+ /K+ /Cl− transporter in the positive inotropic effect of ouabain in cardiac myocytes
Authors: Panet Rivka , Fixler Ruhama , Snyder David , Raz Shmuel , Atlan Henri , Eilam Yael , Hasin Yonathan 
Published: February 2005

AbstractIn this study we have characterized the bumetanide‐sensitive K+ /Na + /Cl− cotransport in cultured rat cardiac myocytes. (1) It carries about 10% of the total K+ influx. (2) It is sensitive to furosemide (Ki0.5 = 10−6M) and bumetanide (Ki0.5 = 10−7M) (3) It is strongly dependent on the extracellular concentrations of Na+ and Cl− (4) It carries out influx of both ions, K+ and Na+. A therapeutic concentration of ouabain (10−7M) stimulated the bumetanide‐sensitive K+ influx (as measured by 86Rb +), in the cultured myocytes, with no effect on the bumetanide‐resistant K+ influx, which was mediated mostly by the Na + /K + pump. Stimulation of the bumetanide‐sensitive Rb+ influx by a low ouabain concentration was strongly dependent on Na+ and Cl− in the extracellular medium. A low concentration of ouabain (10−7M) was found to increase the steady‐state level of cytosolic Na + by 15%. This increase was abolished by the addition of bumetanide or furosemide. These findings suggest that ouabain, at a low (10−7M) concentration, induced its positive inotropic effect in rat cardiac myocytes by increasing Na+ influx into the cells through the bumetanide‐sensitive Na+ / K+ /Cl− cotransporter. In order to examine this hypothesis, we measured the effect of bumetanide on the increased amplitude of systolic cell motion induced by ouabain. Bumetanide or furosemide, added to cultured cardiac myocytes, inhibited the increased amplitude of systolic cell motion induced by ouabain. Neither bumetanide nor furosemide alone has any significant effect on the basal amp itude of systolic cell motion. We propose that stimulation of bumetanidesensitive Na+ influx plays an essential role in the positive inotropic effect in rat cardiac myocytes induced by low concentrations of ouabain.
Bumetanide‐sensitive Na+ /K+ /Cl− transporter is stimulated by phorbol ester and different mitogens in quiescent human skin fibroblasts
Authors: Panet Rivka , Atlan Henri 
Published: February 2005

AbstractIn this study we investigated the correlation between the mitogenic effect and stimulation of Rb + (K + ) fluxes in human skin fibroblasts treated by purified growth factors. Both K+ transporters, bumetanide‐sensitive and ouabain‐sensitive, are stimulated 2‐3‐fold after addition of either fetal calf serum or purified recombinant growth factors to quiescent G0/G1 human skin fibroblasts. Three groups of mitogens were compared: (i) the phorbol ester 2‐O‐tetradecanoyl‐phorbol‐13‐acetate (TPA); (ii) growth factors that stimulate inositol phosphate hydrolysis and subsequently activate protein kinase C—fibroblast growth factor (FGF), platelet derived growth factor (PDGF), and α‐thrombin; and (iii) growth factors that do not activate kinase C—insulin‐like growth factor‐1 (IGF‐1), and transforming like growth‐factor‐α (TGF‐α). The three groups of mitogens stimulated human skin fibroblasts proliferation and Rb+ influxes in a similar dose‐dependent fashion. The results indicate that both the bumetanide‐sensitive and the ouabain‐sensitive Rb+ fluxes are stimulated by protein kinase C‐dependent and by the protein kinase C‐independent pathways of the mitogenic signal.
Stimulation of fibronectin production by TGF‐β1 is independent of effects on cell proliferation: The example of bovine adrenocortical cells
Authors: Shi De Li , Savona Catherine , Chambaz Edmond M. , Feige Jean‐Jacques 
Published: February 2005

AbstractWe reported previously that transforming growth factor β (TGF‐β) does not influence the proliferation of bovine adrenocortical cells but is a very potent inhibitor of their steroidogenic functions (Feige et al.:Journal of Biological Chemistry 262:13491‐13495, 1987). In the present study, we addressed the question of whether these cells modify the synthesis of their extracellular matrix (in particular of fibronectin) in response to TGF‐β1, similarly to the changes observed in cell types whose growth is modified by this factor (e.g., fibroblasts). Immunofluorescence studies using anti‐fibronectin antibodies revealed that TGF‐β1, treatment in serum‐free medium induced the formation of fibronectin‐containing fibrils associated with adrenocortical cells. Metabolic labeling of adrenocortical cells with [35S]‐methionine showed that fibronectin synthesis and secretion were highly stimulated by low concentrations of TGF‐β1. Half‐maximal stimulation was observed for TGF‐β1 concentrations in the range of 0.1 to 0.5 ng/ml and maximal stimulation reached 35‐fold over control at the concentration of 2 ng/ml. The earlier detectable effect was observed after 8 h of treatment (6‐fold stimulation) and the maximal increase was reached after 24 h of treatment. Stimulation of adrenocortical fibronectin synthesis by TGF‐β1 appeared to imply a transcriptional event since it was no longer observed in the presence of DRB, a potent inhibitor of RNA polymerases, and because the level of fibronectin mRNA was stimulated under TGF‐β1 treatment. Taken together, these results indicate that the increased expression of fibronectin is not closely related to growth‐regulatory effects of TGF‐β1 since it is also observed in adrenocortical cells, whose proliferation is unaffected by TGF‐β1.
Osteoblasts display receptors for and responses to leukemia‐inhibitory factor
Authors: Allan E. H. , Hilton D. J. , Brown M. A. , Evely R. S. , Yumita S. , Metcalf D. , Gough N. M. , Ng K. W. , Nicola N. A. , Martin T. J. 
Published: February 2005

AbstractSpecific binding of leukemia‐inhibitory factor (LIF) to osteoblasts, but not multinucleated osteoclasts, was demonstrated by receptor autoradiography by *using cells isolated from newborn rat long bones. The clonal rat osteogenic sarcoma cells, UMR 106‐06, which have several phenotypic properties of osteoblasts, expressed 300 LIF receptors per cell, with an apparent KD of 60 pM. Treatment of calvarial osteoblasts or UMR 106‐01 cells with LIF resulted in a dose‐dependent inhibition of plasminogen activator (PA) activity. Both calvarial osteoblasts and osteogenic sarcoma cells were shown by Western blotting and reverse fibrin autography to produce plasminogen activator inhibitor‐1 (PAI‐1), the production of which was increased by LIF treatment. Northern blot analysis revealed that LIF treatment resulted in a rapid (peak 1 hour), dose‐dependent increase in mRNA for PAI‐1. LIF treatment of the preosteoblast cell line, UMR 201, enhanced the alkaline phosphatase response of these cells to retinoic acid. Each of the osteoblast‐like cell types (calvarial osteoblasts, UMR 106‐06, and UMR 201) was shown to produce LIF by bioassay and, by using the polymerase chain reaction (PCR), was shown to express low levels of mRNA for LIF. These data establish that cells of the osteoblast lineage are targets for LIF action. The reported anabolic effects of this cytokine on bone formation in vivo could be related to inhibition of protease activity. LIF may be an important paracrine modulator in bone, or perhaps an autocrine one, based on the evidence for its production by osteoblasts and osteoblast‐like cells.
Regulation of tubulin synthesis during the cell cycle in the synchronous plasmodia of Physarum polycephalum
Authors: Ducommun B. , Cance J. , Wright M. 
Published: February 2005

AbstractRegulation of α‐ and β‐tubulin isotype synthesis during the cell cycle has been studied in the myxomycete Physarum polycephalum, by subjecting synchronous plasmodia to temperature shifts and pharmacological perturbations. Temperature *shifts interfered with the regulation of tubulin synthesis. Inhibition of DNA synthesis prevents tubulin degradation after completion of the cell cycle (Ducommun and Wright, Eur. J. Cell Biol., 50:48–55, 1989) but did not perturb the initiation of tubulin synthesis. The constant increase of tubulin synthesis in the presence of tubulin‐sequestering drugs and the decrease of tubulin synthesis during a treatment with aphidicolin in late G2 phase suggest the existence of an autoregulatory mechanism of tubulin synthesis. Moreover, the microtubule poison methyl benzimidazole carbamate dissociated synthesis of the α1‐tubulin isotype from the generally strictly coordinated synthesis of all tubulin isotypes during the transient interruption of mitosis. These observations show that a microtubular poison can perturb regulation of the synthesis of specific isotubulins.