bluuebunny/crossref_metadata_2025_split huggingface dataset:
Activation of signal transduction pathways protects quiescent Balb/c‐3T3 fibroblasts against death due to serum deprivation
Authors: Tamm Igor , Kikuchi Toyoko 
Published: February 2005

AbstractPlatelet‐derived growth factor (PDGF), epidermal growth factor (EGF), insulin‐like growth factor‐1 (IGF‐1), and insulin protect density‐inhibited murine Balb/c‐3T3 fibroblasts against death by distinctive mechanisms. Determination of the cell survival‐enhancing activity of growth factors by cell enumeration and neutral red uptake measurement gives equivalent results. PDGF displays a steep doseresponse relationship in the 1−5 ng/ml range. The other factors display shallow log‐linear relationships in the following ranges: EGF: 0.2−5 ng/ml; IGF‐1: 2−80 ng/ml; and insulin: 57−4,500 ng/ml. Agonists that lead to the activation of protein kinase A, including forskolin, 8‐bromoadenosine 3′:5′‐cyclic monophosphate (Br‐cAMP) and N6,2′‐O‐dibutyryladenosine 3′:5′‐cyclic monophosphate (db‐cAMP), markedly increase both short‐term (5‐h) and long‐term (20‐h) survival of cells. 2‐lsobutyl‐1‐methylxanthine (IBMX) markedly enhances short‐term survival, but its effect decays with time. The protein kinase C agonist 12‐O‐tetradecanoyl phorbol‐13‐acetate (TPA) has a moderate protective effect at concentrations of 16‐32 nM, and 64 nM TPA is highly effective. The synthetic diacylglycerols 1,2‐dioctanoylglycerol (DiC8) and 1‐oleoyl‐2‐acetylglycerol (OAG) and the calcium ionophore ionomycin show low activity. Supplemation of EGF with a protein kinase A or C agonist results in a varying additive increase in short‐term (5‐h) cell survival and supplementation of EGF+insulin or PDGF+EGF+insulin increases further the already high level of protection given by the growth factor combinations. Combining a protein kinase A and a protein kinase C agonist in the absence of growth factors gives an approximately additive increase in cell survival. Results obtained with kinase, RNA, and protein synthesis inhibitors suggest that: (1) activated protein kinase C catalyzes one or more phosphorylation events in quiescent Balb/c‐3T3 cells that lead to gene expression with the protein product(s) mediating protection of quiescent cells against death, and (2) phosphorylation events Catalyzed by protein kinase A largely serve to protect cells by a mechanism not requiring de novo RNA and protein biosynthesis.
Xyloside effects on in vitro hematopoiesis: Functional and biochemical studies
Authors: Kirby Suzanne L. , Bentley Stuart A. 
Published: February 2005

AbstractXyloside supplementation of long‐term bone marrow cultures (LTBMCs) has been reported to result in greatly enhanced proliferation of hematopoietic stem cells. This was presumed to be the result of xyloside‐mediated perturbation of proteoglycan synthesis by marrow‐derived stromal cells. To investigate this phenomenon, we first studied the effects of xyloside supplementation on proteoglycan synthesis by D2XRadll bone marrow stromal cells, which support hematopoietic stem cell proliferation in vitro. D2XRadll cells were precursor labelled with 35S‐sulfate, and proteoglycans separated by ion exchange chromatography, isopyknic CsCI gradient centrifugation, and gel filtration HPLC. Xylo‐side‐supplemented cultures showed an approximately fourfold increase in total 35S incorporation, mainly as free chondroitin‐dermatan sulfate (CS/DS) gly‐cosaminoglycan chains in the culture media. Both xyloside supplemented and nonsupplemented cultures synthesized DS1, DS2, and DS3 CS/DS proteoglycans as previously described. In contrast to previous reports, xyloside was found to inhibit hematopoietic cell growth in LTBMC. Inhibitory effects were observed both in cocultures of lL‐3‐dependent hematopoietic cell lines with supportive stromal cell lines and in primary murine LTBMCs. Xyloside was found to have a marked inhibitory effect on the growth of murine hematopoietic stem cells and IL‐3‐dependent hematopoietic cell lines in clonal assay systems and in suspension cultures. In contrast, dialyzed concentrated conditioned media from LTBMCs had no such inhibitory effects. These findings suggest that xyloside‐mediated inhibition of hematopoietic cell growth in LTBMC resulted from a direct effect of xyloside on proteoglycan synthesis by hematopoietic cells.
Generation of superoxide anion by brain endothelial cell xanthine oxidase
Authors: Terada Lance S. , Willingham Irene R. , Rosandich Mary E. , Leff Jonathan A. , Kindt Glenn W. , Repine John E. 
Published: February 2005

AbstractBovine brain endothelial cells (EC) that were isolated and propagated in pure culture had increased ( 20‐fold) levels of xanthine oxidase and xanthine dehydrogenase activity compared to whole brain homogenate. Brain EC also released superoxide anion (O) in to the extracellular medium. Treatment of EC with tungsten decreased (P < 0.05), both XO activity and O release. XO appears to be highly concentrated in cerebral vascular endothelium and may be an important source o; O.
Induction of plasminogen activator inhibitor 1 biosynthesis by hyperthermia
Authors: Bergonzelli Gabriela E. , Kruithof Egbert K. O. 
Published: February 2005

AbstractHyperthermia is a clinical sign of inflammation and constitutes in itself an adaptive defense mechanism. The fibrinolytic system, a highly regulated proteolytic system, a involved in inflammatory processes. Plasminogen activator inhibitor 1 (PAI‐1) is the principal inhibitor of the two activators of the fibrinolytic system: tissue‐ and urokinase‐type PAs (t‐PA and u‐PA). Our present paper provides the first evidence that hyperthermia can directly induce PAI‐1. A moderate heat stress, sufficient to induce heat shock protein 70 mRNA approximately 100‐fold, resulted in a two‐to three‐fold increase in functionally active PAI‐1 in the conditioned medium of human HT‐1080 fibrosarcoma and Hep G2 hepatoma cells. Exposure of these cells to 42°C led to a similar two‐fold and two‐to five‐fold induction of PAI‐1 mRNA expression in HT‐1080 and Hep G2 cells, respectively, as has been determined by using both oligo d(T) selected and total RNA preparations. These results suggest that the observed increase in PAI‐1 accumulation is due to an induction of PAI‐1 biosynthesis. Run‐on transcription analysis indicates that the induction of PAI‐1 biosynthesis by hyperthermia is mediated by a stimulation of PAI‐1 gene transcription. No significant effect of hyperthermia was found on t‐PA or u‐PA at the level of antigen accumulation, mRNA, and gene transcription in human HT‐1080 fibrosarcoma cells. These results point to an additional regulatory mechanism of fibrinolysis in the context of inflammation.
Growth regulated expression of B‐MYB in fibroblasts and hematopoietic cells
Authors: Reiss Krzysztof , Travali Salvatore , Calabretta Bruno , Baserga Renato 
Published: February 2005

AbstractThe B‐myb cDNA has extensive sequence similarities to the c‐myb protooncogene, but, at variance with c‐myb, it is expressed in cells other than hematopoietic cells. In this paper, we show that (1) B‐myb is expressed in mouse, human, and hamster fibroblasts; (2) B‐myb mRNA levels are growth‐regulated in both fibroblasts and peripheral blood mononuclear cells; (3) by its mode of growth regulation (peak of expression, behavior in G1‐specific temperature sensitive (ts) mutants and in the presence of cycloheximide), B‐myb can be classified, like c‐myb, thymidine kinase, PCNA, and others, as a late growth‐regulated gene; (4) B‐myb mRNA levels decrease when HL‐60 cells are induced to differentiate; and (5) the increase in mRNA levels in serum‐stimulated cells is only partially explained by an increase in rate of transcription. The possibility that the B‐myb gene may be the equivalent in fibroblasts and epithelial cells of the c‐myb proto‐oncogene of hematopoietic cells is discussed.
Expansion of myelopoietic precursors and inhibition of B‐cell precursors in mice that express a T‐cell receptor gamma (Vγ 1.1JγM4Cγ4) transgene
Authors: Ferrick David A , Cumano Ana , Furlonger Caren , Min Xia , Iscove Norman N. , Paige Christopher J. , Mak Tak W. 
Published: February 2005

AbstractKnowledge of the genetic determinants that can affect renewal of multipotential stem cells and their commitment to specific cell lineages is essential to our understanding of multicellular development. However, despite the vast amount of accumulated knowledge in this area, genetic determinants that affect renewal and commitment of precursor cells are unknown. In this study, we demonstrate that three independently derived founder mouse strains, transgenic for the TcR Vγ1.1Jγ4Cγ4 (TcRγ4) chain gene, differed significantly from normal mice in their development of T and B cells as well as myelopoietic precursor cells. Ontogenic programs consistent with an acceleration of T‐cell development and a delayed appearance and suppressed levels of pre‐B‐ and B‐cell precusors were evident in these transgenic mice. In addition, TcRγ4 transgenic mice possessed a significantly elevated level of myelopoietic pluripotent precursors. 3H‐thymidine cell suicide studies suggest that higher percentages of pluipotential precursors from the bone marrow of the TcRγ4 transgenic mice were in the S phase of the cell cycle. These modulations of the lymphoid and myelopoietic compartments, however, were not found in other T‐cell receptor transgenic mice (e.g., TcR Vγ1.2Jγ2Cγ2, TcRγ2; or Vβ8.1Dβjβ2.4Cβ2, TcRβ) constructed with the same or similar cDNA expression vector. The results suggest that the expression of a specific T‐cell receptor γ chain gene, and/or an elevated level of particular subset of TcRγδL cells, may affect the proliferation and relative proportions of haemopoietic and lymphoid precursors.
DNA‐mediated transfer of a human gene that confers resistance to mitomycin C
Authors: Buchwald Manuel , Clarke Carol 
Published: February 2005

AbstractAttempts to complement the defect in the mitomycin C (MMC)‐sensitive Chinese hamster ovary (CHO) mutant MMC3 led to the isolation of hybrids with high resistance to the cytotoxic action of the drug. Hybrid cells selected with MMC after fusion of MMC3 cells to human diploid fibroblasts were approximately five times more resistant to MMC than wild‐type CHO cells but retained the original MMC3 sensitivity to another DNA cross‐linking agent, diepoxybutane. To confirm that the MMC resistance was genetically determined and was of human origin, DNA from the resistant hybrids was introduced into MMC3 cells, and transfectants were selected in MMC. These cells had the same level of MMC resistance as the hybrids. Thus we have identified a human gene that can confer MMC resistance to CHO cells. Identification of the gene should help understand the mechanisms of MMC resistance in mammalian cells.
GM‐CSF in association with IL‐1 triggers day‐8 CFU‐S into cell cycle: Role of histamine
Authors: Piquet‐Pellorce Claire , Schneider Elke , Dy Michel 
Published: February 2005

AbstractOur recent evidence for the requirement of endogeneous histamine in IL‐3‐induced proliferation of day‐8 CFU‐S has prompted us to investigate whether or not GM‐CSF, which shares with IL‐3 the ability to stimulate bone marrow histamine synthesis, could also affect the cell cycle status of CFU‐S via this mediator. We show herein that recombinant GM‐CSF alone fails to trigger day‐8 CFU‐S into S phase, but supports their survival. However, in the same experimental conditions, GM‐CSF in combination with IL‐1 induces a CFU‐S proliferation similar to that obtained in response to IL‐3, while IL‐1 by itself has no effect on this biological activity. We further provide evidence that this phenomenon is completely abolished: (i) by preventing GM‐CSF‐induced histamine synthesis by α‐FMH, the specific inhibitor of histidine decarboxylase (HDC), or (ii) by blocking the binding sites of H2 histamine receptors with their specific antagonist oxmetidine. Similar results are obtained when progenitor‐enriched bone marrow cells are used instead of the unfractionated population. In addition, we provide an argument in support of a histamine receptor modulation by GM‐CSF that could explain the lack of effect of factor‐induced histamine on day‐8 CFU‐S cell cycling. Indeed, the entry of these progenitors into S phase that is normally promoted by dimaprit, a specific histamine H2 receptor agonist, is abolished by a preincubation with GM‐CSF. Taken together, our data support the conclusion that IL‐1 makes CFU‐S sensitive to GM‐CSF‐induced endogeneous histamine that will trigger them into cell cycle, while GM‐CSF alone has no such effect on this biological activity.
Isolation of variants of chinese hamster ovary cells with abnormally low levels of GSH: Decreased ability to cleave endocytosed disulfide bonds
Authors: Mandel Richard , Ryser Hugues J.‐P. , Niaki Bijan , Ghani Farooq , Shen Wei‐Chiang 
Published: February 2005

AbstractMutants of Chinese hamster ovary cells were selected for resistance to a 3 hour exposure to 4 μ/ml N‐methyl‐N'‐nitro‐N‐nitrosoguanidine and tested for glutathione (GSH) levels. Six of eight clones that survived the initial treatment had reduced GSH levels ranging from 26 to 61% of wild‐type values. These eight cell lines were tested for their susceptibility to a drug conjugate in which methotrexate (MTX) is disulfide‐linked to poly(D‐lysine) (MTX‐SS‐PDL) to test their capacity to cleave the endocytosed disulfide bond and release free MTX from this otherwise undegradable drug conjugate. We had shown that wild‐type cells were killed by ∼1 × 10−7 M MTX given as free drug, as MTX‐poly(L‐lysine) or as MTX‐SS‐PDL, but were not affected by MTX‐poly(D‐lysine). All six lines with abnormally low levels of GSH were resistant to MTX‐SS‐PDL. The variants with the lowest levels of GSH (MNR‐5) and MNR‐10 were tested further and showed near‐normal sensitivity to MTX and MTX poly(L‐lysine). As expected, both lines were hypersensitive to melphalan. They were, however, normally sensitive to diphtheria toxin and ricin, indicating that some cleavage opf the interchain disulfides in these protein toxins occurs even when cellular GSH is abnormally low. The lesser GSH requirement for toxin activation may be due to their extraordinary potency.
Interleukin‐6 and transforming growth factor‐β synergistically stimulate chondrosarcoma cell proliferation
Authors: Guerne Pierre‐André , Lotz Martin 
Published: February 2005

AbstractThis study examines the regulation of Swarm rat chondrosarcoma (SRC) cell proliferation in vitro. In serum‐free cultures, SRC cells showed only transient DNA synthesis and this was increased by serum. Transforming growth factor‐β (TGF‐β) was identified as an essential serum component, since the mitogenic effect of sera was related to their TGF‐β content and neutralized by antibody to TGF‐β.Among a large panel of agents tested, TGF‐β was the only factor that stimulated proliferation in serum‐free media. The TGF‐β isoforms TGF‐β1 and TGF‐β2 induced similar dose‐dependent increases with maximal 62.5‐fold stimulation at 10 ng/ml.Interleukin‐6 (IL‐6) was identified as a new factor that stimulated SRC proliferation. IL‐6 effects were serum‐dependent and their magnitude correlated with the TGF‐β content in different serum preparations. In serum‐free cultures where IL‐6 by itself had no detectable effect it caused up to 7.6‐fold increased proliferation in the presence of small doses of TGF‐β (0.01–0.1 ng/ml). This synergy was unique, since no other factor tested synergized with IL‐6 or TGF‐β. In examining potential mechanisms for this synergy it was found that TGF‐β increased IL‐6 receptor expression.In summary, these results identify IL‐6 as a new and TGF‐β as the most potent growth factor for chondrosarcoma cells and describe novel interactions between these factors in the regulation of cell growth.
Maintenance and mobility of hemoglobin and water within the human erythrocyte after detergent disruption of the plasma membrane
Authors: Cameron Ivan L. , Cox Laura A. , Liu Xi Rong , Fullerton Gary D. 
Published: February 2005

AbstractIs an intact plasma membrane responsible for keeping hemoglobin and water within the human erythrocyte? If not, what is responsible? How free is Hb to move about within the erythrocyte? To answer these questions erythrocytes were taken for phase contrast microscopy, transmission electron microscopy (TEM), determination of water‐holding capacity, and proton NMR studies both before and after membrane disruption with a nonionic detergent (Brij 58). Addition of 0.2% Brij to a D2O saline solution of hemoglobin (Hb) caused particles of Hb to appear and to aggregate. This aggregation of Hb caused the amplitude of the Hb proton NMR spectra to decrease. Thus, the less mobile the Hb the lower the Hb proton spectra amplitude. Erythrocytes washed in D2O saline showed proton NMR spectra of relatively low amplitude. Addition of Brij (0.2%) to these erythrocytes caused increased Hb mobility within these erythrocytes. The TEM of fixed and thin‐sectioned erythrocytes treated with Brij showed disruption of the plasma membrane of all erythrocytes regardless of whether or not they had lost Hb. Brij‐permeabilized erythrocytes washed in D2O saline or in a D2O K buffer maintained a higher heavy water‐holding capacity upon centrifugation as compared to nonpermeabilized erythrocytes. The TEM of Brij‐treated and washed erythrocyte “shells” revealed a continuous submembrane lamina but no other evidence of cytoskeletal elements. The water‐holding capacity of the erythrocyte can be accounted for by the water‐holding capacity of hemoglobin. The evidence favors a relatively immobile state of Hb and of water in the erythrocyte that is not immediately dependent on an intact plasma membrane but is attributed to interactions between Hb molecules and the submembrane lamina.
The increase of calmodulin in PC12 cells induced by NGF is caused by differential expression of multiple mRNAs for calmodulin
Authors: Bai Guang , Weiss Benjamin 
Published: February 2005

AbstractA rat pheochromocytoma cell line (PC12 cells) was used as a model to investigate the role of calmodulin and its multiple mRNAs in NGF‐induced neuronal differentiation. The effect of NGF on the degree of differentiation was assayed using a simple differentiation scoring system. Significant increases in the differentiation score were seen by one day, and the scores increased about 10‐fold by 8 days of treatment. NGF also increased calmodulin in the PC12 cells; significant increases were seen by 2 days of treatment, and a maximum increase of 3‐fold was seen by 4 days. Northern blot analysis using a calmodulin riboprobe revealed that all five calmodulin mRNAs found in rat tissue were present in PC12 cells. The relative abundance of the calmodulin mRNAs was 1.7 ≥ 1.4 ≥ 2.3 ≥ 4.1 ≥ 0.9 kb. NGF treatment caused a differential increase in these mRNAs. The 1.4 kb transcript (from Gene II) was increased earlier (at 1 day) and to a greater extent (3‐fold) than any of the other mRNAs. Studies of the half‐lives (t1/2) of these mRNAs suggested that the t1/2 varied with the mRNA; the smaller the mRNA, the shorter the t1/2. However, there were no significant effects of NGF on the t1/2 of any of the mRNAs. These studies indicate that NGF elevates calmodulin in PC12 cells by causing a differential increase in the multiple mRNAs for calmodulin and that the increase in calmodulin may play some part in NGF‐induced neuronal differentiation in PC12 cells.
Sheep amniotic fluid has a protein factor which stimulates human fibroblast populated collagen lattice contraction
Authors: Rittenberg T. , Longaker M. T. , Adzick N. S. , Ehrlich H. P. 
Published: February 2005

AbstractSutured incisional wounds made in fetal sheep and rabbits heal without scarring. Fetal sheep excisional wounds can close by contraction, but those in fetal rabbits do not. In vivo and in vitro evidence suggests that rabbit amniotic fluid inhibits wound contraction. The question arises: does sheep amniotic fluid promote wound contraction because their fetal wounds close by contraction? Sheep amniotic fluid (SAF) from 100 and 125 days gestation was tested in fibroblast populated collagen lattice (FPCL) system, an in vitro model of wound contraction. SAF stimulated FPCL contraction in a dose responsive manner. SAF from a 100 day fetus was more stimulating than a 125 day SAF. SAF enhanced FPCL contraction in the presence or absence of serum. SAF was fractionated by size, using column chromatography. It yielded a fraction with an estimated molecular weigh near 40,000 daltons, which stimulated FPCL contraction. The factor was inactivated by proteolytic digestion and heat denaturation. This protein fraction which stimulates FPCL contraction is not related to (1) actin‐myosin filaments enhanced contraction by ATP‐induced cell contraction, (2) promotion of fibroblast elongation on glass surface or in collagen, or (3) increased cell number by enhanced fibroblast duplication in a collagen matrix. A mechanism for SAF promotion of FPCL contraction was investigated but not identified.
Glutamine synthetase expression in rat oligodendrocytes in culture: Regulation by hormones and growth factors
Authors: Fressinaud Catherine , Weinrauder Halina , Delaunoy Jean‐Pierre , Tholey Georges , Labourdette Géarard , Sarliève Louis Léon 
Published: February 2005

AbstractGlutamine synthetase (GS, EC 6.3.1.2.) has long been considered as a protein specific for astrocytes in the brain, but recently GS immunoreactivity has been reported in oligodendrocytes both in mixed primary glial cell cultures and in vivo. We have investigated its expression and regulation in “pure” cligodendrocyte cultures. “Pure” oligodendrocyte secondary cultures were derived from newborn rat brain primary cultures enriched in oligodendrocytes as described by Besnard et al. (1987) and were grown in chemically defined medium. These cultures contain more than 90% galactocerebroside‐positive oligodendrocytes and produce “myelin” membranes (Fressinaud et al., 1990) after 6–10 days in subcultures (30–35 days, total time in culture). The presence of GS in oligodendrocytes from both primary glial cell cultures and “pure” oligodendrocyte cultures was confirmed by double immunostaining with a rabbit antisheep GS and guinea pig antirat brain myelin 2′, 3′‐cyclic nucleotide 3′‐phosphodiesterase. In “pure” oligodendrocyte cultures, about half of cells were labeled with anti‐GS antibody. Furthermore, on the immunoblot performed with a rabbit antisheep GS, the GS protein in “pure” oligodendrocyte secondary cultures was visualized as a single band with an apparent molecular mass of about 43 KDa. In contrast, two protein bands for GS were observed in cultured astrocytes. On the immunoblot performed with a rabbit antichick GS, two immunopositive protein bands were observed: a major one migrating as the purified adult chick brain GS and a minor one with a lower molecular mass. Two similar immunoreactive bands were also observed in pure rat astrocyte cultures. Compared to pure rat astrocyte cultures, “pure” oligodendrocyte cultures of the same age displayed an unexpectedly high GS specific activity that could not be explained by astrocytic contamination of the cultures (less than 5%). As for cultured astrocytes, treatment of oligodendrocyte cultures with dibutyryl‐adenosine 3′: 5′‐cyclic monophosphate, triiodothyronine, or hydrocortisone increased significantly GS specific activity. Interestingly, epidermal growth factor, basic fibroblast growth factor, and platelet‐derived growth factor that increase the GS activity in astrocytes do not affect this activity in oligodendrocytes. Thus we confirm the finding of Warringa et al. (1988) that GS is also expressed in oligodendrocytes. We show that its activity is regulated similarly in astrocytes and oligodendrocytes by hormones, but that it is regulated differently by growth factors in these two cell types.