bluuebunny/crossref_metadata_2025_split huggingface dataset:
Differential regulation of glucose transporter 1 and 2 mRNA expression by epidermal growth factor and transforming growth factor‐beta in rat hepatocytes
Authors: Mischoulon David , Rana Basabi , Kotliar Natalio , Pilch Paul F. , Bucher Nancy L. R. , Farmer Stephen R. 
Published: February 2005

AbstractWe have examined by Northern blot analysis the expression of two members of the glucose transporter family of genes (GLUT‐1 and GLUT‐2) in regenerating liver and in hepatocytes cultured under various conditions. GLUT‐1, although thought to be a growth‐associated gene, is not expressed in normal or regenerating liver, whereas GLUT‐2, a liver‐specific gene, is abundant in normal liver and gradually up‐regulated during liver regeneration. Conversely, in hepatocytes cultured conventionally on dried rat tail collagen (RTC) in the presence of EGF and insulin, which potentiate proliferation, GLUT‐1 mRNA is rapidly and abundantly expressed, whereas GLUT‐2 is depressed. To investigate the causes of this “switch” in glucose transporter expression seen when hepatocytes are removed from the liver and cultured under the conventional proliferative conditions, we examined the effects of specific growth factors and extracellular matrices on cultured hepatocytes. EGF, a potent liver mitogen, although causing a threefold induction of GLUT‐1, was found to have no effect on GLUT‐2 expression, suggesting that the increase in GLUT‐2 seen in regenerating liver is not due to EGF. Inhibition of protein synthesis by cycloheximide in cultured hepatocytes does not prevent the induction of GLUT‐1 mRNA. In addition, treatment of cells with cycloheximide appears to stabilize the GLUT‐2 mRNA, preventing the usual down‐regulation of this gene in cultured hepatocvtes. The expression of the two glucose transporter mRNAs also differed when the hepatocytes were adherent to particular cell matrices. Culture of hepatocytes on a reconstituted basement membrane gel matrix (EHS) is known to restrain their growth and mediate high levels of differentiated hepatocytic functions that are lost under conventional culture conditions. Unlike cells on RTC, hepatocytes on EHS expressed low levels of GLUT‐1 mRNA, and decreased GLUT‐2 mRNA. TGF‐β, an attenuator of DNA synthesis, when added to cultures on RTC, substantially down‐regulated GLUT‐2 but had no effect on GLUT‐1. We propose that the effectors, EGF, TGF‐β and basement membrane components, play a significant role in the regulation of expression of GLUT‐1 and GLUT‐2 in hepatocytes. © 1992 Wiley‐Liss, Inc.
Effect of L‐fucose on proliferation and myo‐inositol metabolism in cultured cerebral microvessel and aortic endothelial cells
Authors: Stefani Mark R. , Dunlap Joyce A. , Yorek Mark A. 
Published: February 2005

AbstractDecreased myo‐inositol metabolism possibly contributes to the development of diabetic complications including micro and macrovascular disease. Previous studies have shown that hyperglycemia may be partially responsible for this defect. We have found that L‐fucose, a monosaccharide present in low concentrations in normal circulation and found to be elevated in diabetes, causes defects in cultured endothelial cells, including alterations in myo‐inositol metabolism and proliferation. Murine cerebral microvessel and bovine aortic endothelial cells take up L‐fucose from the medium in a time and concentration‐dependent manner. Both acute and chronic exposure of these cultured endothelial cells to media containing L‐fucose at concentrations that may exist in diabetic sera cause a significant decrease in the accumulation of myo‐inositol and its incorporation into inositol phospholipids. There is a concomitant decrease in the intracellular levels of myo‐inositol. Kinetic analysis of the effect of L‐fucose on myo‐inositol uptake suggests that L‐fucose competitively inhibits the transport of myo‐inositol, exhibiting a Ki in the range of 1.6–4.1 mM for both cell types. Endothelial cells exposed to L‐fucose concentrations of 0.5–20 mM exhibit depressed rates of proliferation in a concentration‐dependent fashion. Furthermore, L‐fucose causes a concentration‐dependent decrease in synthesis of proteoglycan by cultured cerebral microvessel endothelial cells as measured by incorporation of 35S; however, this effect is not observed in the aortic endothelia. These data suggest that L‐fucose at concentrations that may exist in diabetic sera may impair myo‐inositol metabolism and proliferation of the vascular endothelium. © 1992 Wiley‐Liss, Inc.
Different organizational states of fodrin in cultured MDCK cells are induced by treatment with low pH, calmodulin antagonist TFP, and tumor promoter PMA
Authors: Huotari Virva , Sormunen Raija , Lehto Veli‐Pekka , Eskelinen Sinikka 
Published: February 2005

AbstractWe have investigated the molecular mechanisms underlying dynamic organization of the fodrin network by treating the epithelial MDCK cells with various agents affecting intracellular pH, intracellular calcium ion concentration, intracellular calmodulin, and protein kinase C (PKC) activity. Elevation of intracellular calcium level by A23187 or treatment with trifluoperazine (TFP), a calmodulin inhibitor, did not have any drastic effect on the fodrin distribution as judged by immunofluorescence microscopy. A long‐term incubation with phorbol‐12‐myristate‐13‐acetate (PMA), a protein kinase C activator, in contrast, released fodrin from the lateral walls of the MDCK cells, leading to a diffuse cytoplasmic distribution. TFP, along with PMA, accelerated destabilization of the fodrin skeleton. Treatment with TFP alone rapidly released the cells from the substratum, which, however, could be prevented by PMA. We have previously shown that lowering of intracellular pH (<6.5) leads to a removal of fodrin from its basolateral residence (Eskelinen et al., 1992) and that this translocation is reversed upon returning normal pH. We now show that the rebuilding of the membrane skeleton can be prevented if TFP is added to the acidified cells. Moreover, in TFP‐treated acidified cells, fodrin shows a clusterlike organization similar to that observed in resting lymphocytes. We also noticed that interconversions between these different organizational states of fodrin are independent of the intracellular calcium concentration. Thus manipulation of the intracellular pH and treatment with TFP and PMA reveals different organizational states of the fodrin skeleton. This suggests that fodrin may participate in PMA‐, TFP‐ and pH‐sensitive signal transduction pathways. © 1992 Wiley‐Liss, Inc.
Growth factors produced by human embryonic kidney cells that influence megakaryopoiesis include erythropoietin, interleukin 6, and transforming growth factor‐beta
Authors: Withy Raymond M. , Rafield Lori F. , Beck Anton K. , Hoppe Henry , Williams Neil , McPherson John M. 
Published: February 2005

AbstractPartially purified protein preparations containing megakaryocyte growth factor activity were prepared from human embryonic kidney (HEK) cell conditioned medium using ammonium sulfate precipitation, Cibicron blue affinity chromatography, and wheatgerm lectin affinity chromatography. Treatment of these preparations with neutralizing antibodies directed against erythropoietin (EPO) and interleukin 6 (IL6) resulted in a dramatic reduction in their capacity to stimulate megakaryocyte maturation in vitro. The presence of EPO in these preparations was confirmed by both immunoblotting and use of a mouse spleen erythroid progenitor cell proliferation assay routinely used to quantitate EPO activity in vitro. Northern blot analysis of HEK cell‐derived mRNA with IL6 DNA probes revealed the presence of an IL6 transcript with a molecular size of 1.3 kb. Analysis of the HEK cell‐derived preparation by ELISA confirmed the presence of immunologically reactive IL6. In addition, it was shown that purified recombinant human EPO and IL6 stimulated megakaryocyte maturation in the in vitro assay used in this study. These data indicate that the activity in HEK cell conditioned medium that stimulates megakaryocyte maturation in vitro is predominantly due to the presence of IL6 and EPO. Immunoneutralization studies of another HEK cell‐derived preparation, which was inhibitory in the megakaryocyte maturation assay, demonstrated that it contained transforming growth factor beta (TGFβ), a potent inhibitor of megakaryocyte maturation. Taken together, these studies indicate that HEK cell conditioned medium, which has previously been reported to contain megakaryocyte growth factor activity, is comprised of a complex mixture of growth and differentiation factors, some of which promote and others that inhibit the process of megakaryopoiesis. © 1992 Wiley‐Liss, Inc.
Transformed growth phenotype of mouse mammary epithelium in primary culture induced by specific fetal mesenchymes
Authors: Kanazawa Takuya , Hosick Howard L. 
Published: February 2005

AbstractWhen mesenchyme from fetal mammary or salivary gland is implanted into adult mouse mammary gland, adjacent epithelium responds with intense hyperplasia. The hyperplastic cells are more vulnerable than are non‐stimulated cells to transformation in vivo by a chemical carcinogen or by mammary tumor virus. This system provides a potentially useful model for determining how stroma contributes to mammary tumorigensis. We have developed co‐culture systems and used them to investigate in more detail the nature of the signal produced by the mesenchyme cells. Monolayers of mesenchyme cells were prepared on tissueculture wells. The mesenchyme cells were trapped on the surface by a thin overlay of agarose. Primary mammary epithelial cells were cultured atop this barrier layer, either as organoids in collagen gels for assessment of anchorage‐dependent growth, or as single‐cell dispersions in soft agarose for assessment of anchorage independent growth. Our procedures for assay of anchorage‐independent growth allow us for the first time to detect and measure this transformation‐defining characteristic in non‐immortalized mammary epithelial cells in primary culture. Fetal mammary fat pad precursor tissue and fetal salivary mesenchyme both stimulated anchorage‐dependent growth of mammary epithelium with cell number increasing as much as fifteenfold during a 6‐day culture period. These same fetal tissues also stimulated anchorage‐independent growth of the mammary epithelial cells, with colony‐forming efficiencies of up to 40% in co‐cultures with salivary mesenchyme. No colonies formed in the absence of mesenchyme. Cells of colonies contained keratin, which indicates that the colonies grew from epithelial cells and not from a contaminant of another cell type. When co‐cultured epithelial cells were subsequently re‐cultured in the absence of mesenchyme, they lost their ability to grow independent of anchorage. No colonies grew in co‐cultures with fetal cells from heart, kidney, or lung, which is consistent with the lack of stimulation by these tissues in the mammary gland in vivo. A tumor promoter, 12‐O‐tetradecanoylphorbol acetate (TPA), also caused anchorage‐independent growth of the dispersed mammary epithelial cells. Culture medium conditioned by primary or early‐passage salivary mesenchyme cells was capable of stimulating growth under both anchorage‐dependent and anchorage‐independent conditions, confirming that these effects are mediated by a paracrine factor. The results indicate that stimulatory fetal mesenchymes produce soluble molecules that act analogously to transforming growth factors. © 1992 Wiley‐Liss, Inc.
Biphasic effect of the mitotoxin bFGF‐saporin on bovine lens epithelial cell growth: Effect of cell density and extracellular matrix
Authors: David Thierry , Tassin Jacqueline , Lappi Douglas A. , Baird Andrew , Courtois Yves 
Published: February 2005

AbstractWe have studied the effect of a specific FGF receptor suicide antagonist on the growth of bovine epithelial cells (BEL cells) in culture. This basic fibroblast growth factor‐saporin conjugate (bFGF‐SAP) has a biphasic effect on bovine lens epithelial cells (BEL cells). Whereas 0.01 nM and 0.1 nM bFGF‐SAP stimulate BEL cells proliferation, 1 nM and 10 nM bFGF‐SAP have the predicted toxic effects on BEL cell growth. The toxicity of bFGF‐SAP is observed 2 to 3 days after the initial treatment and depends on cell density. Accordingly, the sensitivity of confluent cells to bFGF‐SAP is reduced compared to sparse cells. A time course analysis reveals that bFGF‐SAP is effective after a short exposure to cells and that its effects are not increased with longer treatments. Cell growth on bFGF‐SAP pretreated extracellular matrix (ECM) or posterior lens capsule (PLC) is also affected. Basic FGF‐SAP has been shown to bind to the extracellular material, allowing a modulation of lens cells migration and survival by a single treatment in vitro. This finding raises the possibility of its use in vivo to prevent capsules invasion by lens cells after cataract surgery. © 1992 Wiley‐Liss, Inc.
Cortical photons in early development of frog eggs: Comparative study of photon emission between nuclear division and cytoplasmic fission
Authors: Oonuki Makoto , Ryuzaki Masashi 
Published: February 2005

AbstractIn the study of cell division in the early development of frog eggs, cortical photon emission was investigated by converting small light emission from living cells into digital pulses of potentials and recording the integrals of these pulses (analog method), counting the number of pulses (photon‐counting method), and counting the number of integrated pulses (improved photon‐counting method). By the analog and improved photon‐counting methods, changes in photon emission due to cell division could be clearly detected. The emitted light increased about 5.10−19W at the start of a cleavage furrow. Rapid changes in chemical reactions causing photon emission were compared during nuclear division and cytoplasmic phases. This emission occured mainly in cytoplamic fission, the rate being greater than in nuclear division by a factor of about 2.9. Chemical reaction rates were shown to differ according to bulk emission, thus indicating the mechanisms for the reactions to also differ. © 1992 Wiley‐Liss, Inc.
Vascular endothelial growth factor induces interstitial collagenase expression in human endothelial cells
Authors: Unemori Elaine N. , Ferrara Napoleone , Bauer Eugene A. , Amento Edward P. 
Published: February 2005

AbstractVascular endothelial growth factor (VEGF) is a 45kDa secreted peptide that has potent mitogenic activity specific for endothelial cells in vitro and the ability to induce a strong angiogenic response in vivo. In the present study, 24 h treatment with VEGF resulted in a stimulation of expression of the metalloproteinase, interstitial collagenase, at the protein and mRNA levels 2.5–3.0‐fold in human umbilical vein endothelial cells but not in human dermal fibroblasts. The dose response curve for collagenase induction was biphasic with the peak stimulatory response obtained by treatment of cells with 10–100 ng/ml (0.2–2 nM) VEGF. The dose response curve for collagenase induction overlapped with, but was not identical to, the response curve for proliferation, which showed VEGF mitogenic activity between ≤ 0.1–50 ng/ml (≤ 0.002–1 nM). There was no induction seen in expression of other members of the matrix metalloproteinase family, including the 72kDa type IV collagenase, the 92kDa type V collagenase, or stromelysin. Expression of transcripts for the major metalloproteinase inhibitor, tissue inhibitor of metalloproteinases, was also unaltered by treatment with VEGF (1–200 ng/ml). These studies demonstrate that in addition to stimulating proliferation of endothelial cells, VEGF can also induce the expression of the only metalloproteinase that can initiate degradation of interstitial collagen types I‐III under normal physiological conditions. Both responses are likely to contribute to the angiogenic potential of this peptide. © 1992 Wiley‐Liss, Inc.
Nonreceptor mediated nuclear accumulation of insulin in H35 rat hepatoma cells
Authors: Harada Shuko , Loten Ernest G. , Smith Robert M. , Jarett Leonard 
Published: February 2005

AbstractWe previously demonstrated that insulin accumulated in the nucleus in several cell types and partially characterized the uptake mechanisms and pathways in H35 rat hepatoma cells. Nuclear accumulation of insulin was energy independent, time, temperature, and insulin concentration dependent, but apparently nonsaturable. This study investigated further the initial endocytotic pathways that contribute to the nuclear accumulation of insulin using trypsin treatment of the cells to prevent insulin binding to its plasma membrane receptor. Total cell‐associated, intracellular, and nuclear insulin were compared in control and trypsin‐treated H35 hepatoma cells. Trypsin treatment markedly decreased total cell‐associated and intracellular insulin as well as the nuclear accumulation of insulin when cells were incubated with 2.8 ng/ml insulin. When the cells were incubated with 100 ng/ml insulin, trypsin treatment totally inhibited insulin binding to the plasma membrane for at least 90 min. However, intracellular accumulation of insulin was reduced by only 50% at 60 min, and trypsin treatment failed to inhibit the nuclear accumulation of insulin. Chemical extraction and Sephadex G‐50 chromatography revealed nuclear associated insulin in trypsin‐treated cells was identical to that in control cells incubated with either 2.8 or 100 ng/ml insulin. These results suggest that a nonreceptor mediated uptake pathway, i.e., fluid‐phase endocytosis, contributed significantly to the nuclear accumulation of insulin at high insulin concentrations, but at lower insulin concentrations the receptor‐mediated pathway predominated. No matter which initial endocytotic route was used to internalize insulin, the insulin apparently associated with the same nuclear matrix proteins. This association of insulin with the nuclear matrix may be involved in regulation of nuclear events such as cell growth and differentiation or gene transcription. © 1992 Wiley‐Liss, Inc.
Alterations in cellular calcium handling as a result of systemic calcium deficiency in the developing chick embryo: II. Ventricular myocytes
Authors: Miyahara Takashi , Akins Robert E. , Tuan Rocky S. 
Published: February 2005

AbstractWe have previously shown that cardiovascular anomalies, such as hypertension and tachycardia, develop in Ca2+‐deficient, shell‐less (SL) chick embryos cultured ex ovo, accompanied by elevated circulating catecholamines and higher α‐adrenergic sensitivity of cardiovascular functions. Results described in the preceding work, using erythrocytes as an experimental system, show that cellular Ca2+ handling properties are also altered as a result of long‐term calcium deficiency. To examine the relevance of these findings to cells of the cardiovasculature, we have analyzed and compared the Ca2+ handling characteristics of the heart cells of SL and normal (NL) embryos. For this study, isolated and cultured ventricular myocytes of SL and NL embryos were loaded with Fura‐2 via transient membrane damage with glass beads. Compared to Fura‐2/AM, bead loading yielded similar values and kinetic profiles of [Ca2+]i‐dependent differential fluorescence and, in addition, did not affect cell viability and beating activity. The Fura‐2 loaded ventricular myocytes were washed in Ca2+‐free buffer and then analyzed by ratiometric fluorescence (350 nm/380 nm) microscopy for kinetic changes in [Ca2+]i (R350/380 values) as a function of [Ca2+]o and adrenergic modifiers. At 0.5 and 1.0 mM [Ca2+]o, SL cells showed significantly higher [Ca2+]i, higher beating rates, and faster rate of increase in [Ca2+]i compared to NL cells. At higher [Ca2+]o (3–5 mM), there was no significant difference in [Ca2+]i and beating rate between NL and SL cells. Treatment with norepinephrine (NE; 0.01–1 μM) at 1 mM [Ca2+]o substantially increased [Ca2+]i in both NL and SL cells. In the former, the NE effect was completely inhibited by β‐blockade (1 μM propranolol). In contrast, in SL cells, NE remained effective after β‐blockade, and combined α‐blockade (1 μM prazosin) and β‐blockade was needed to inhibit completely the NE effect. In both NL and SL cells, treatment with NE substantially increased beating rates in a similar manner. Taken together, these findings suggest that Ca2+ handling and adrenergic regulation of the heart cells are significantly altered in the SL embryos, and that these alterations may be related to the development of impaired cardiovascular functions resulting from systemic Ca2+ deficiency. © 1992 Wiley‐Liss, Inc.
Identification of the cell surface and nuclear receptors for NGF in a breast carcinoma cell line
Authors: Rakowicz‐Szulczynska Ewa M. 
Published: February 2005

Abstract125I‐nerve growth factor (NGF) was found to be internalized and translocated to the nucleus of SKBr5 breast carcinoma cells. The cytoplasm and chromatin isolated from nonmitotic cells accumulated two‐and five‐fold, respectively, more of 125I‐NGF than the cells undergoing mitosis. MAb 20.4 developed against the NGF cell surface receptor immunoprecipitated the 80,000 Mr receptor from plasma membrane and two protein species from the chromatin; 90,000 Mr (major band) and 200,000 Mr (minor band). In SKBr5 cells, binding of NGF to the chromatin did not affect synthesis of rRNA. Proliferation of SKBr5 cells was slightly stimulated by NGF. In control melanoma A875 cells, which express the 230,000 Mr chromatin receptor, NGF inhibited both rRNA synthesis and cell proliferation. We suggest that the 90,000 Mr chromatin receptor expressed by SKBr5 cells represents a “nonactive”, ligand‐binding subunit of the high molecular weight receptor for NGF. The critical role of the chromatin receptor for NGF in rRNA‐dependent cell proliferation is discussed. © 1993 Wiley‐Liss, Inc.
Growth inhibitory activity in extracts from human fibroblasts
Authors: Macieira‐Coelho Alvaro , Söderberg Anita 
Published: February 2005

AbstractIndirect evidence for the presence of a growth inhibitor in normal human fibroblasts has been obtained previously; the inhibitory activity has been found associated with crude cell extracts, but the molecule responsible for the growth inhibition has never been isolated. We have isolated a glycopeptide fraction from human fibroblast cultures, whose synthesis decreases when the cells are stimulated into the division cycle. It was separated by electric charge, lectin affinity, and molecular mass. When added to quiescent cells simultaneously with a growth stimulus, the glycopeptide reduces DNA synthesis activity. The relationship of the kinetics of the synthesis of the glycopeptide with the cell division cycle and its molecular weight are different from what has been described so far for other growth regulators. The decreased synthesis of this inhibitor, induced by growth factors, seems to be one of the requirements for the initiation of the division cycle by human fibroblasts. This response to growth factors was stable during the lifespan of the fibroblast population and became less pronounced only in cells at the end of their replicative potential. © 1993 Wiley‐Liss, Inc.