bluuebunny/crossref_metadata_2025_split huggingface dataset:
Epidermal growth factor dependence and TGFα autocrine growth regulation in primary rat tracheal epithelial cells
Authors: Ferriola Patrice C. , Robertson Alice T. , Rusnak David W. , Diaugustine Richard , Nettesheim Paul 
Published: February 2005

AbstractWe have examined dependence of primary rat tracheal epithelial (RTE) on exogenous epidermal growth factor (EGF) and determined whether a TGFα autocrine pathway is operating in these cells. Primary RTE cells plated in serum free media (SFM) without EGF and bovine pituitary factor (BPE) show little proliferation compared to cultures propagated in media containing EGF/BPE (CSFM). Removal of EGF/BPE shortly after plating, however, results in significant proliferation, although plateau cell densities are reduced and cell morphology is significantly altered compared to cells propagated in CSFM. Addition of EGF and/or BPE to cultures propagated in SFM minus EGF/BPE restores maximum cell density. The concentration of TGFα peptide in media conditioned by cells propagated without EGF/BPE is lower than the concentration in the media of CSFM cultures. TGFα mRNA and protein levels are also significantly lower in cells late in culture compared to logarithmically growing cells regardless of the presence or absence of EGF/BPE. The proliferation of primary RTE cells propagated without EGF/BPE is inhibited by neutralizing TGFα antiserum and by a tyrphostin compound that blocks TGFα/EGF receptor tyrosine kinase activity. These results indicate that primary RTE cells utilize TGFα as an autocrine growth factor and that the autocrine pathway is regulated as a function of growth state of the cells. However, this pathway does not provide growth autonomy to primary RTE cells, since cultures remain dependent on exogenous EGF/BPE for sustained proliferation. © 1992 Wiley‐Liss, Inc.
Transforming growth factor‐β1 is a heparin‐binding protein: Identification of putative heparin‐binding regions and isolation of heparins with varying affinity for TGF‐β1
Authors: McCaffrey Timothy A. , Falcone Domenick J. , Du Baoheng 
Published: February 2005

AbstractPrevious studies indicated that a major factor in heparin's ability to suppress the proliferation of vascular smooth muscle cells is an interaction with transforming growth factor‐β1 (TGF‐β1). Heparin appeared to bind directly to TGF‐β1 and to prevent the association of TGF‐β1 with α2‐macroglobulin (α2‐M). The present studies indicate that 20–70% of iodinated TGF‐β1 binds to heparin‐Sepharose and the retained fraction is eluted with ∼0.37 M NaCI. Native, unlabelled platelet TGF‐β1, however, is completely retained by heparin‐Sepharose and eluted with 0.9–1.2 M NaCI. Using synthetic peptides, the regions of TGF‐β1 that might be involved in the binding of heparin and other polyanions were examined. Sequence analysis of TGF‐β1 indicated three regions with a high concentration of basic residues. Two of these regions had the basic residues arranged in a pattern homologous to reported consensus heparin‐binding regions of other proteins. The third constituted a structurally novel pattern of basic residues. Synthetic peptides homologous to these three regions, but not to other regions of TGF‐β1, were found to bind to heparin‐Sepharose and were eluted with 0.15 M‐0.30 M NaCI. Only two of these regions were capable of blocking the binding of heparin to 125I‐TGF‐β. Immobilization of these peptides, followed by affinity purification of heparin, indicated that one peptide was capable of isolating subspecies of heparin with high and low affinity for authentic TGF‐β1. The ability of TGF‐β1 to bind to heparin or related proteoglycans under physiological conditions may be useful in understanding the biology of this pluripotent growth and metabolic signal. Conversely, a subspecies of heparin molecules with high affinity for TGF‐β1 may be a factor in some of the diverse biological actions of heparin. © 1992 Wiley‐Liss, Inc.
Dissociation between parathyroid hormone‐stimulated cAMP and calcium increase in UMR‐106‐01 cells
Authors: Merritt Bradley S. , Yamaguchi Dean T. , Green Jacob , Kleeman Charles R. , Muallem Shmuel 
Published: February 2005

AbstractWe used the osteogenic sarcoma cell line, UMR‐106‐01, to determine whether the rise in free cytosolic Ca2+ concentration ([Ca2+]i) and cellular cAMP following PTH stimulation are able to be regulated independently. For this purpose, we compared the effect of a PTH antagonist, stimulation of protein kinase C, augmentation by prostaglandins, and the time course of desensitization of the two cellular responses. Two × 10−7 M of the PTH antagonist 8,18Nle 34Tyr‐bPTH(3–34) amide ([Nle, Tyr]bPTH(3–34)A) was required to inhibit 10−9 M bPTH(1–34)‐stimulated cAMP generation by 50%. 10−7 M bPTH(1–34) completely overcame the inhibition induced by 10−6 M [Nle, Tyr]bPTH(3–34)A. Only 7 × 10−8 M and 2.7 × 10−7 M [Nle, Tyr]bPTH(3–34)A were required to half maximally inhibit the [Ca2+]i increase evoked by 3 × 10−8 and 10−7 M bPTH(1–34), respectively. In addition, dissociation between [Ca2+]i and cAMP signals was observed when modulation by protein kinase C and prostaglandins was tested. Preincubation of the cells with 10 nM TPA for 5 minutes markedly inhibited the PTH‐evoked [Ca2+]i increase. Short incubation with PGF2α augmented the PTH‐evoked [Ca2+]i increase. Similar pretreatments had no effect on the PTH‐stimulated cAMP increase. Finally, preincubation with 1.5 × 10−9 M bPTH(1–34) for 20 minutes almost completely blocked the effect of 10−7 M bPTH(1–34) on [Ca2+]i, while preincubation with 5 × 10−9 M bPTH(1–34) for 4 hours was required to inhibit the effect of 10−8 M bPTH(1–34) on cAMP production by 50%. The differences in the regulation of the two PTH‐stimulated cellular signaling systems, in particular, the response to antagonists and the time course of desensitization, could be at the level of the PTH receptor(s) or at a postreceptor domain. © 1992 Wiley‐Liss, Inc.
Restoration of proliferating cell nuclear antigen (PCNA) complex formation in xeroderma pigmentosum group a cells following cis‐diamminedichloroplatinum (II)‐treatment by cell fusion with normal cells
Authors: Miura Masahiko , Domon Masaharu , Sasaki Takehito , Kondo Seiji , Takasaki Yoshinari 
Published: February 2005

AbstractWe examined the role of the factor deficient in xeroderma pigmentosum group A (XP‐A) cells in the formation of proliferating cell nuclear antigen (PCNA) complex with DNA in the DNA repair process in human fibroblasts following cisdiamminedichloroplatinum (CDDP)‐treatment. Immunofluorescence staining after methanol fixation was used to detect the PCNA complex formation. When quiescent normal cells were PCNA‐stained at 3 h after 100 μM CDDP treatment for 1 h, almost all nuclei of the cells showed a punctuated staining pattern. On the other hand, nuclei of XP‐A cells were not stained. These results were the same with the findings following 10J/m2 of ultraviolet light (UV)‐irradiation. The quantitative analysis of the PCNA immunofluorescence intensity of normal cells revealed that the mean intensity was increased by 4.8 times by the CDDP‐treatment and 6.1 times by the UV‐irradiation, compared with that of untreated cells. The intensities among nuclei ranged widely in both treatments. In contrast, the mean intensity was not increased in XP‐A cells by the same treatments. However, when XP‐A cells were fused with normal cells with polyethylene glycol (PEG) treatment, the nuclei of the XP‐A cells showed positive PCNA‐staining following CDDP‐treatment or UV‐irradiation in almost all cases. These results suggest that the PCNA complex formation may play a role in the DNA repair process after the step where the factor deficient in XP‐A cells is involved following CDDP‐treatment as well as following UV‐irradiation. © 1992 Wiley‐Liss, Inc.
Repetitive 5‐azacytidine treatments of Fao cells induce a stable and strong expression of gamma‐glutamyl transpeptidase
Authors: Baik Ja‐Hyun , Chikhi Naima , Bulle Frédérique , Giuili Galicia , Guellaën Georges , Siegrist Sylvie 
Published: February 2005

AbstractThe role of DNA methylation in the expression of the rat gamma‐glutamyl transpeptidase (GGT) gene was assessed in the Fao cell line using a hypomethylating agent, 5‐azacytidine. Ten repetitive treatments of the cells, with 8 μM 5‐azacytidine for 24 h, led to 13‐ and 80‐fold increases, respectively, in GGT activity and in GGT mRNA level. The DNA methylation patterns generated by the isoschizomeric restriction enzymes Hpa II and Msp I indicated that the GGT gene, highly methylated in Fao cells, became strongly demethylated after 5‐azacytidine treatments. Thus, DNA demethylation increases the expression of the GGT gene. 5‐Azacytidine treatments also increased, but to a lesser extent, mRNAs level for actin, albumin, mitochondrial aspartate aminotransferase, aldolase BmRNAs (12‐ to 16‐fold) as wells as for tubulin, gluthathione transferese, and tyrosine aminotransferase mRNAs (2‐to 5‐fold). The GGT gene expression was further studied in B4 cells, cloned from the demethylated Fao cell population. This clone B4 exhibited a stable and strong GGT activity and a highly demethylated GGT gene. Among the three GGT mRNA I, II, or III, transcribed from three different promoters of the single rat GGT gene, only mRNA III was detected in Fao cells and was increased in clone B4, indicating that the demethylation acts on the promoter for mRNA III. The analysis of the differentiation state of B4 cells, as compared to Fao cells, showed a loss of the regulation of GGT and aspartate aminotransferase genes by dexamethasone, as well as a loss of the gluconeogenic pathway. Interestingly, B4 cells have retained many other specific functions of hepatic differentiation and have acquired α‐fetoprotein expression; thus this clone exhibits the characteristics of a hepatic fetal phenotype. © 1992 Wiley‐Liss, Inc.
Platelet endothelial cell adhesion molecule, PECAM‐1, modulates cell migration
Authors: Schimmenti Lisa A. , Yan Horng‐Chin , Madri Joseph A. , Albelda Steven M. 
Published: February 2005

AbstractCell migration is an important process in such phenomena as growth, development, and wound healing. The control of cell migration is orchestrated in part by cell surface adhesion molecules. These molecules fall into two major categories: those that bind to extracellular matrix and those that bind to adjacent cells. Here, we report on the role of a cell‐cell adhesion molecule, platelet‐endothelial cell adhesion molecule‐1, (PECAM‐1), a member of the lg superfamily, in the modulation of cell migration and cell‐cell adhesion. PECAM‐1 is a 120–130 kDa integral membrane protein that resides on endothelial cells and localizes at sites of cell‐cell contact. Since endothelial cells express PECAM‐1 constitutively, we studied the effects of PECAM‐1 on cell‐cell adhesion and migration in a null‐cell population. Specifically, we transfected NIH/3T3 cells with the full length PECAM‐1 molecule (two independent clones). Transfected cells containing only the neomycin resistance gene, cells expressing a construct coding for the extracellular domain of the molecule, and cells expressing the neu oncogene were used as controls. The PECAM‐1 transfectants appeared smaller and more polygonal and tended to grow in clusters. Indirect immunofluorescence of PECAM‐1 transfectants showed peripheral staining at sites of cell‐cell contact, while the extracellular domain transfectants and the control cells did not. In two quantitative migration assays, the full‐length PECAM‐1 transfectants migrated more slowly than control cells. Thus, PECAM‐1 transfected into a null cell appears to localize to sites of cell‐cell contact, promote cell‐cell adhesion, and diminish the rate of migration. These findings suggest a role for this cell‐cell adhesion molecule in the process of endothelial cell migration. © 1992 Wiley‐Liss, Inc.
In vitro model of angiogenesis using a human endothelium‐derived permanent cell line: Contributions of induced gene expression, G‐proteins, and integrins
Authors: Bauer Jeffrey , Margolis Michael , Schreiner Clara , Edgell Cora‐Jean , Azizkhan Jane , Lazarowski Eduardo , Juliano R. L. 
Published: February 2005

AbstractThe EA hy926 cell line is a continuous, clonable, human cell line that displays a number of features characteristic of vascular endothelial cells (Edgell et al., 1983). Here we report that when EA hy926 cells (EA cells) are plated on an extracellular matrix material [Matrigel®], they undergo a process of morphological re‐organization leading to the formation of a complex network of cord or tubelike structures. These events seem to resemble, in some respects, an in vitro process of angiogenesis. The morphological re‐arrangement occurs within a 12–16 hr period and seems to require expression of new messenger RNA and protein, since it is completely blocked when actinomycin D or cycloheximide are present at the time the cells are plated on Matrigel. This is not due to overt toxicity of the drugs, since exposure of cells to actinomycin D at 2 hr or more after plating on Matrigel has little effect on the formation of the tubelike structures. The process of Matrigelinduced tube formation also apparently involves a G‐protein mediated signal. Treatment of the EA cells with pertussis toxin completely blocks the process and causes the ADP‐ribosylation of a 42 kD protein that is recognized by an antibody to Gi‐alpha subunits. In contrast, concentrations of pertussis toxin sufficient to block tube formation have only modest effects on the adhesion or motility of EA cells on purified matrix components such as laminin or collagen IV. The process of Matrigel‐induced tube formation also involves integrins since monoclonal antibodies to integrin alpha6 or beta 1 subunits can completely block the process. The concentrations of anti‐integrin antibodies needed to block tube formation are much lower than those required to block cell adhesion on purified matrix components and are sufficient to occupy less than 10% of the alpha6 or beta 1 subunits available at the cell surface. These results suggest that integrins may be involved in this potential model of angiogenesis in processes beyond their usual role in cell adhesion. Based on these results, it seems likely that the EA hy 926 cell line will prove to be a useful model for in vitro study of angiogenic processes. © 1992 Wiley‐Liss, Inc.
Electrochemical potentials of potassium in skeletal muscle under different metabolic states
Authors: Khuri Raja N. , Agulian Samuel K. , Abdulnour‐Nakhoul Solange , Nakhoul Nazih L. 
Published: February 2005

AbstractIntracellular potassium and membrane potential were measured simultaneously by means of double‐barrelled liquid ion‐exchange microelectrodes in single fibers of rat thigh muscle in vivo in rats maintained in seven different metabolic states. The K+ equilibrium potential (Ek) was more negative than the simulataneously measured membrane potential (Em) in the normal state by 18.4 mV. K+ loading, acute and chronic, resulted in depolarization of Em due to increased serum K+ (hyperkalemia) with no increase in intracellular K+. K+ depletion resulted in hyperpolarization of Em as plasma K+ decreased proportionately more than intracellular K+. Low Na+ diet had no effect. Intracellular K+ was decreased in acute acidosis but not in the chronic state. Thus K+ depletion and acute acidosis are associated with intracellular K+ decrease. The fact that hyperpolarization exists in the former and not the latter is a reflection that hypokalemia accompanies the former condition. The hyperpolarizing states of K+ depletion and chronic acidosis are accompanied by decreased excitability and muscle weakness. © 1992 Wiley‐Liss, Inc.
Establishment of a chinese hamster ovary cell line that expresses grp78 antisense transcripts and suppresses A23187 induction of both GRP78 and GRP94
Authors: Li Li‐Jing , Li Xuan , Ferrario Angela , Rucker Natalie , Liu Eva S. , Wong Sam , Gomer Charles J. , Lee Amy S. 
Published: February 2005

AbstractGRP78, a 78,000 dalton protein residing in the endoplasmic reticulum, is postulated to play important roles in protein folding and cell survival during calcium and other physiologcial stress. Here we describe the construction of an eukaryotic expression vector for the constitutive expression of grp78 antisense RNA and the creation of a CHO cell line, 78WO, which expresses high levels of the grp78 antisense RNA through amplification of the stably transfected antisense vector. We observed that whereas 78WO maintains a basal levelof GRP78 similar to that of control cells, GRP78 is no longer inducible by A23187. The 78WO cells have undergone a compensatory increase in grp78 transcription such that the effects of antisense are cancelled out at the protein level under nonstressed conditions. In these same cells, GRP94, a 94,000 dalton ER protein, is also rendered noninducible by A23187. This provides the first evidence that the regulation of two ER proteins might be coupled such that the failure to induce GRP78 results in the down‐regulation of GRP94. The 78WO cell line grows with a doubling time of about 26 hr and exhibits decreased tolerance to A23187, suggesting the GRPs contribute to cell viability under calcium stress. The establishment of this cell line, which can be stably maintained, will provide a useful tool for testing whether the induction of the GRPs is important for protein folding or transport and whether their enhanced synthesis is the cause or consequence of a variety of physiological adaptations. © 1992 Wiley‐Liss, Inc.
Interaction of endothelial cells with a laminin A chain peptide (SIKVAV) in vitro and induction of angiogenic behavior in vivo
Authors: Grant Derrick S. , Kinsella James L. , Fridman Rafael , Auerbach Robert , Piasecki Barbara A. , Yamada Yoshihiko , Zain Mona , Kleinman Hynda K. 
Published: February 2005

AbstractEndothelial cells are known to bind to laminin, and two peptides derived from the laminin A (CTFALRGDNP) and B1(CDPGYIGSR) chains block the capillary‐like tube formation on a laminin‐rich basement membrane matrix, Matrigel. In the present study, we have used various in vitro and in vivo assays to investigate the angiogenic‐biologic effects of a third active site in the laminin A chain, CSRARKQAASIKVAVSADR (designated PA22‐2) on endothelial cells. The SIKVAV‐containing peptide was as active as the YIGSR‐containing peptide for endothelial cell attachment but was less active than either the RGD‐containing peptide or intact laminin. Endothelial cells seeded on this peptide appeared fibroblastic with many extended processes, unlike the normal cobblestone morphology observed on tissue culture plastic. In addition, in contrast to normal tube formation on Matrigel, short irregular structures formed, some of which penetrated the matrix and sprouting was more apparent. Analysis of endothelial cell conditioned media of cells cultured in the presence of this peptide indicated degradation of the Matrigel and zymograms demonstrated active collagenase IV (gelatinase) at 68 and 62 Kd. A murine in vivo angiogenesis assay and the chick yolk sac/chorioallantoic membrane assays with the peptide demonstrated increased endothelial cell mobilization, capillary branching, and vessel formation. These data suggest that the ‐SIKVAV‐site may play an important role in initiating branching and formation of new capillaries from the parent vessels, a behavior that is observed in vivo in response to tumor growth or in the normal vascular response to injury. © 1992 Wiley‐Liss, Inc.
Cell cycle regulated expression of nucleolar antigen P120 in normal and transformed human fibroblasts
Authors: Fonagy Anna , Swiderski Carol , Wilson Amy , Bolton Wade , Kenyon Norma , Freeman James W. 
Published: February 2005

AbstractNormal and SV40 virus‐transformed WI‐38 human lung fibroblasts were serum starved and refed, or synchronized by double thymidine block and released from the block. At different time points in the cell cycle, steady state levels of P120 mRNA and P120 protein content of the cells were determined by densitometric scans of Northern and Western blots. At the same time points, [3H]thymidine uptake was measured and flow cytometric analysis performed for DNA content and P120 antigen staining. Levels of P120 protein and P120 mRNA were approximately 4 times greater in non‐synchronous, exponentially growing transformed cells than in similarly growing normal cells. Early G1‐cells, synchronized either with serum deprivation or with metabolic block, contained only a trace amount of P120 protein and mRNA. The P120 gene was transcribed early in G1 and P120 protein synthesis initiated in middle G1. A dramatic increase of P120 protein level occurred in S‐phase with a corresponding mRNA peak preceding the P120 protein peak. These results indicate that P120 is overexpressed in transformed WI‐38 cells and that P120 is temporally regulated during the cell cycle of both transformed and normal fibroblasts. The dramatic increase in P120 protein expression at the G1 to S boundary suggests that P120 may play a role in the regulation of cell cycle and increased nucleolar activity that is associated with cell proliferation. © 1993 Wiley‐Liss, Inc.
Pulsatile and steady flow induces c‐fos expression in human endothelial cells
Authors: Hsieh Hsyue‐Jen , Li Nan‐Qian , Frangos John A. 
Published: February 2005

AbstractThe effects of pulsatile and steady fluid flow on the mRNA levels of proto‐oncogenes c‐fos, c‐jun, and c‐myc in cultured human umbilical vein endothelial cells (HUVEC) were investigated. c‐fos mRNA levels in stationary cultures were very low. A 1 Hz pulsatile flow with an average shear stress of 16 dynes/cm2 induced a dramatic increase of c‐fos mRNA levels in HUVEC 0.5 h after the onset of flow, which declined rapidly to basal levels within 1 h. Steady flow with a similar shear stress also induced a transient increase of c‐fos mRNA levels, but to a lesser extent. In addition, increased, c‐fos mRNA levels were observed when low shear (2–6 dynes/cm2) was replaced by high shear (16–33 dynes/cm2). Pulsatile and steady flow caused a slight increase of c‐jun and c‐myc mRNA levels. The role of pulsatility was also investigated in platelet‐derived growth factor (PDGF) expression. Pulsatile flow induced a transient increase of PDGF A‐ and B‐chain mRNA levels with peaks at 1.5–2 h. Pulsatile flow, which was more stimulatory in mediating c‐fos expression, however, was less stimulatory than steady flow in mediating PDGF expression. By using various inhibitors, protein kinase C was found to be an important mediator in flow‐induced c‐fos expression, with the involvement of G proteins, phospholipase C, and intracellular calcium. Protein kinase C was previously shown as a possible major mediator in flow‐induced PDGF expression which, at least partly, appeared to follow the induction mechanism of c‐fos, suggesting a possible connection between c‐fos and PDGF induction. However, the c‐fos antisense treatment, which significantly inhibited c‐fos transcription, failed to block the flow‐induced PDGF expression, suggesting that flow‐induced c‐fos expression may not play an important role in the mechanism of flow‐induced PDGF expression. The difference in the induction of c‐fos and PDGF expression under pulsatile as compared to steady flow indicates of c‐fos and PDGF expression under pulsatile as compared to steady flow indicates that a complex, flow‐mediated regulatory mechanism of gene expression exists in HUVEC. The increased expression of these proto‐oncogenes mediated by flow may be important in regulating long‐term cellular responses. © 1993 Wiley‐Liss, Inc.