bluuebunny/crossref_metadata_2025_split huggingface dataset:
EGF and TGF‐alpha are potent chemoattractants for endothelial cells and EGF‐like peptides are present at sites of tissue regeneration
Authors: Grotendorst Gary R. , Soma Yoshinao , Takehara Kazuhiko , Charette Marc 
Published: February 2005

AbstractWe report here that epidermal growth factor (EGF) isolated from male mouse salivary glands is active as a potent chemoattractant for a rat heart vascular endothelial cell line (RHEC). Human EGF and transforming growth factor‐alpha (TGF‐alpha) produced in Escherichia coli are also active as chemoattractants for these cells. The chemotactic response of the cells is induced by concentrations of murine EGF that ranged from 0.01 to 1.0 nM with similar specific activities for both the human EGF and TGF‐alpha. Other growth factors such as fibroblast growth factor (FGF), TGF‐beta, platelet‐derived growth factor (PDGF), insulin, or IGF were not active as chemoattractants for these cells. The chemotactic response exhibited similar kinetics to those reported for connective tissue cells responding to PDGF (2–6 h) and required both RNA and protein synthesis. However, in contrast to NIH/3T3 cells responding to PDGF, the proliferating cultures of RHEC respond as well as growth‐arrested cultures in the chemotaxis assay. In in vivo studies of connective tissue regeneration it appears that EGF‐like factors are present in wound fluid collected from rats. Acid extracts of wound fluid contained a RHEC chemotactic activity that was neutralized with anti‐EGF antisera. These studies indicate that EGF‐like factors may function in vivo to control in part the angiogenic events that occur during tissue repair.
Bone marrow‐derived stromal cell line expressing osteoblastic phenotype in vitro and osteogenic capacity in vivo
Authors: Benayahu D. , Kletter Y. , Zipori D. , Wientroub S. 
Published: February 2005

AbstractMarrow stroma has been shown to have osteogenic potential. Here we report the characterization of a unique stromal cell line derived from mouse bone marrow (MBA‐15), which expresses osteoblastic phenotype in vitro and forms bone in vivo. More than 70% of cells in culture were histochemically positive for alkaline phosphatase. The enzyme levels were enhanced threefold when cultures were treated with dexamethasone. Gel electrophoresis of [3H]‐proline‐labeled cultures showed that MBA‐15 cells produced only type I collagen. These cells were responsive to PTH, as indicated by a 50‐fold increase in intracellular cAMP. Prostaglandin E2, but not calcitonin, stimulated cAMP up to 70‐fold. When cultures were grown to confluence and fed daily with ascorbic acid and β‐glycerophosphate, the cells formed a Von Kossa positive, thick extracellular matrix, shown to contain hydroxyapatite crystals. MBA‐15 cells produced mineralized bone when implanted in diffusion chambers. These results indicate that the MBA‐15 cell line possesses osteoblastic features in vitro and osteogenic capacity in vivo.
Basic fibroblast growth factor is released from endothelial extracellular matrix in a biologically active form
Authors: Presta M. , Maier J. A. M. , Rusnati M. , Ragnotti G. 
Published: February 2005

AbstractBasic fibroblast growth factor (bFGF) binds to heparin‐like molecules present in the extracellular matrix (ECM) of transformed fetal bovine aortic endothelial GM 7373 cells. Binding of bFGF to ECM can be competed by heparin or heparan sulfate, and ECM‐bound bFGF can be released by treating the cells with heparinase or heparatinase. After binding to ECM, bFGF is slowly released into the medium in a biologically active form, as shown by its capacity to induce an increase of cell‐associated plasminogen activator activity and cell proliferation. The increase is prevented upon removal of ECM‐bound bFGF by a neutral 2 M NaCI wash. Soluble heparin and heparan sulfate reduce the amount of ECM‐bound bFGF released into the medium, possibly competing with ECM polysaccharides for heparinase‐like enzymes produced by endothelial cells, suggesting that these enzymes are involved in the mobilization of ECM‐bound bFGF.
Regulation of the sodium‐potassium pump in cultured rat skeletal myotubes by intracellular sodium ions
Authors: Brodie Chaya , Sampson S. R. 
Published: February 2005

AbstractThe properties of the Na‐K pump and some of the factors controlling its amount and function were studied in rat myotubes in culture. The number of Na‐K pump sites was quantified by measuring the amount of [3H]ouabain bound to whole‐cell preparations. Activity of the pump was determined by measurement of oua‐bain‐sensitive 86Rb‐uptake and component of membrane potential. Chronic treatment of myotubes with tetrodotoxin (TTX), which lowers [Na]i, decreased the number of Na‐K pumps, the ouabain‐sensitive 86Rb uptake, and the size of the electrogenic pump component of Em. In contrast, chronic treatment with either ouabain or veratridine, which increases [Na+]i, resulted in an elevated level of Na‐K pump sites. This effect was blocked by inhibitors of protein synthesis. Neither rates of degradation nor affinity of pump sites in cells treated with TTX, veratridine, or ouabain differred from those in control cells. The number and activity of Na‐K pump sites were unaffected by chronic elevation in [Ca]i or chronic depolarization. We conclude that alterations in the level in intracellular Na ions play the major role in regulation of Na‐K pump synthesis in cultured mammalian skeletal muscle.
Enhanced messenger RNA stability and differentiation of HL 60 cells treated with 1, 25‐dihydroxyvitamin D3 and cordycepin
Authors: Mathew Poruneloor A. , Ellis Linda K. , Studzinski George P. 
Published: February 2005

AbstractThe effect of inhibitors of RNA synthesis on 1,25(OH)2 vitamin D3‐induced monocytic differentiation was studied in a well‐differentiating clone AB 47 of HL 60 cells. The concentrations of these inhibitors were chosen to permit the maintenance of cell viability for at least 48 hours, and resulted in 40‐60% inhibition of total cellular RNA synthesis. No impairment of 1,25(OH)2 vitamin D3‐induced monocytic differentiation was observed with all inhibitors tested, and the presence of cordycepin actually enhanced differentiation. The phenotypic evidence of monocytic differentiation correlated with the increased levels of mRNA for c‐fos and c‐fms measured by hybridization to appropriate nick‐translated cDNA probes. In contrast, nuclear run‐on experiments showed the expected inhibition of transcription of these genes by the compounds used. The data suggest that interference by external agents with transcription of genes essential for a differentiation program brings into play compensatory mechanisms which permit the program to continue. Thus, differentiation appears to have a high priority among various competing intracellular pathways in 1,25(OH)2 vitamin D3‐treated HL 60 cells. Stabilization of messenger RNA levels evident in this study may therefore represent a general cellular mechanism for the correction of unwanted effects of xenobiotics on the cell.
Growth‐inhibitory effect of TGF‐B on human fetal adrenal cells in primary monolayer culture
Authors: Riopel Lise , Branchaud Charlotte L. , Goodyer Cynthia G. , Adkar Vinita , Lefebvre Yves 
Published: February 2005

AbstractWe examined the effects of transforming‐growth factor‐B (TGF‐B) on growth ([3H]‐thymidine uptake) and function (dehydroepiandrosterone sulfate [DHAS] and cortisol production) of human fetal zone adrenal cells. Results indicate that TGF‐B significantly inhibits, in a dose‐related manner, both basal and epidermal growth factor (EGF)‐stimulated cell growth: IC50= 0.1–;0.25 ng/ml. EGF is ineffective in overcoming the inhibitory effect of TGF‐B, suggesting a noncompetitive antagonism between the two factors. Also, the inhibitory effect of TGF‐B is additive to that of adrenocorticotropic hormone (ACTH). On the other hand, TGF‐B (1 ng/ml) does not significantly change basal or ACTH‐stimulated DHAS or cortisol secretion. We conclude that, unlike its effect on other steroid‐producing cells, TGF‐B inhibits growth of fetal zone cells and does not appear to have a significant inhibitory effect on steroidogenesis.
Induction of glucose‐regulated proteins in Xenopus laevis A6 Cells
Authors: Winning R. S. , Heikkila J. J. , Bols N. C. 
Published: February 2005

AbstractWe have characterized the induction of glucose‐regulated proteins (GRPs) in Xenopus laevis A6 cells, a kidney epithelial cell line. Exposure of A6 cells to medium in which 2‐deoxyglucose replaced galactose resulted in enhanced synthesis of two proteins at 78 and 98 kd. The 78 kd protein was determined by two‐dimensional PAGE to consist of two isoelectric variants with pls of 5.3 and 5.2 whereas the 98 kd protein resolved into a single spot with a pl of 5.1. The 78 kd protein cross‐reacted with antiserum against chicken GRP78 (glucose‐regulated protein), suggesting that the Xenopus protein shares homology with a previously characterized GRP. This was supported by the finding that a rat GRP78 probe hybridized with a 2‐deoxyglucose‐inducible mRNA. Synthesis of the two proteins was also induced by tunicamycin, 2‐deoxygalactose, and dithiothreitol. However, the GRPs were not induced by glucosamine or calcium ionophore A23187 at concentrations and exposure periods that have previously been shown to elicit a GRP response in mammalian and avian cells. Enhanced synthesis of the two GRPs by 2‐deoxyglucose was transient, reaching maximal levels by 12‐24 h and decreasing to near control levels by 48 h. Removal of the stress at the point of peak synthesis resulted in decreased synthesis of both proteins within 6 h and a return to control levels within 24 h of recovery. These data suggest that Xenopus cells have a GRP response that is similar, but not identical, to that found in mammalian cells.
Effects of transforming growth factor β and epidermal growth factor on cell proliferation and the formation of bone nodules in isolated fetal rat calvaria cells
Authors: Antosz Mark E. , Bellows Carlton G. , Aubin Jane E. 
Published: February 2005

AbstractWhen cells enzymatically isolated from fetal rat calvaria (RC cells) are cultured in vitro in the presence of ascorbic acid and Na β‐glycerophosphate, discrete three‐dimensional nodules form with the histologic, immunohistochemical, and ultra‐structural characteristics of bone (Bellows et al; Calcified Tissue International 38:143–154, 1986; Bhargava et al., Bone, 9:155–163, 1988). Quantitation of the number of bone nodules that forms provides a colony assay for osteoprogenitor cells present in the RC population (Bellows and Aubin, Develop. Biol., 133:8–13, 1989). Continuous culture with either epidermal growth factor (EGF) or transforming growth factor beta (TGF‐β) results in dose‐dependent inhibition of bone nodule formation; however, the former causes increased proliferation and saturation density, while the latter reduces both parameters. Addition of EGF (48 h pulse, 2‐200 ng/ml) to RC cells at day 1 after plating results in increased proliferation and population saturation density and an increased number of bone nodules formed. Similar pulses at confluence and in postconfluent multilayered cultures when nodules first begin forming (approx. day 11) inhibited bone nodule formation and resulted in a smaller stimulation of cell proliferation. Forty‐eight hour pulses of TGF‐β (0.01‐1 ng/ml) reduced bone nodule formation and proliferation at all times examined, with pulses on day 1 causing maximum inhibition. The effects of pulses with TGF‐β and EGF on inhibition of nodule formation are independent of the presence of serum in the culture medium during the pulse. The data suggest that whereas EGF can either stimulate or inhibit the formation of bone nodules depending upon the time and duration of exposure, TGF‐B inhibits bone nodule formation under all conditions tested. Moreover, these effects on osteoprogenitor cell differentiation do not always correlate with the effects of the growth factors on RC cell proliferation.
Inositol 1,3,4,5‐tetrakisphosphate stimulates the initiation of DNA synthesis in Ca2+‐deprived rat liver cells
Authors: Hill Timothy D. , Zwiller Jean , Boynton Alton L. 
Published: February 2005

AbstractThe G1‐S boundary of non‐neoplastic cells requires extracellular Ca2+ for successful transition. Inositol 1,3,4,5‐tetrakisphosphate but not inositol 1,4,5‐trisphosphate can partially replace Ca2+ and stimulate the initiation of DNA synthesis of Ca2+‐deprived T51B rat liver cells but only if sufficient extracellular Ca2+ (i.e., 0.075 mM) is present. The potent tumor promoter and protein kinase C activator 12‐O‐tetradecanoylphorbol acetate is also capable of replacing ex‐tracellular Ca2+ and partially stimulating the initiation of DNA synthesis. In addition, both inositol‐1,3,4,5‐tetrakisphosphate and 12‐O‐tetradecanoylphorbol acetate added together elicit a full DNA synthetic response.
Asymmetric distribution of phospholipids in the membrane of vesicles released during in vitro maturation of guinea pig reticulocytes: Evidence precluding a role for “aminophospholipid translocase”
Authors: Vidal Michel , Sainte‐Marie Josette , Philippot Jean R. , Bienvenue Alain 
Published: February 2005

AbstractGuinea pig reticulocytes lose their transferrin (Tf) binding activity during maturation, in the form of vesicles (exosomes) released into the extracellular medium. Vesicles were prepared from cultures of reticulocytes to study the possible externalization of a particular membrane‐associated activity, i.e., that of “aminophospholipid translocase.” Analysis of the peptide composition of these vesicles revealed that the major proteins are the Tf receptor and another peptide (70kDa), which is probably the “clathrin‐uncoating ATPase” described by johnstone et al. (1987). The exosome had a lipid composition similar to erythrocyte membrane, although with a lightly but significantly lower phosphatidylethanolamine content. The aminophospholipid distribution in the vesicle membrane was determined by fluorescamine labeling. The exosomes showed an asymmetric aminophospho‐lipid distribution similar to that of erythrocytes. “Aminophospholipid translocase” activity was absent, as no transverse diffusion of spin‐labeled phospholipids occurred over more than 2 hours at 37°C.
Ultrastructural analysis of acute immune thrombocytopenia in mice: Dissociation between alterations in megakaryocytes and platelets
Authors: Stenberg Paula E. , Levin Jack 
Published: February 2005

AbstractThrombopoiesis was studied in mice after the induction of acute immune throm‐bocytopenia with platelet antiserum (PAS). Utilizing electron microscopy, we examined platelets and megakaryocytes (MK) obtained 4, 8, 12, 24, 48, 72, and 120 hr after administration of PAS. Four to 24 hr after injection of PAS, the majority of bone marrow MK were normal in size and organelle distribution. The demarcation membrane system (DMS) extended normally throughout the mature cell cytoplasm at these times. However, approximately 50% of MK observed 48 hr and 72 hr after injection of PAS were significantly larger than normal, and often had demarcation membranes confined to an area between a peripheral organelle‐deficient zone and a central nuclear zone. The median values for sectional areas of platelets obtained 8–72 hr after administration of PAS were significantly greater than the median value for sectional areas of platelets in a pooled control sample. The proportion of cytoplasm to surface‐connected canalicular system appeared greater than normal in most large platelets from the PAS samples; and increased numbers of profiles of Golgi complex and endoplasmic reticulum were observed. By 48 hr post‐injection of PAS (at which time the modal ploidy class of MK has shifted from 16N to 32N; Corash et al.,Blood 70:177, 1987), most platelets were normal in size and cytoplasmic appearance. At 120 hr post‐injection of PAS, virtually all platelets exhibited a normal size and complement of organelles, and MK also had returned to normal. Our data indicate that in response to acute thrombocytopenia, MK prematurely release platelets which differ from normal platelets in size and cytoplasmic appearance. There was a marked dissociation between alterations in platelets and MK, since a statistically significant increase in platelet sectional area occurred 40 hr before the shift in modal ploidy class of MK, and platelet size subsequently decreased toward normal during the period that has been shown to be associated with the maximum shift in MK ploidy. These results strongly suggest that the characteristics of platelet release do not depend on the ploidy or cytoplasmic characteristics of MK.
Modulation of TGF‐β type 1 receptor: Flow cytometric detection with biotinylated TGF‐β
Authors: Newman Walter , Beall L. Dawson , Bertolini Donald R. , Cone James L. 
Published: February 2005

AbstractTransforming growth factor β‐ type 1 (TGF‐β) was reacted with NHS‐biotin to yield a derivative of TGF‐β1 which was biotinylated on lysine residues. The biotinylated form of TGF‐β1 was separated from the unreacted material by reverse phase chromatography. In three separate bioassays, the derivatized peptide was as active as the starting material. The use of FITC‐avidin in conjunction with flow cytometry demonstrated that the binding of biotinylated TGF‐β to its receptor is saturable, competable, and specific. A 100‐fold molar excess of unde‐rivatized TGF‐β1 gave 85% inhibition of binding of the biotinylated peptide to the mink lung cell line CCL‐64, while TGF‐β2 showed no inhibition of binding, nor did insulin, calcitonin, or TGF‐α. Both CCL‐64 cells and human umbilical vein endothelial cells showed a density‐dependent down‐regulation of receptor expression in culture. Several factors were examined that might mediate this effect. The down‐regulation was shown not to be due to the secretion of an active form of TGF‐β1. The extracellular matrix from high‐density cells did not decrease expression of the receptor. Fibronectin, collagen, and gelatin were also unable to signal changes in receptor expression, even though in other systems such matrix components can regulate the responsiveness of cells to TGF‐β1. Lastly, staining simultaneously for DNA content and TGF‐β1receptor expression showed that there was no correlation between cell cycle and receptor levels.
Different pattern of differentiation in two LLC‐PK1 Clones
Authors: Van Den Bosch Ludo , De Smedt Humbert , Borghgraef Roger 
Published: February 2005

AbstractTwo clones (LD3 and LC3) were isolated from the established renal cell line LLC‐PK1. They differed with respect to the development of the Na+‐dependent α‐methyl‐D‐glucoside (AMG) uptake. The higher uptake capacity in LD3 as compared to LC3 was owing to the expression of a higher number of carrier molecules as was shown by the specific phlorizin binding. The intracellular cyclic AMP level, the D‐glucose concentration in the growth medium and the cell density could be excluded as the causes of the difference between both clones. We found that the faster development of the Na+‐dependent hexose carrier in LD3 was parallelled by a faster expression of trehalase in this clone. This suggests that the expression of both apical proteins is correlated. From the growth curves it was concluded that renewal of the undifferentiated population was roughly the same in both clones. The recruitment of cells from the undifferentiated to the growth arrested state seems also very similar as was deduced from the development of tight junctions and from the down‐regulation of the α‐methylaminoisobutyric acid (meAIB) uptake. The start of differentiation was identical as was shown by the similar rate of expression found for γ‐glutamyl transferase. The difference between both clones is most likely situated at the traverse to a fully differentiated cell. This process takes more time in LC3 than in LD3. Also the fully differentiated state seemed to be different in both clones. We conclude that the pattern of differentiated characteristics found in both clones is different and a model incorporating these differences is presented.
Cytochemical and molecular properties of simian virus 40 transformed Kaposi's sarcoma‐derived cells: Evidence for the secretion of a member of the fibroblast growth factor family
Authors: Werner Sabine , Hofschneider Peter Hans , Stürzl Michael , Dick Irene , Roth Willi Kurt 
Published: February 2005

AbstractContact‐inhibited Kaposi's sarcoma‐derived cells (KS cells) were transfected with Simian Virus 40 (SV40) DNA. Transformed cells (SV‐KSC) were selected for their capacity to form foci on monolayers of the low‐malignant KS cells. Isolated SV‐KSC foci were found to contain integrated SV40 DNA sequences and to express SV40 large T‐antigen. Several differentiation properties of KS cells are retained in the SV40 transformants, e.g., expression of vimentin and the endothelial cell marker BMA 120. In contrast to the maternal KS cells, SV‐KSC are capable of growing in platelet‐derived growth factor (PDGF)‐depleted plateletpoor‐plasma serum (PPPS) and in soft agar. However, they are not tumorigenic in nude mice. Expression of the oncogenes c‐myc, c‐N‐ras, c‐Ha‐ras, and p53 is significantly elevated in SV‐KSC, whereas c‐fos and c‐erb B expression is comparable to that of KS cells and fibroblasts. Conditioned medium from SV‐KSC can substitute for PDGF when PDGF‐dependent, nontransformed KS cells are grown in PPPS. Biochemical analysis of the SV‐KSC supernatant and PDGF A and B mRNA expression analysis provide evidence that the mitogenic activity is not due to a PDGF‐like growth factor. On the other hand, there is evidence to indicate that the SV‐KSC mitogen is a member of the fibroblast growth factor family. SV‐KSC represent an interesting model system for the study of different degrees of malignancy of cultured mesenchymal cells and especially provide an important source for the isolation of a potent growth factor for KS cells and other mesenchymal cells in vitro.
Internal binding sites for MSH: Analyses in wild‐type and variant Cloudman melanoma cells
Authors: Orlow Seth J. , Hotchkiss Sara , Pawelek John M. 
Published: February 2005

AbstractCloudman S91 mouse melanoma cells express both external (plasma membrane) and internal binding sites for MSH. Using 125I‐beta melanotropin (β‐MSH) as a probe, we report here an extensive series of studies on the biological relevance of these internal sites. Cells were swollen in a hypotonic buffer and lysed, and a particulate fraction was prepared by high‐speed centrifugation. This fraction was incubated with 125 l‐β‐MSH with or without excess nonradioactive β‐MSH in the cold for 2 hours. The material was then layered onto a step‐wise sucrose gradient (8–80%) and centrifuged (156,000g, 60 min); fractions were collected and counted in a gamma counter or assayed for various enzymatic activities. The following points were established: (1) Specific binding sites for MSH were observed sedimenting at an average density of 50% sucrose in amelanotic cells and at higher densities in melanotic cells. (2) These sites were similar in density to those observed when intact cells were labeled externally with 125I‐β‐MSH and then warmed to promote internalization of the hormone. (3) Most of the internal binding sites were not as dense as fully melanized melanosomes. (4) In control experiments, the MSH binding sites were not found in cultured hepatoma cells. (5) Variant melanoma cells, which differed from the wild‐type in their responses to MSH, had reduced expression of internal binding sites even though their ability to bind MSH to the outer cell surface appeared normal. (MSH‐induced responses included changes in tyrosinase, dopa oxidase, and dopachrome conversion factor activities, melanization, proliferation, and morphology.) (6) Isobutylmethylxanthine, which enhanced cellular responsiveness to MSH, also enhanced expression of internal binding sites. The results indicate that expression of internal binding sites for MSH is an important criterion for cellular responsiveness to the hormone.