bluuebunny/crossref_metadata_2025_split huggingface dataset:
Signal perception of fibroblasts for directional migration to platelet‐derived growth factor in Boyden‐type chambers
Authors: Adelmann‐Grill Bernhard C. , Cully Zuzana 
Published: February 2005

AbstractFibroblast chemotaxis has usually been determined in Boyden‐type chambers with polycarbonate filters, assuming that a stable concentration gradient of the attractant develops that causes directional migration of the cells. This view has been repeatedly challenged, and development of such gradients in vivo is unlikely. The present experiments were designed to test if a stable concentration gradient was required for normal dermal fibroblasts to migrate toward platelet‐derived growth factor. It was found that a brief pulse of the attractant was required and sufficient to induce chemotaxis. The pulse had to contain a specific concentration of attractant and was ineffective when not unilateral. The observed effects could not be attributed to induction of random migration or migration on a mediator‐coated surface. It is not clear which machinery is regulating this cellular behaviour, but it is suggested that cells may migrate in vivo by similar mechanisms, because the establishment of stable concentration gradients of attractants in tissues is deemed unlikely.
Cyclic adenosine monophosphate‐mediated induction of F9 teratocarcinoma differentiation in the absence of retinoic acid
Authors: Goldstein Beth , Rogelj Snezna , Siegel Susan , Farmer Stephen R. , Niles Richard M. 
Published: February 2005

AbstractF9 teratocarcinoma stem cells differentiate into parietal endoderm‐like cells when given retinoic acid (RA) and dibutyryl cyclic adenosine monophosphate (DB‐cAMP). It is generally accepted that the stem cells are resistant to the action of cAMP alone and need to be primed by RA in order to respond to cAMP. In this report, we demonstrate that F9 stem cells differentiate into parietal endoderm‐like cells in the absence of exogenous RA when treated with cholera toxin and 1‐methyl,3‐isobutyl xanthine (CT/MIX) or 8‐bromo‐cAMP/MIX (8B2‐cAMP/MIX). Cells treated with CT/MIX or 8B2‐cAMP/MIX were morphologically similar to parietal endoderm‐like cells, produced high amounts of plasminogen activator, and synthesized both type IV collagen and laminin mRNA. Conversely, markers made in abundance by stem cells such as stage‐specific embryonic antigen (SSEA‐1) and an mRNA species of 6.8 kb (pST6‐135) were markedly reduced in CT/MIX‐treated cells. To prove that cAMP alone could induce differentiation Lipidex‐1000, a hydrophobio gel, was used to remove 80–;90% of the endogenous serum retinoids. F9 cells grown in this retinoid‐depleted serum and treated with 8B2‐cAMP/MIX differentiated to parietal endoderm‐like cells as shown by both dramatic changes in morphology and induction of type IV collagen mRNA. Our results indicate that the differentiation of F9 to parietal endoderm‐like cells can be induced by increased intracellular cAMP and is not strictly dependent on the addition of RA.
Low temperature blocks exit of pro‐opiomelanocortin from the endoplasmic reticulum but not subsequent delivery to the site of prohormone processing
Authors: Wattenberg Binks W. 
Published: February 2005

AbstractThe effect of reduced temperature on the delivery of the prohormone pro‐opiomelanocortin (POMC) to the site of prohormone processing was investigated in the mouse anterior pituitary cell line AtT20. At 20°C processing was substantially inhibited and was almost completely arrested at 18°C. Earlier studies with membrane glycoproteins indicated that at these temperatures protein movement was blocked at the level of exit from the Golgi apparatus. In contrast it was found here that the inhibition of processing at reduced temperature was due to the retention of POMC in the endoplasmic reticulum. When POMC was allowed to progress to the Golgi before temperature was reduced, subsequent processing was only slightly retarded by incubation at 18°C. This indicates either that Golgi exit is not inhibited at this temperature, or that the processing apparatus exists in the Golgi. A surprising incidental result was that when held in the endoplasmic reticulum at low temperature POMC is apparently subject to post‐translational N‐linked glycosylation.
Quantification of protein transcytosis in the human colon carcinoma cell line CaCo‐2
Authors: Heyman Martine , Crain‐Denoyelle Anne‐Marie , Nath Samir Kumar , Desjeux Jehan‐Francois 
Published: February 2005

AbstractThe transepithelial absorption of food‐type proteins has been shown to proceed by endocytosis along two functional pathways: a minor direct pathway allowing transport of intact protein and a major lysosomal degradative pathway. The human colon carcinoma cell line CaCo‐2 grown on Millipore filters was used here further to characterize these pathways by measuring HRP transport across the cell monolayer in Ussing chambers. In the apical‐basal direction, this transport mainly occurred along the degradative pathway and was inhibited at 4°C (7.41 ± 1.26 pmoles/h·cm2 vs. 27.40 ± 8.91 at 37°C). The amount conveyed via the direct pathway was very small (0.89 ± 0.35 pmoles/h·cm2) and did not diminish at 4°C (1.43 ± 0.59 pmoles/h·cm2). In the basal‐apical direction, HRP transport along the degradative pathway at 37°C was similar to the apical‐basal value and was inhibited at 4°C (16.40 ± 4.50 vs. 2.72 ± 2.52 pmoles/h·cm2), but along the direct pathway, it was eight times the apical‐basal value (8.36 ± 3.11 pmoles/h·cm2) and was inhibited at 4°C (2.43 ± 0.78 pmoles/h·cm2). Intact HRP fluxes were not correlated with the eletrical resistance of the filters, indicating transport via a transcellular route. Monensin at 10−5 M did not affect direct or degradative transport in the apical‐to‐basal direction. These results suggest that in CaCo‐2 cells HRP undergoes bidirectional transcytosis by a fluid‐phase mechanism, but the extent of degradation during that transport varies according to the membrane (apical or basal) where it is presented.
Volume regulation in lens epithelial cells and differentiating lens fiber cells
Authors: Beebe David C. , Parmelee Judith T. , Belcher Karla S. 
Published: February 2005

AbstractPrevious studies from our laboratory have led us to conclude that lens cell elongation is caused by an increase in cell volume. This volume increase results from an increase in the potassium content of the cells due to decreased potassium efflux. In contrast, an increase in the volume of most cggers a regulatory volume decrease (RVD) that is usually mediated by increased potassium efflux. For this reason, chicken embryo lens epithelial cells were tested to see whether they were capable of typical cell volume regulation. Changes in cell volume during lens fiber differentiation were first estimated by 3H2O water uptake. Cell water increased in proportion to cell length in elongating lens cells. Treatment of epithelial cells cultured in basal medium with dilute or concentrated medium, or with medium containing 50 mM sucrose, resulted in typical volume regulatory responses. Cells lost or gained volume in response to osmotic stress, then returned to their previous volume. In addition, the elongation and increase in cell volume that accompanies lens fiber cell differentiation occurred normally in either hypoor hypertonic media. This observation showed that the activation of mechanisms to compensate for osmotic stress did not interfere with the increase in volume that accompanies elongation. The ability of elongating cells to volume regulate was also tested. Lens epithelial cells were stimulated to elongate by exposure to embryonic vitreous humor, then challenged with hypotonic medium. These elongating cells regulated their volume as effectively as unstimulated cells. Therefore, cells that were increasing their volume due to reduced potassium efflux could adjust their volume in response to osmotic stress, presumably by increasing potassium efflux. This suggests that the changes in potassium efflux that occur during differentiation and RVD are regulated by distinct mechanisms.
Impaired carcinoembryonic antigen release during the process of suramin‐induced differentiation of the human colic adenocarcinoma cell clone HT29‐D4
Authors: Fantini Jacques , Rognoni Jean‐Baptiste , Theveniau Magali , Pommier Gilbert , Marvaldi Jacques 
Published: February 2005

AbstractThe establishment of a differentiated state of the human colic adenocarcinoma cell clone HT29‐D4 can be obtained by two ways: 1) the removal of glucose and its replacement by galactose in the culture medium (Fantini et al.:Biology of the Cell65:163–169, 1989); 2) the addition of suramin, a polyanionic compound, in the glucose‐containing medium (Fantini et al.:Journal of Biological Chemistry264:10282–10286, 1989). We investigated the release of CEA in the culture medium of glucose‐deprived HT29‐D4 cells (HT29‐D4‐Gal) and studied its alteration in suramin‐treated HT29‐D4 cells (HT29‐D4‐S). The amount of CEA released in the medium in function of time in culture of undifferentiated HT29‐D4‐Glu cells was very low (5 to 8 ng/106 cells/24 hours) and almost constant throughout the experiment whereas it increased sharply during differentiation of HT29‐D4‐Gal cells (380 ng/106 cells/24 hours after 9 days in culture). Surprisingly the amount of CEA released by differentiated HT29‐D4‐S cells remained very low and comparable with the one of HT29‐D4‐Glu cells. Moreover suramin, when added to CEA‐producing HT29‐D4‐Gal cells, strongly inhibited its release.Radioiodination of cell surface proteins followed by immunoprecipitation using an anti‐CEA monoclonal antibody showed the presence of a 180 kDa poly‐peptide, i.e., CEA, predominantly labeled in HT29‐D4‐Gal and ‐S cells. The total CEA cellular content was higher in HT29‐D4‐Glu and HT29‐D4‐S cells than in HT29‐D4‐Gal cells. When HT29‐D4‐Gal or ‐S cells were treated with the bacterial phosphatidylinositol phospholipase C (PI‐PLC) a similar level of CEA was released suggesting a similar type of CEA anchorage. The present data demonstrate that a decrease in CEA release (i.e., in HT29‐D4‐Glu and ‐S cells) corresponds to an increase in its overall cellular expression. These results are in favour of a regulatory mechanism, impaired by suramin, which determines the balance between the soluble and the membrane bound forms of CEA.
Frequent expression of receptors for granulocyte‐macrophage colony‐stimulating factor on human nonhematopoietic tumor cell lines
Authors: Miyagawa Kiyoshi , Chiba Shigeru , Shibuya Kyoichi , Piao Yun‐Feng , Matsuki Shigeru , Yokota Jun , Terada Masaaki , Miyazono Kohei , Takaku Fumimaro 
Published: February 2005

AbstractReceptors for granulocyte‐macrophage colony‐stimulating factor (GM‐CSF) were identified on 9 of 35 (26%) human nonhematopoietic tumor cell lines including non‐small cell lung cancer, stomach cancer, colon cancer, and osteosarcoma cells. GM‐CSF receptors distributed on these human tumor cells were low affinity types with an equilibrium dissociation constant of 1.5–10.0 nM. Cross‐linking studies revealed that the molecular weights of the low affinity GM‐CSF receptors were 65–85 kilodaltons. The high affinity receptors identified on hematopoietic cells were not detected on human nonhematopoietic tumor cells which we studied, and we could detect no effects of GM‐CSF on cell growth of these tumor cells.
Insulin‐like growth factor‐1 (IGF‐1), insulin, and epidermal growth factor (EGF) are survival factors for density‐inhibited, quiescent Balb/c‐3T3 murine fibroblasts
Authors: Tamm Igor , Kikuchi Toyoko 
Published: February 2005

AbstractThe great majority of murine Balb/c‐3T3 fibroblasts in density‐inhibited, quiescent cultures disintegrate and die rapidly when cells are deprived of serum in the medium. Platelet‐derived growth factor (PDGF, 5 ng/ml) used alone and insulin‐like growth factor (IGF‐1, 40 ng/ml) + epidermal growth factor (EGF, 10 ng/ml) prevent most of this cell death and all three factors used together protect close to all cells in the confluent monolayer as determined by counting trypsinized cell suspensions in a Coulter counter. IGF‐1 used alone affords a high level of protection during the first 5 hours of incubation in serum‐free medium but the protective effect declines subsequently unless EGF is also present. EGF alone has little protective activity. The survival‐promoting activity of PDGF used alone or of PDGF + EGF + IGF‐1 is not significantly decreased by selective inhibition of messenger precursor RNA transcription with 5,6‐dichloro‐1‐β‐D‐ribofuranosylbenzimidazole (DRB,20 or 40 μM), which prevents G1 traverse of the cells mediated by the combination of the three growth factors. DRB also does not interfere with the early protective effect of IGF‐1 + EGF, but decreases the late protective effect of this growth factor combination. DRB by itself decreases cell viability in the absence of growth factors or serum. In these experiments viability was assayed by neutral red uptake by using an automated microplate reader. Inhibition of protein synthesis with cycloheximide (CHX, 1 or 5 μg/ml) over a 20‐hour period was associated with decreased survival of cells protected by IGF‐1 + EGF or PDGF + EGF + IGF, but also with decreased survival of cells incubated in the absence of growth factors or serum. The decrease in survival was somewhat more marked when IGF + EGF was present than when PDGF + EGF + IGF‐1 was present. Insulin (1,500 ng/ml) mimics the action of IGF‐1 (40 ng/ml). The cell survival‐enhancing activities of growth factors are concentration dependent. The evidence presented indicates that PDGF, EGF, and IGF‐1 (or insulin) act through distinctive mechanisms in affording protection of cells against death. The short‐term protective effects of the growth factors are independent of gene expression and may be mediated via metabolic events. Long‐term protection may be dependent on gene expression, especially in the case of IGF‐1 + EGF.
Lipoxin A4 and lipoxin B4 stimulate the release but not the oxygenation of arachidonic acid in human neutrophils: Dissociation between lipid remodeling and adhesion
Authors: Nigam Santosh , Fiore Stefano , Luscinskas Francis W. , Serhan Charles N. 
Published: February 2005

AbstractThe profiles of actions of lipoxin A4 (LXA4) and lipoxin B4 (LXB4), two lipoxygenase‐derived eicosanoids, were examined with human neutrophils. At nanomolar concentrations, LXA4 and LXB4 each stimulated the release of [1‐14C]arachidonic acid from esterified sources in neutrophils. Lipoxin‐induced release of [1‐14C]arachidonic acid was both dose‐ and time‐dependent and was comparable to that induced by the chemotactic peptide f‐met‐leu‐phe. Time‐course studies revealed that lipoxin A4 and lipoxin B4 each induced a biphasic release of [1‐14C]arachidonic acid, which was evident within seconds (5–15 sec) in its initial phase and minutes (>30 sec) in the second phase. In contrast, the all‐trans isomers of LXA4 and LXB4 did not provoke [1‐14C]AA release. Lipoxin‐induced release of arachidonic acid was inhibited by prior treatment of the cells with pertussis toxin but not by its β‐oligomers, suggesting the involvement of guaninine nucleotide‐binding regulatory proteins in this event. Dual radiolabeling of neutrophil phospholipid classes with [1‐14C]arachidonic acid and [3H]palmitic acid showed that phosphatidylcholine was a major source of lipoxin‐induced release of [1‐14C]arachidonic acid. They also demonstrated that lipoxins rapidly stimulate both formation of phosphatidic acid as well as phospholipid remodeling. Although both LXA4 and LXB4 (10−8‐10−6M) stimulated the release of [1‐14C]arachidonic acid, neither compound evoked its oxygenation by either the 5‐or 15‐lipoxygenase pathways (including the formation of LTB4, 20‐COOH‐LTB4, 5‐HETE, or 15‐HETE). LXA4 and LXB4 (10−7M) each stimulated the elevation of cytosolic Ca2+ as monitored with Fura 2‐loaded cells, albeit to a lesser extent than equimolar concentrations of FMLP. Neither lipoxin altered the binding of [3H]LTB4 to its receptor on neutrophils. In addition, they did not stimulate aggregation or induce adhesion of neutrophils to human endothelial cells. Results indicate that both LXA4 and LXB4 stimulate the rapid remodeling of neutrophil phospholipids to release arachidonic acid without provoking either aggregation or the formation of lipoxygenase‐derived products within a similar temporal and dose range. Together they indicate that LXA4 and LXB4 display selective actions with human neutrophils and suggest that these eicosanoids possess unique profiles of action which may regulate neutrophil function during inflammation.
Studies on the cell‐cycle dependency of the actions of insulin‐like growth factor‐I in Balb/c 3T3 cells
Authors: Kojima Itaru , Kitaoka Masafumi , Ogata Etsuro 
Published: February 2005

AbstractThe present study was conducted to determine the cell‐cycle dependency of various actions of IGF‐I in Balb/c 3T3 cells. When autophosphorylation of the IGF‐I receptor was determined in [32P]‐labelled cells, IGF‐I increased radioactivity in a 100 K‐Da phosphoprotein, presumably β‐subunit of the IGF‐I receptor, both in quiesent and in primed competent cells. Likewise, IGF‐I stimulated uptake of [3H]deoxyglucose independent of the cell cycle. In contrast, IGF‐I increased calcium entry, radioactivity in [3H]diacylglycerol, and [3H]thymidine incorporation in primed competent cells while these reactions were not induced by IGF‐I in quiescent cells. The latter three reactions were attenuated when cells were pretreated with pertussis toxin. These results indicate that some, but not all, of the actions of IGF‐I are dependent on the cell cycle in Balb/c 3T3 cells. They also suggest that a pertussis‐toxin‐sensitive G protein may be involved in the cellcycle‐dependent actions of IGF‐I.
Differential effects of transforming growth factor‐β and epidermal growth factor on the cell cycle of cultured rabbit articular chondrocytes
Authors: Vivien Denis , Galéra Philippe , Lebrun Emmanuel , Loyau GéRard , Pujol Jean‐Pierre 
Published: February 2005

AbstractWe examined the effect of transforming growth factor (TGF‐β) on the proliferative rate and cell cycle of cultured rabbit articular chondrocytes using cell counting, cytofluorometry, and [3H]‐thymidine incorporation. In the presence of 2% or 10% FCS (fetal calf serum), TGF‐β at 0.01, 0.1, 1, and 10 ng/ml had an inhibitory effect on cell proliferation after 24 h exposure with a dose dependence only for 2% FCS. Flow cytometric analysis of cell DNA content at that time showed that a high proportion of cells were arrested in late S‐phase (SQ or G2Q) in either 2% or 10% FCS‐containing medium. In both cases, a disappearance of the cell blockage occurred between 24 and 48 h after TGF‐β addition. However, whereas a stimulation of cell proliferation rate was observed at that time in cultures containing 10% FCS, a dose‐dependence inhibition of cell growth was detected, in contrast, for 2% FCS‐treated cells. Presence of TGF‐β during the last 24 h was not necessary to release the arrested cells. Furthermore, platelet‐poor plasma at 10% produced the same effects as FCS, suggesting that platelet‐derived factors, such as platelet‐derived growth factor (PDGF), could not be responsible for the release of blocked cells in this case. We compared the effect of TGF‐β to that of epidermal growth factor (EGF), used at an optimal concentration (10 ng/ml). In both slowly growing (2% FCS) and proliferating chondrocytes (10% FCS), EGF caused a significant increase of cell proliferation as early as 24 h. No arrest in late S‐phase but an augmentation of the percentage of cells in S‐ and G2M‐phases were observed. When combined, TGF‐β and EGF did not induce synergistic effect on the chondrocyte proliferation, as estimated by cell counting. [3H]‐thymidine labeling showed that the factors induced identical maxima of incorporation but the peak occurred earlier for TGF‐β than for EGF (approximately 6 h versus 12 h, respectively). Although both factors induce similar cell‐number increases at 48 h in 10% FCS‐containing medium, these proliferative effects were due to different actions on the cell cycle. The present study indicates that TGF‐β induces first a recruitment of chondrocytes in noncycling SQ‐ or G2Q‐blocked cells. Then, the release of these cells may produce either apparent stimulation of cell proliferation if sufficient levels of an unknown serum factor are present (10% FCS) or an inhibition of growth rate when only reduced amounts of this factor are available (2% FCS). This mechanism is quite different from that of EGF, which make more cells enter S‐phase, whatever the serum concentration in medium, and might be related to the transforming capacity of TGF‐β.
Ionic regulation of the plasma membrane potential of rainbow trout (Salmo gairdneri) spermatozoa: Role in the initiation of sperm motility
Authors: Gatti Jean‐Luc , Billard Roland , Christen Richard 
Published: February 2005

AbstractThe ionic dependence of the trout sperm plasma membrane potential was analysed by measuring the accumulation of the lipophilic ions 3H‐tetraphenylphosphonium (TPP) and 14C‐thiocyanate (SCN) following dilution in artificial media isotonic to the seminal fluid. Our data showed that the trout sperm plasma membrane has a mixed conductance: the plasma membrane potential is sensitive upon the transmembrane gradients of K+, Na+, and H+. This potential is negative (< −40mV) in a 125 mM choline chloride media (ChM) at pH 8.5. Replacement of choline by sodium has a small depolarizing effect. The membrane potential is about −15 mV in a 125 mM potassium chloride and falls near zero mV only if valinomycin is added. In ChM changing the external pH (pHe) greatly affects the membrane potential: its value rises from less than −40 mV at pHe 9.0 to −17 mV at pHe 5.0. This pH effect is observed also in presence of sodium or potassium. A decrease in the transmembrane proton gradient produced by increasing internal pH without changing pHe induces also a depolarisation of the plasma membrane. In the different media in which trout sperm remain immotile after dilution (media with [K+]>20–40 mM or a pH <7.5) the plasma membrane is more depolarized than in media allowing motility, suggesting a relationship between the state of membrane polarization and the intracellular effectors of the axonemal movement.
Spontaneous and cAMP‐dependent induction of a resting phase and neurite formation in cell hybrids between human neuroblastoma cells and thymidine auxotrophs of rat nerve‐like cells
Authors: Ishii Kazuhiro , Adachi Yoshifumi , Hatanaka Masakazu , Sakamoto Hiromi , Furuyama Jun‐Ichi 
Published: February 2005

AbstractCell hybrids (BIM) were produced between human neuroblastoma cells (IMR‐32) and thymidine auxotrophs (B3T) of rat nerve‐like cells (B103) in order to obtain cell lines undergoing stable neuronal differentiation. BIM cells exhibited the growth properties of partial transformation: 1) When the cell growth reached a plateau, BIM cells ceased to proliferate and expressed a differentiated phenotype. The shape of the cells changed from flat to round and they extended neurites. 2) When cultured in methylcellulose, BIM cells formed colonies, indicating that BIM cells have the ability of anchorage‐independent growth. By Southern blot analysis, BIM cells had both human and rat types of N‐myc genes. The human N‐myc genes were amplified, but the extent of the amplification was lower in BIM cells than that in the parental cell line IMR‐32. The rat N‐myc gene was detected at a similar level in BIM, B3T, B103, and rat fibroblastic cells 3Y1. Therefore, the decrease in amplification of human N‐myc genes may be involved in the properties of partial reverse‐transformation in BIM cells. When treated with various drugs such as db‐cAMP, forskolin, and cAMP with isobutyl‐methylxanthine, BIM cells expressed a nerve‐like phenotype. These findings indicate that cell hybridization yielded partial normalization of transformed nerve‐like cells.