bluuebunny/crossref_metadata_2025_split huggingface dataset:
Density‐related changes of potassium (86Rb) uptake by amphibian endothelial cells
Authors: Fava P. , Geck P. , Brändle K. , Bereiter‐Hahn J. 
Published: February 2005

AbstractPotassium influx has been investigated in XTH‐2 cells, a line derived from tadpole heart endothelia. In this line, the density at which the cultures become confluent is clearly separated from the density at which growth arrest takes place. Densityrelated changes in K+ influx were monitored by determining the uptake of 86Rb into well adhering cells kept in culture medium. The main observations were (1) 86Rb uptake is highest in single cells, and on confluency it reaches a low level, which is kept constant at higher cell density regardless of whether the cultures are stationary or still in logarithmic growth phase; (2) the relative amount of 86Rb taken up via the Na+−K+−2Cl− cotransport pathway and via the Na+/K+ pump changes from low cell density to confluent cultures; 86Rb uptake of single cells is nearly insensitive to ouabain, a maximum of ouabain sensitivity is reached around confluency, whereas piretanide‐sensitive 86Rb uptake is highest in single cells and seems to reach a minimum at the onset of confluency; (3) the variations in Na+/K+ pumping rate reflect neither differences in the amount of enzyme present nor changes in enzyme repartition between apical and basolateral plasma membranes; they seem to result from either “masking” or “unmasking” of the enzyme; (4) no alterations in K+ uptake occur that would be characteristic of the “stationary growth phase.” The only changes that seem to be related to arrest of proliferation are concerned with the Na+/K+ ‐ATPase, which achieves an extraordinary susceptibility to stimulation by monensin and exhibits an increase in PNPPase activity.
Laminin inhibits human keratinocyte migration
Authors: Woodley David T. , Bachmann Peter M. , O'Keefe Edward J. 
Published: February 2005

AbstractA quantitative migration assay for human keratinocytes was developed to assess the influence of extracellular matrix molecules on cell motility independently from their effect on cell proliferation. Fibronectin and collagen types I and IV markedly promoted keratinocyte migration, but albumin, type V collagen, and heparan sulfate proteoglycan had little effect. In contrast, laminin inhibited keratinocyte motility and dramatically reduced type IV collagen‐induced migration in a concentration‐dependent manner. Laminin was not toxic, since it had no apparent effect on morphology, growth, or ability of cells to be passaged. Laminin, a major component of the lamina lucida, may inhibit motility of keratinocytes in vivo. Absence of contact with laminin, which accompanies wounding, may facilitate motility and healing in the epidermis.
Effect of serum on the intracellular pH of BALB/c‐3T3 cells: Serum deprivation causes changes in sensitivity of cells to serum
Authors: Martinez R. , Gillies R. J. , Giuliano K. A. 
Published: February 2005

AbstractOne of the earliest responses of quiescent mammalian cells to the addition of serum is an increase in intracellular pH (pHin). This pHin change is generally believed to be due to an increased activity of Na+/H+ exchange. A number of investigators have observed steady‐state differences in pHin between cells in the presence and absence of serum. However, no one has examined differences in pHin regulation that may exist between cells chronically exposed to, or deprived of serum. In this study, we investigated the effects of serum deprivation to identify those components of pHin regulation that were associated with quiescence. To do this, we examined pHin in cells growing chronically in 10% serum as well as in cells that were either acutely (1.5–2 hr) or chronically (48 hr) deprived of serum. Intracellular pH was monitored using the fluorescence of intracellularly loaded pyranine dye. Our results indicate that the resting pHin values of chronically or acutely serum‐deprived cells were not significantly different from each other yet, in both cases, were lower than those observed in cells exposed to 10% serum. Furthermore, we observed significant increases in pHin of both acutely or chronically serum‐deprived cells in response to the addition of serum at various concentrations, in the presence of 24 mM bicarbonate. Chronically serum‐deprived cells had slightly smaller responses and were more sensitive to lower concentrations of serum than were acutely deprived cells. Therefore, our data suggest that long‐term serum deprivation affects the magnitude and sensitivity of pHin to serum stimulation and causes the loss of some form of pHin regulatory mechanism(s).
Enhancement of transformed cell growth in agar by serine protease inhibitors
Authors: Cook Jeffry R. , Chen Jan‐Kan 
Published: February 2005

AbstractWe investigated the effects of three serine protease inhibitors (leupeptin, soybean trypsin inhibitor, and aprotinin) on the serum‐free growth of two transformed cell lines in soft agar. Aprotinin markedly enhanced the growth of rat embryo fibroblasts that had been transformed by polyoma middle T antigen (PyMLV‐REF52), while having only a slight effect on the colonial growth of SV40 transformed Balb/c 3T3 cells (SV3T3‐Aga). Leupeptin and soybean trypsin inhibitor, on the other hand, significantly enhanced the growth of SV3T3‐Aga cells while having little effect on PyMLV‐REF52 growth. We observed no stimulatory effect of any of the protease inhibitors on serum‐free monolayer growth. Under conditions of excess aprotinin, PyMLV‐REF52 cells were found to be unresponsive to epidermal growth factor (EGF) at a concentration that would normally stimulate agar colony growth. However, aprotinin was not capable of supporting colony formation with transforming growth factor‐beta. These results indicate that aprotinin acts primarily as a protease inhibitor in spite of its structural homology to EGF and that EGF may promote the soft agar growth of these cell lines either by inhibiting proteolysis directly or by enhancing the synthesis of a serine protease inhibitor.
Growth‐dependent subcellular redistribution of protein kinase C in cultured porcine aortic endothelial cells
Authors: Uratsuji Yuko , Dicorleto Paul E. 
Published: February 2005

AbstractWe have previously observed major differences in the phosphorylation of membrane proteins in sparse, proliferating versus confluent, quiescent pig aortic endothelial cells (EC) (Kazlauskas and DiCorleto, 1987). In the present study we examined whether EC growth state can influence the activity of a specific phosphorylating enzyme, protein kinase C (PKC) in cytosolic and membrane fractions of pig aortic EC. Levels of PKC were measured using two methods: (1) Ca2+ and phospholipid‐dependent phosphorylation of exogenous histones using gamma‐labeled [32P]ATP, and (2) [3H]phorbol‐12, 13‐dibutyrate (PDBu) binding activity. The total amount of PKC activity in the quiescent versus proliferating cells was similar but the percentage of PKC activity in the membrane fraction correlated with the proliferative index of the cells: confluent, quiescent cultures exhibited a majority of PKC activity in the cytosolic fraction (67%), whereas sparse, proliferating cultures contained principally membrane‐bound PKC (70%). We also examined the role of PKC in the mitogenic response of pig aortic EC to fetal calf serum. Following serum stimulation of sparse, serum‐deprived pig aortic EC, PKC activity redistributed from the cytosolic to the membrane fraction in a rapid process that correlated with subsequent DNA synthesis. A potent activator of PKC, 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA), induced a minimal mitogenic response in pig aortic EC when added alone but acted synergistically with low concentrations of fetal calf serum to greatly stimulte DNA synthesis. Furthermore, pig aortic EC treated with TPA for 24 h to down‐regulate PKC exhibited only 25% of the serum‐stimulated mitogenic activity of control cultures. These results suggest a role for PKC activation and translocation in the proliferation of pig aortic EC.
Recombinant gibbon interleukin‐3 stimulates megakaryocyte colony growth in vitro from human peripheral blood progenitor cells
Authors: Mazur Eric M. , Cohen Janet L. , Bogart Lee , Allan Mufson R. , Gesner Thomas G. , Yang Yu‐Chung , Clark Steven C. 
Published: February 2005

AbstractGibbon interleukin‐3 (rlL‐3) has recently been cloned and found to have a high degree of homology with the human IL‐3 molecule. In this investigation, we evaluated the effects of gibbon rlL‐3 on normal human peripheral blood megakaryocyte progenitor cell growth in vitro. Gibbon rlL‐3 exhibited substantial megakaryocyte colony stimulatory activity (Meg‐CSA), supporting peak colony numbers at a concentration of 1 U/ml. Megakaryocyte colony growth induced by rlL‐3 reached 58% of the maximum achieved with the active, Meg‐CSA‐containing protein fraction of aplastic canine serum. Increasing gibbon rlL‐3 concentrations also stimulated a 4–5‐fold increase in megakaryocyte colony size and resulted in a decrease in geometric mean megakaryocyte ploidy. Ploidy values fell from 8.5N ± 1.4 (±SEM) at an rlL‐3 concentration of 0.1 U/ml to a minimum of 2.9N ± 0.3 at 10 U/ml. In the presence of rlL‐3 at 1.0 U/ml, megakaryocyte colony growth was linear with cell plating density and the regression line passed approximately through the origin. The effects of rlL‐3 on megakaryocyte colony growth were independent of the presence of T‐lymphocytes in the cultures. Cross‐species evaluation of murine and gibbon lL‐3 indicated that its bioactivity is species restricted. Murine lL‐3 did not support colony growth from human megakaryocyte progenitors and gibbon rlL‐3 showed no activity in stimulating acetylcholinesterase production by murine bone marrow cells. Gibbon rlL‐3 is a potent stimulator of the early events of human megakaryocyte progenitor cell development promoting predominantly mitosis and early megakaryocytic differentiation.
Cells with marrow and spleen repopulating ability and forming spleen colonies on day 16, 12, and 8 are sequentially ordered on the basis of increasing rhodamine 123 retention
Authors: Ploemacher R. E. , Brons N. H. C. 
Published: February 2005

AbstractMouse bone marrow cells (BMC) were subjected to countercurrent centrifugal elutriation and subsequently separated on the basis of light scatter and fluorescence intensity after being labeled with the supravital dye Rhodamine 123 (Rh‐123). The sorted cells were then assayed for their in vivo spleen colony‐forming ability (day −8, −12, and −16 CFU‐S) and their ability to repopulate the bone marrow or spleen over a 13‐day period with CFU‐S‐12, CFU‐GM, or nucleated cells. Cells with marrow repopulating ability (MRA), as measured by the ability of the sorted cells to repopulate the marrow with secondary CFU‐S‐12 or CFU‐GM, had low affinity for Rh‐123. These cells showed minimal spleen colony‐forming ability, and the ratio of MRA to CFU‐S‐12 in this preparation was 309. Cells with spleen repopulating ability (SRA), CFU‐S‐16, CFU‐S‐12, and CFU‐S‐8 retained increasing amounts of Rh‐123, respectively, and CFU‐S‐8 were almost exclusively found among cells with high Rh‐123 affinity. These cells also included about half of all day‐12 CFU‐S, and the ratio of MRA to day‐12 CFU‐S was 0. The results show that MRA cells, SRA cells, CFU‐S‐16, CFU‐S‐12, and CFU‐S‐8 can be sequentially ordered on the basis of increasing mitochondrial activity. The data also demonstrate for the first time, and without the application of negative selection by the use of cytostatic agents, that MRA cells are a separate class of primitive hemopoietic stem cells that fully meet the criteria of pre‐CFU‐S.
Extent and properties of nonbulk “bound” water in crystalline lens cells
Authors: Cameron I. L. , Contreras E. , Fullerton G. D. , Kellermayer M. , Ludány A. , Miseta A. 
Published: February 2005

AbstractCrystalline lenses provided good material to study and measure the properties of cellular water. Different methods were used to establish the extent and properties of nonbulk water in mammalian lenses. These methods include: NMR titration analysis, a test of the osmotic properties, a test of dye exclusion In lenses with intact cell membranes and in lenses with disrupted cell membranes, and the water‐holding capacity of lenses subjected to 40,000 × g for 1 hour with intact cell membranes and in lenses with disrupted cell membranes. The data from these methods, as well as other data from the literature, lead to the conclusion that most, if not all, of the water in lens cells (up to 2.2 g water/g dry mass) has motional and osmotic properties that distinguish it from bulk water. These findings call into question the common and convenient assumption that all but a small proportion of cellular water is like that in dilute solution.
Characterization of a high‐molecular‐weight protein immunoprecipitated by platelet‐derived growth factor antisera
Authors: Graves Dana T. , Antoniades Harry N. 
Published: February 2005

AbstractWe previously demonstrated that a high‐molecular‐weight glycoprotein could be immunoprecipitated from metabolically labeled U‐2 OS cells with platelet‐derived growth factor (PDGF) antiserum and that it appears to be derived from a different precursor than is the 30 kD PDGF‐like mitogen produced by these cells. These findings were unexpected, since the molecular weight of this glycoprotein is too large to be encoded by the PDGF structural genes. From experiments with metabolically labeled U‐2 OS human osteosarcoma, fibroblasts, and NRK cells, we report here that a 185 kD protein immunoprecipitated with PDGF antiserum has the following characteristics. (1) It is a PDGF binding protein that is unrelated to α2‐macroglobulin. (2) It is phosphorylated in response to PDGF stimulation. (3) It is immunoprecipitated by phosphoryrosine antibodies. (4) It is not a substrate of epidermal growth factor‐induced tyrosine kinase activity. These studies indicate that high‐molecular‐weight proteins immunoprecipitated by antiserum to PDGF represent a complex between PDGF and a binding protein capable of being phosphorylated by a PDGF‐induced tyrosine kinase. These characteristics are identical to those of the PDGF receptor.
DNA replication fork progression rate and temporal organization of S phase in normal epidermis and in basal cell carcinoma
Authors: Galand Paul , Heenen Michel 
Published: February 2005

AbstractThe double‐pulse labeling technique for DNA fiber autoradiography was applied to epidermal cells from normal human skin and from human basal cell carcinoma (BCC). We aimed to measure the size and replication rate of the replication unit (RU) for both types of cell and to account, from these results, for our previous observation of a near doubling of S‐phase duration in BCC, compared with normal skin. The mean RU size was 76 ± 4 m̈m in BCC, not significantly different from the 68 ± 6 μm value found in normal skin, so the mean of those two values (i.e., 72 μm), was used in further calculations. The rate of replication fork progression was 0.59 ± 0.005 μmlm/min in the normal epidermis and 0.33 ± 0.03 μm/min in BCC, corresponding to a replication time of the average RU equal to 61 min and 109 min, respectively. Thus, with an unchanged RU size in BCC, the observed 1.8‐fold decrease in the rate of fork progression in the tumor can account entirely for our previous observation of a 1.8‐fold increase in S‐phase duration in this tumor, without requiring the assumption of any change in the temporal organization of DNA synthesis in the malignant cells. Considering S phase as an ordered process in which a major part, if not all, of the genome replicates at genetically determined times, we suggest that the clusters of replication units are, in turn, organized into temporally defined “sets.” These sets are composed of all the clusters (whatever their chromosomal location) that are programmed to initiate replication during the same fraction of the S period. This hypothesis implies that DNA synthesis in a given set is triggered by some event coupled to progression of replication in the immediately preceding set. Based on a S‐phase duration of 10.2 hours in normal skin and of 19.2 hours in BCC (our previous data), and assuming perfect synchrony and homogeneity of the clusters within each set and of each cluster's constitutive RUs, the minimum number of sequentially replicating sets, in both instances, can be estimated as roughly equal to 10.
Dual effect of dihydropyridines on 45Ca uptake induced by the K+ —channel blocker 4‐aminopyridin on mast cells
Authors: Eleno N. , Botana L. , Espinosa J. 
Published: February 2005

AbstractThe 1,4‐dihydropyridines (DHP) are calcium antagonists and represent a new class of drugs which act by a selective inhibition of Ca++ influx through voltage‐operated calcium channels. We report the effect of nifedipine (Bay A 1040), nisoldipine (Bay K 5552) ad nitrendipine (Bay E 5009) on the histamine release and on the 45Ca uptake promoted by 4‐aminopyridin in mast cells. These cells treated with DHP (10−12−10−3 M) activated the secretory response in a dose‐dependent manner in the range of concentrations 10−6−10−3 M did not significantly inhibit the secretion. 4‐Aminopridin, a known K+‐channel blocker, induced 45Ca uptake. Pretreatment of mast cells with DHP prior to 4‐aminopyridin stimulation inhibited or stimulated 45Ca uptake depending on concentration; thus, concentrations of DHP below 10−12 of nitrendipine and 10−9 for nisoldipine and nifedipine were inhibitory, while higher doses potentiated 45Ca uptake. These results demonstrate a diversity of pharmacological effects of DHP on mediator secretion and 45Ca uptake in mast cells and throw into question their only properties as Ca++ antagonists.
Effect of endothelial‐cell‐conditioned medium on proteoglycan synthesis in cultured smooth muscle cells from pig aorta
Authors: Berrou Eliane , Breton Monique , Deudon Elisabeth , Picard Jacques 
Published: February 2005

AbstractThe effect of porcine endothelial‐cell‐conditioned medium on proteoglycan synthesis by pig aorta smooth muscle cells was studied under serum‐free conditions. Maximal stimulation of [35S]‐sulfate incorporation (50%) into medium‐secreted and cell layer proteoglycans was observed after 20 min and 4 h incubation, respectively. This stimulation can be explained neither by increased secretion nor by oversulfation of medium‐secreted [35S]‐labeled proteoglycans. Those [35S]‐proteoglycans secreted (for 24 h) in the presence of endothelial cell‐conditioned medium were characterized by a higher hydrodynamic size than those secreted in the presence of control medium, without modification of glycosaminoglycan chain length. Agreement between the stimulation of incorporation of [35S]‐sulfate into glycanic chains (50.1%) and [14C]‐serine residues associated with glycosaminoglycans (49.9%) involved an increase in the number of glycanic chains linked to protein cores. The lesser stimulation of [14C]‐serine incorporation into secreted proteins (18%) suggested that stimulation of glycosaminoglycan synthesis was not the direct consequence of enhanced protein synthesis. Proteoglycan synthesis was studied in the presence of para‐nitrophenyl‐b̃‐D‐xyloside. Fractionation of medium‐secreted [35S]‐proteoglycans and xyloside‐initiated glycosaminoglycans revealed that stimulation of [35S]‐glycosaminoglycan synthesis by endothelial‐cell‐conditioned medium required a protein core acceptor for glycanic chain initiation. Our results suggest that the factor(s) secreted by endothelial cells are able to modify smooth muscle cell proteoglycan synthesis by stimulating the first step of protein core glycosylation. This stimulation was accompanied by an increase in proteoglycan hydrodynamic size.
Ca‐dependent slow action potentials in human skeletal muscle
Authors: Siri L. Nicola , Dubrovsky A. L. , Uchitel O. D. 
Published: February 2005

AbstractSlow Ca‐action potentials (CaAP) were studied in normal human skeletal muscle fibers obtained during surgery (fibers with both ends cut). Control studies also were carried out with intact as well as cut rat skeletal muscle fibers. Experiments were performed in hypertonic Cl‐free saline with 10 or 84 mM Ca and K‐channel blockers; muscles were preincubated in a saline containing Cs and tetraethylammonium. A current‐clamp technique with two intracellular microelectrodes was used. In human muscle, 14.5% of the fibers showed fully developed CaAPs, 21% displayed nonregenerative Ca responses, and 64.5% showed only passive responses; CaAPs were never observed in 10 mM Ca. In rat muscle, nearly 90% of the fibers showed CaAPs, which were not affected by the cut‐end condition. Human and rat muscle fibers had similar membrane potential and conductance in the resting state. In human muscle (22–32± C, 84 mM Ca), the threshold and peak potential during a CaAP were + 26 ± 6 mV and + 70 ± 3 mV, respectively, and the duration measured at threshold level was 1.7 ± 0.5 sec. In rat muscle, the duration was four times longer During a CaAP, membrane conductance was assumed to be a leak conductance in parallel with a Ca and a K conductance. In human muscle (22–32° C, 84 mM Ca, 40 μS, fiber diameter), values were 0.4 ± 0.1 μS, 1.1 ± 0.7 μS, and 0.9 ± 0.4 μS, respectively. Rat muscle (22–24° C, 84 mM Ca) showed leak and K conductances similar to those found in human fibers. Ca‐conductance in rat muscle was double the values obtained in human muscle fibers.
Transferrin endocytosis and iron uptake in developing myogenic cells in culture: Effects of microtubular and metabolic inhibitors, sulphydryl reagents and lysosomotrophic agents
Authors: Sorokin L. M. , Morgan E. H. , Yeoh G. C. T. 
Published: February 2005

AbstractThe experiments described in this study were designed to investigate receptor‐mediated endocytosis of transferrin and its role in iron uptake by cultured chick presumptive myoblasts (dividing and non‐dividing) and myotubes. The effects of a variety of inhibitors on the internalization of transferrin and iron were investigated and three main effects were found: (i) sulphydryl reagents and microtubular inhibitors reduced the rate of transferrin and iron internalization to similar degrees, (ii) metabolic inhibitors reduced the rate of iron uptake more than that of transferrin endocytosis, and (iii) lysosomotrophic agents almost completely abolished iron accumulation by the cells without any effect on the rate of transferrin internalization. The results suggest that metabolic energy is required not only for the endocytosis of transferrin but also for subsequent steps in the iron uptake process, and that iron release from transferrin occurs in acidified endosomes. Overall, these experiments show that all or virtually all of the iron taken up by developing muscle cells from transferrin occurs as a consequence of receptor‐mediated endocytosis of the protein.
Stable variant of LM fibroblast defective in fluid‐phase but competent in receptor‐mediated endocytosis
Authors: Ryser Hugues J.‐P. , Li Wande , Mandel Richard , Shen Wei‐Chiang 
Published: February 2005

AbstractThe F‐40 cell line, a stable variant of LM fibroblasts selected for its resistance to polyethylene glycol (PEG)‐induced fusion (Roos and Davidson: Somatic Cell and Molecular Genetics 6:381‐‐391, 1980), has a decreased capacity to internalize fluid‐phase markers and nonspecifically surface‐bound macromolecules. It is not defective in exocytosis since, after a short sucrose pulse, it releases the same fraction of ingested sucrose into the medium as does the parental line. F40 cells have a normal capacity to carry out receptor‐mediated endocytosis, as tested with 125I‐alpha‐2 macroglobulin (alpha‐2 MG) and 125I‐transferrin (Tf), and to recycle Tf receptor to the cell surface. These data demonstrate that receptor‐mediated and non‐receptor mediated endocytosis are distinct processes that can be altered independently. Of the many membrane fusions occurring in the course of endocytosis, the only one that appears associated with the defect in cell fusion characteristic of F40 cells is the formation of primary endocytotic vesicles engaged in non‐receptor‐mediated internalizations.