bluuebunny/crossref_metadata_2025_split huggingface dataset:
In vitro expression of a 38,000 dalton heparin‐binding glycoprotein by morphologically differentiated smooth muscle cells
Authors: Millis Albert J. T. , Hoyle Marian , Kent Lorraine 
Published: February 2005

AbstractIn vitro, high density monolayer cultures of vascular smooth muscle cells can be induced to form multicellular nodules. The nodular cells appear to be morphologically differentiated smooth muscle cells. Sodium dodecyl sulfate (NaDodSO4)‐polyacrylamide gel electrophoresis was used to compare the proteins synthesized and secreted by monolayer and nodular cultures of smooth muscle cells. Although most proteins appeared to be similar, the nodular cultures contained a unique heparin binding protein of Mr = 38,000 (38kD protein) (Millis, A.J.T., Hoyle, M., Reich, E., and Mann, D.M., 1985, J. Biol. Chem., 260: 3754–3761). The 38kD protein was glycosylated and its apparent molecular weight was shifted to Mr = 32,500 after synthesis in the presence of tunicamycin or digestion with endoglycosidase F. The production of 38kD protein by nodular cell cultures did not appear to result from the degradation of a high molecular weight precursor in nodular conditioned medium. Further, it was not detected in monolayer cell conditioned medium that had been incubated with nodular cells. Finally, its synthesis was not induced in monolayer cell cultures that had been labeled in nodular cell conditioned medium. The 38kD protein appears to be uniquely associated with nodular cultures of smooth muscle cells.
Mechanisms of cytoskeletal regulation: Modulation of aortic endothelial cell protein band 4.1 by the extracellular matrix
Authors: Leto Thomas L. , Pratt Bruce M. , Madri Joseph A. 
Published: February 2005

AbstractThe bovine aortic endothelial cell (BAEC) cytoskeleton is a complex structure modulated by many stimuli including release from contact inhibition and various components of the extracellular matrix (ECM). Transduction of information from the ECM to the cell nucleus proceeds via several complex pathways including the cytoskeleton. We have demonstrated the presence of an immunoreactive isoform of the human erythrocyte cytoskeletal protein band 4.1 (4.1) in BAEC. BAEC 4.1 is similar in molecular weight to the erythroid protein by immunoblot analyses and produces a similar pattern of cysteine specific cleavage products consistent with a cluster of cysteine residues previously described in the erythroid molecule. We have also examined the effects of defined ECM proteins on the distributions of cultured BAEC 4.1 and actin filaments (AF) at confluency and following release from contact inhibition. The distribution of 4.1 in BAEC on a plasma fibronectin substrate is complex, having partial codistribution with cytoplasmic AF and a unique perinuclear staining. In contrast, on a collagen type I/III substrate, 4.1 is localized, in part, to peripheral areas of cell‐cell contact distinct from the dense peripheral band staining of AF. During migration on this substrate, 4.1 had a filamentous distribution having partial codistribution with AF. Indirect immunofluorescence staining of cross‐sections of bovine calf aortae revealed a cortical staining pattern in the aortic endothelial cells with staining noted on the luminal and basolateral aspects of the cells. These data suggest that, in endothelial cells, protein 4.1 is a cortical membrane protein which may function to link actin filaments to other skeletal proteins such as spectrin. These findings also suggest an active role for protein 4.1 in cytoskeletal reorganization events which can occur in response to external stimuli, such as the extracellular matrix or contact with other cells.
Alternative interpretation for the osmotic response of human erythrocytes
Authors: Massaldi H. A. , Richieri G. V. , Mel H. C. 
Published: February 2005

AbstractIn a recent publication, Heubusch et al. (J Cell. Physiol, 122:266–272, 1985) reported changes of erythrocyte volume measured by the Coulter counter technique over a wide range of osmolalities (160 to 3000 mOsm). Their results showed a partially hindered, nonlinear response, in contrast to classical observations made over more restricted osmolality ranges, using other methods. The authors suggested the underlying cause of this behavior to be a mechanical resistance of the membrane cytoskeleton. In this paper, we wish to offer a different interpretation of their results on erythrocyte osmotic behavior, based on similar experiments carried out in our laboratory, and supported by previous analyses from the literature. In particular, it is shown that the shape‐factor correction to the electronic sizing measurement can correctly account for the observed deviations from linearity in the hypotonic range. In contrast, increased chemical nonideality and eventual hemolysis are the likely factors responsible for the behavior in the hypertonic range.
Electrophysiological responses of osteoclasts to hormones
Authors: Ferrier Jack , Ward Angela , Kanehisa Junya , Heersche Johan N. M. 
Published: February 2005

AbstractElectrophysiological measurements were carried out on osteoclasts in vitro. Such isolated osteoclasts are able to resorb bone in vitro and contract in response to calcitonin (CT). Our measurements show that individual osteoclasts respond to CT with a significant transient hyperpolarization of membrane potential. Application of parathyroid hormone (PTH) and dibutyryl cAMP produced a transient hyperpolarization in some osteoclasts. Measurements on an osteoblastlike line (ROS 17/2.8) showed a sustained hyperpolarizing response to CT, which is similar to but smaller than the hyperpolarizing response to PTH and dibutyryl cAMP in this and some other osteoblastlike lines. In contrast to osteoblastlike cells, the osteoclasts have no long term membrane potential response to CT, to PTH, or to dibutyryl cAMP. These results show that there are distinct differences between osteoclasts and osteoblasts in their ion transport responses to hormones.
Regulation of inositol phosphate levels by prostaglandins in cultured endometrial cells
Authors: Orlicky D. J. , Silio M. , Williams C. , Gordon J. , Gerschenson L. E. 
Published: February 2005

AbstractStimulation of cultured rabbit endometrial cells by one of the rabbit endometrial cell culture proliferation factors, prostaglandin F2α (PGF2α), resulted in a very rapid increase in the intracellular levels of [3H]‐inositol triphosphate (IP3), [3H]‐inositol biphosphate (IP2), and [3H]‐inositol monophosphate (IP1) in cells prelabeled with [3H]‐inositol. These increases in inositol phosphate levels were detected in periods of stimulation as short as 30 seconds, reached a maximum by 1 1/2−2 min and declined to control levels by 6–10 min. The stimulation was dose‐dependent with maximal increases observed near 10−6 M PGF2α. The cholinergic agent, carbachol, also led to time and dose‐independent increases in IP3. Lithium, cadmium, silver, copper, and zinc ions had no effect either on the breakdown of IP3 or on the accumulation of IP1. In contrast, vanadate at 10−6 or 10−5 M did lead to a decrease in the breakdown of IP1 and a concomitant increase in IP1, IP2, and IP3. PGF2α was found previously to induce an increase in rabbit endometrial cell DNA synthesis which was inhibited by concomitant or prior addition of prostaglandin E1 (PGE1). PGE1, in a dose‐dependent manner, was found to inhibit the observed IP3 increase by PGF2α at 1 1/2 min of stimulation. PGF2α treated and control cultures did not differ in cAMP or cGMP levels, cellular 45Ca uptake, nor cellular 22Na uptake. We propose that IP3 may be one of the intracellular messenger(s) synthesized following the treatment of rabbit endometrial cell cultures with the proliferation agent PGF2α and that it may play a crucial role with cAMP in growth regulation.
Energy‐dependent volume regulation in primary cultured cerebral astrocytes
Authors: Olson James E. , Sankar Raman , Holtzman David , James Adrian , Fleischhacker Deborah 
Published: February 2005

AbstractCell volume regulation and energy metabolism were studied in primary cultured cerebral astrocytes during exposure to media of altered osmolarity. Cells suspended in medium containing 1/2 the normal concentration of NaCl (hypoosmotic) swell immediately to a volume 40–50% larger than cells suspended in isoosmotic medium. The cell volume in hypoosmotic medium then decreases over 30 min to a volume approximately 25% larger than cells in isoosmotic medium. In hyperosmotic medium (containing twice the normal concentration of NaCl), astrocytes shrink by 29%. Little volume change occurs following this initial shrinkage. Cells resuspended in isoosmotic medium after a 30 min incubation in hypoosmotic medium shrink immediately to a volume 10% less than the volume of cells incubated continuously in isoosmotic medium. Thus, the regulatory volume decrease (RVD) in hypoosmotic medium involves a net reduction of intracellular osmoles. The RVD is partially blocked by inhibitors of mitochondrial electron transport but is unaffected by an inhibitor of glycolysis or by an uncoupler of oxidative phosphorylation. Inhibition of RVD by these metabolic agents is correlated with decreased cellular ATP levels. Ouabain, added immediately after hypoosmotic induced swelling, completely inhibits RVD, but does not alter cell volume if added after RVD has taken place. Ouabain also inhibits cell respiration 27% more in hypoosmotic medium than in isoosmotic medium indicating that the (Na, K)‐ATPase‐coupled ion pump is more active in the hypoosmotic medium. These data suggest that the cell volume response of astrocytes in hypoosmotic medium involves the net movement of osmoles by a mechanism dependent on cellular energy and tightly coupled to the (Na, K)‐ATPase ion pump. This process may be important in the energy‐dependent osmoregulation in the brain, a critical role attributed to the astrocyte in vivo.
Development of a neural phenotype in differentiating ganglion cell‐derived human neuroblastoma cells
Authors: Scott Ian G. , Åkerman Karl E. O. , Heikkilä Jari E. , Kaila Kai , Andersson Leif C. 
Published: February 2005

AbstractHuman neuroblastoma cells (clone SHSY‐5Y) induced to differentiate by 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA) are shown to possess properties characteristic of mature ganglion cells. Elevation of the external K+ concentration, exposure to Ca2+ ionophore A23187, and acetylcholine all stimulate the release of preloaded 3H‐noradrenaline in the presence but not in the absence of added Ca2+. Acetylcholine causes a fall in the 86Rb+ or 14C‐TPMP equilibrium potential across the plasma membrane and stimulates 86Rb+ efflux. These responses are prevented by atropine. Acetylcholine and muscarine but not nicotine stimulate an increase in 45Ca2+ influx, an effect blocked by atropine. None of these responses have been observed in nondifferentiating cells. Muscarinic receptors, however, as measured by the binding of tritiated quinuclidinyl benzilate (3H‐QNB), were present to a similar extent in control and differentiated cells. Both cell types also exhibit an accelerated release of Ca2+ in response to acetylcholine, but the control cells were at least 1 order of magnitude more sensitive to the agonist.
Human skeletal growth factor stimulates collagen synthesis and inhibits proliferation in a clonal osteoblast cell line (MC3T3‐E1)
Authors: Linkhart Susan , Mohan Subburaman , Linkhart Thomas A. , Kumegawa Masayoshi , Baylink David J. 
Published: February 2005

AbstractHuman skeletal growth factor (hSGF), an 11‐kD polypeptide purified from human bone, has been proposed to be a local regulator of bone formation. To investigate the underlying cellular mechanisms in an in vitro model system, we examined the effects of hSGF on proliferation and collagen synthesis in cells of the clonal osteoblast cell line MC3T3‐E1. This line was isolated from newborn mouse calvarial cells and retains many characteristics of mature osteoblasts (Sudo, H., et al., (1984) J. Cell Biol. 96:191). A 14‐hr treatment with hSGF increased noncollagenous protein synthesis to 215% of unstimulated controls and increased collagen synthesis to 630% of controls as determined by [3H]proline incorporation and high‐pressure liquid chromatographic separation of [3H]proline and [3H]hydroxyproline in acid hydrolysates of trichloroacetic acid‐insoluble protein. HSGF did not increase cell number over a 48‐hr period and caused a reversible inhibition of DNA synthesis. Half‐maximal hSGF concentration for stimulation of [3H]proline incorporation and inhibition of [3H]thymidine incorporation was 100 ng/ml. HSGF also inhibited DNA synthesis in cells stimulated by serum. In contrast, hSGF stimulated both collagen synthesis and DNA synthesis in primary cultures of chick embryo bone cells, which may be developmentally less mature than MC3T3‐E1 cells. The results suggest that hSGF directly stimulated mature osteoblast matrix synthetic activity and that hSGF has differential effects on proliferation of osteoblast progenitor cells and mature osteoblasts.
Reversible inactivation of the Ah receptor associated with changes in intracellular ATP levels
Authors: Gudas Jean M. , Hankinson Oliver 
Published: February 2005

AbstractWe have previously demonstrated that incubation of Hepa‐1 cells in the presence of cytochalasin B (CB) results in a time‐ and temperature‐dependent loss of 2,3,7,8‐tetrachlorodibenzo‐p‐dioxin (TCDD) binding activity (Gudas et al., 1986). We show here that this loss of binding activity is probably attributable to a CB induced inhibition of glucose transport, as incubation of cells in the presence of glycolytic inhibitors or in glucose free medium caused a similar effect. All conditions leading to loss of binding also caused a marked reduction in cellular ATP concentration, suggesting that ATP (or perhaps another energy‐dependent molecule) is required for maintaining the receptor in the active state. Inactivation of the Ah receptor occurred in the cytosol but not when it had translocated to the nucleus. Reactivation of receptor binding activity occurred readily in vivo and did not require de novo protein synthesis. However, attempts to restore receptor binding activity in vitro were not successful. To our knowledge this is the first reported evidence indicating that the TCDD binding Ah receptor can exist in both an inactive and an active form, with the amount present in the active ligand binding form being coupled to the energy state of the cells.
Mc3T3‐G2/PA6 preadipoctyes support in vitro proliferation of hemopoietic stem cells through a mechanism different from that of interleukin 3
Authors: Kodama Hiro‐Aki , Hagiwara Hajime , Sudo Hiroko , Amagai Yuji , Yokota Takashi , Arai Naoko , Kitamura Yukihiko 
Published: February 2005

AbstractBoth MC3T3‐G2/PA6 preadipocytes and interleukin 3 (IL 3) can support in vitro proliferation of mouse hemopoietic stem cells (CFU‐S). We examined whether MC3T3‐G2/PA6 cells produce IL 3 and whether a common mechanism might underlie the action of both of these agents. We used cultured mast cells, DA‐1 cells, and FDC‐P2 cells as the targets of IL 3 and conditioned medium (CM) of WEHI‐3 cells as a source of IL 3. MC3T3‐C2/PA6 CM did not support the growth of the above cells. IL 3 mRNA was not detected in the preadipocytes. Since CM obtained from the cocultures of bone marrow cells and MC3T3‐G2/ PA6 cells did not have a significant effect on the growth of the IL 3‐dependent cells, none of the bone marrow cells seem to produce IL 3 under the influence of the preadipocytes. When the factor‐dependent cells were cocultured with MC3T3‐G2/PA6 cells, the former did not survive, whereas mast cells and DA‐1 cells intimately associated with the preadipocytes. Even when bone marrow cells, mast cells, and MC3T3‐G2/PA6 cells were cocultured, the number of CFU‐S increased, but not that of mast cells. These results seem to exclude the possibility of the action of IL 3 in the microenvironment provided by MC3T3‐G2/PA6 preadipocytes.
Modulation of WI‐38 cell proliferation by elevated levels of CaCl2
Authors: Praeger Frank C. , Cristofalo Vincent J. 
Published: February 2005

AbstractElevating the level of extracellular calcium (CaEx2+) increases the saturation density achieved by the normal human diploid cell line, WI‐38, but does not change the growth rate. Day 7 cell yields remain unchanged when [CaEx2+] is between 0.5 mM and 3.0 mM, decrease when [CaEx2+] < 0.5 mM, and increase when [CaEx2+] > 3.0 mM. Combining hydrocortisone with additional CaCl2 results in an additive effect on the saturation density relative to that obtained with each treatment separately. The stimulatory effect of elevated [CaCl2] is independent of serum concentration but is lost when WI‐38 cells are grown in conditioned medium. Stimulation is recovered when conditioned medium is diluted with serum‐free medium. In the case of young cultures grown in conditioned medium, stimulation can also be recovered when higher than usual levels of additional CaCl2 are used (2–3 mM). A glutamine supplementation to the conditioned medium potentiates cell response to elevated [CaCl2]. These results indicate that the loss of an enhanced saturation density when cells are grown in conditioned medium is not due to serum depletion but is more likely the effect of metabolites and/or nutrient depletion. When older or less vigorously growing cultures are grown in conditioned medium, additions of up to 3 mM CaCl2 only lead to inhibition, suggesting an age‐related change in proliferative regulation. Elevated levels of CaEx2+ also enhance the proliferative response of quiescent monolayers to serum stimulation. This finding, along with the increase in saturation density of Ca2+‐treated cultures, suggests that an elevated level of CaEx2+ affects cell entry into and exit from quiescence brought on by density‐dependent inhibition.
Na+‐dependent amino acid transport is a major factor determining the rate of (Na+, K+)‐ATPase mediated cation transport in intact HeLa cells
Authors: Zibirre Reinhard , Poronnik Philip , Koch Gebhard 
Published: February 2005

AbstractLittle is known concerning the effects of Na+‐coupled solute transport on (Na+, K+)‐ATPase mediated cation pumping in the intact cell. We investigated the effect of amino acid transport and growth factor adition on the short term regulation of (Na+, K+)‐ATPase cation transport in HeLa cells. The level of pump activity in the presence of amino acids or growth factors was compared to the level measured in phosphate buffered saline. These rates were further related to the maximal pump capacity, operationally defined as ouabain inhibitable 86Rb+ influx in the presence of 15 μM monensin. Of the growth factors tested, only insulin was found to moderately (22%) increase (Na+, K+)‐ATPase cation transport. The major determinant of pump activity was found to be the transport of amino acids. Minimal essential medium (MEM) amino acids increased ouabain inhibitable 86Rb+ influx to a level close to that obtained with monensin, indicating that the(Na+, K+ )‐ATPase is operating near maximal capacity during amino acid transport. This situation may apply to tissue culture conditions and consequently measurements of (Na+, K+)‐ATPase activity in buffer solutions alone may yield little information about cation pumping under culture conditions. This finding applies especially to cells having high rates of amino acid transport. Furthermore, rates of amino acid transport may be directly or indirectly involved in the long‐term regulation of the number of (Na+, K+)‐ATPase molecules in the plasma membrane.
Control of heme synthesis during Friend cell differentiation: Role of iron and transferrin
Authors: Laskey Jennifer D. , Ponka Prem , Schulman Herbert M. 
Published: February 2005

AbstractIn many types of cells the synthesis of δ‐aminolevulinic acid (ALA) limits the rate of heme formation. However, results from our laboratory with reticulocytes suggest that the rate of iron uptake from transferrin (Tf), rather than ALA synthase activity, limits the rate of heme synthesis in erythroid cells. To determine whether changes occur in iron metabolism and the control of heme synthesis durihg. Erythroid cell development Friend erythroleukemia cells induced to erythroid differentiation by dimethylsulfoxide (DMSO) were studied. While added ALA stimulated heme synthesis in uninduced Friend cells (suggesting ALA synthase is limiting) it did not do so in induced cells. Therefore the possibility was investigated that, in induced cells, iron uptake from Tf limits and controls heme synthesis. Several aspects of iron metabolism were investigated using the synthetic iron chelator salicylaldehyde isonicotinoyl hydrazone (SIH). Both induced and uninduced Friend cells take up and utilize Fe for heme synthesis directly from Fe‐SIH without the involvement of transferrin and transferrin receptors and to a much greater extent than from saturating levels of Fe‐Tf (20 μM). Furthermore, in induced Friend cells 100 μM Fe‐SIH stimulated 2‐l4C‐glycine incorporation into heme up to 3.6‐fold as compared to the incorporation observed with saturating concentrations of Fe‐Tf. In contrast, Fe‐SIH, even when added in high concentrations, did not stimulate heme synthesis in uninduced Friend cells but was able to do so as early as 24 to 48 h following induction. In addition, contrary to previous results with rabbit reticulocytes, Fe‐SIH also stimulated globin synthesis in induced Friend cells above the level seen with saturating concentrations of transferrin. These results indicate that some step(s) in the pathway of iron from extracellular Tf to protoporphyrin, rather than the activity of ALA synthase, limits and controls the overall rate of heme and possibly hemoglobin synthesis in differentiating Friend erythroleukemia cells.
Elevation of intracellular glutathione content associated with mitogenic stimulation of quiescent fibroblasts
Authors: Shaw Jill P. , Chou Iih‐Nan 
Published: February 2005

AbstractThe relationship between total glutathione (GSH) content and cell growth was examined in 3T3 fibroblasts. The intracellular GSH level of actively growing cultures gradually decreases as these cells become quiescent by either serum deprivation or high cell density. Upon mitogenic stimulation of sparse, quiescent (G0/G1) cultures with serum, there is a rapid 2.3‐fold elevation in intracellular GSH levels which, is maximal by 1 h and returns to baseline by 2 h. This is followed by a more gradual increase in GSH content as cells enter the S phase. In addition, the elevation in GSH content is required for maximum induction of DNA synthesis. Treatments that prevent the early increase in intracellular GSH levels do not affect protein synthesis but result in a reversible dose‐dependent decrease in the percent of cells capable of entering S phase. These results indicate that GSH may be important in the regulation of cellular proliferation.