bluuebunny/crossref_metadata_2025_split huggingface dataset:
Modulation of cell attachment to culture support by pH, fibronectin, hemin, and cobalt protoporphyrin
Authors: Imbenotte José , Verger Claude , Sassa Shigeru 
Published: February 2005

AbstractWhen chick embryo fibroblasts were seeded in the presence of minimum essential medium supplemented with 1% (v/v) horse serum, the rate of cell attachment, after 1 hr incubation, was less than 5% at pH 6.5 and about 50% and 80% at pH 7.5 and pH 8.3, respectively. If, however, cultures were pretreated with fibronectin, a substance that promotes cell adhesion, a high rate of cell attachment was also observed at pH 6.5. Two other compounds of totally different chemical nature, cobalt‐protoporphyrin (CoPP) and hemin, also enhanced cell attachment at pH 6.5. CoPP was shown to increase the synthesis of proteins, but it did not affect the intracellular heme content of cells incubated at pH 6.5. The possibility that CoPP, and presumably also hemin, induce cell attachment by promoting the synthesis of a fibronectin like protein is discussed.
Effects of interaction between estradiol‐17β and progesterone on the proliferation of cloned breast tumor cells (MCF‐7 and T47D)
Authors: Schatz Robert W. , Soto Ana M. , Sonnenschein Carlos 
Published: February 2005

AbstractWe have previously reported that the proliferation of cloned MCF‐7 and T47D human mammary tumor cells can be inhibited by increasing concentrations of charcoal‐dextran stripped female human serum (CDFHS). The maximal proliferation rate was restored by the addition of 3 × 10−11 M estradiol‐17β to the culture media. These observations suggest that the proliferation of T47D and MCF‐7 cells is regulated by a blood‐borne inhibitor whose effects are neutralized by estrogens. In the present report we explore the possibility that progesterone alters the estrogenic response. MCF‐7 cells were grown in DME containing 2–40% CDFHS. Progesterone, at 3 × 10−7 M to 3 × 10−12 M, had no effect on the yield of MCF‐7 or T47D cells that were cultured in the presence or absence of estradiol‐17β.
Platelet‐derived growth factor stimulates rapid polyphosphoinositide breakdown in fetal human fibroblasts
Authors: Chu Shu‐Heh W. , Hoban Carolyn J. , Owen Albert J. , Geyer Robert P. 
Published: February 2005

AbstractThe addition of human platelet‐derived growth factor (PDGF) to confluent, quiescent cultures of human diploid fibroblasts induced the rapid breakdown of cellular polyphosphoinositides. The levels of 32P‐labeled phosphatidylinositol 4,5‐bisphosphate (PIP2), phosphatidylinositol 4‐phosphate (PIP), and phosphatidylinositol (PI) decreased by 30 to 40% within 1 min after exposure of the cells to PDGF. The levels of PIP and PIP2 returned to their initial values within 3 and 10 min, respectively, after PDGF addition. The level of PI continued to increase after it had returned to control values and was up threefold within 30 min after PDGF addition. In cells prelabeled with myo‐[3H]inositol PDGF caused an eightfold increase in the levels of inositol trisphosphate (IP3) within 2 min. Lesser increases, twofold and 1.3‐fold, respectively, were seen in levels of inositol bisphosphate (IP2) and inositol monophosphate (IP). Within 10 min after PDGF addition the levels of all three inositol phosphates had decreased to control values. The levels of IP3 measured 2 min after PDGF addition depended on the PDGF concentration and were maximal at 5–10 ng/ml of PDGF. Similar concentrations of PDGF stimulate maximal cell growth and DNA synthesis in these cells.
Insulin receptor capping and its correlation with calmodulin‐dependent myosin light chain kinase
Authors: Majerick Mary H. , Bourguignon Lilly Y. W. 
Published: February 2005

AbstractBoth fluorescence microscopy and fluorometric analysis techniques have been used to characterize insulin receptor capping in IM‐9 human lymphoblastoid cells. Morphologically, insulin caps appear similar to lectin or antiimmuno‐globulin‐induced caps displaying a preferential accumulation of actin, myosin, and actin‐binding protein directly underneath the cap structure. Using the fluorescent calcium indicator quin2 we have detected no change in the calcium activity following insulin stimulation. However, in the presence of a number of calmodulin inhibitors, such as W‐5, W‐7, W‐12, and trifluoperazine (TFP), insulin capping is significantly inhibited, which implies that a calmodulin‐regulated process is involved. Using double immuonofluorescence microscopy, we have found that the calmodulin‐dependent myosin light chain kinase (MLCK) is concentrated directly beneath insulin caps. Upon treatment with trifluoperazine (TFP), the redistribution of both MLCK and insulin receptors are inhibited concomitantly. Our data indicate that the calmodulin‐dependent myosin light chain kinase may be directly responsible for the activation of actomyosin‐mediated contractility during insulin receptor capping.
The in vitro autocrine secretion of CSFs alone does not account for the longterm growth of murine myeloid leukemic cells in suspension cultures
Authors: Fichelson Serge , Heard Jean‐Michel , Levy Jean‐Paul 
Published: February 2005

AbstractTo investigate the mechanisms supporting the in vitro longterm growth of murine leukemic myeloblastic cells, factor‐dependent and autonomous myeloblastic cells have been examined for their CSF responsiveness, CSF secretion, and autostimulation of growth. Purified CSF‐1, GM‐CSF, and IL‐3 stimulated cloning and proliferation of both autonomous (ACL) and factor‐dependent cell lines (FDCL), and were unable to induce differentiation of these cells. Sensitivities to CSFs were similar for ACL, FDCL and normal bone marrow cells. Most of the cell lines secreted CSFs, stimulating colony formation from normal bone marrow cells in bilayer agar cultures. The number of induced granulo‐macrophagic colonies was similar in the presence of either ACL or FDCL. The ability to stimulate their own proliferation was similar for both ACL and FDCL in a longterm suspension culture assay as well as in a shortterm cloning assay. These results strongly suggest that the autocrine secretion of CSF‐related activities are not sufficient to fully account for the in vitro longterm proliferation of virus‐transformed myelomonocytic cells.
A self‐renewing, bipotential erythroid/mast cell progenitor in continuous cultures of normal murine bone marrow
Authors: Wendling Françoise , Shreeve Mona , McLeod David , Axelrad Arthur 
Published: February 2005

AbstractWe have established permanent lines of nonadherent cells from fresh normal mouse bone marrow in media containing pokeweed mitogen‐stimulated spleen cell conditioned medium (PWSCM). These lines continuously produced erythropoietic progenitor cells (detected by their ability to form erythroid bursts in semi‐solid medium containing erythropoietin) together with cells having characteristics of the mast cell lineage (as demonstrated by metachromatic staining with toluidine blue, histamine content and membrane receptors for IgE). Sixteen such cell lines have been established in sixteen attempts. Cloning experiments were carried out to determine the nature of the progenitor cell(s) responsible for the permanence of these cultures. When cells were cultured in methylcellulose medium containing PWSCM, colonies were observed which reached macroscopic size after 4 weeks of incubation. Replating of individual primary colonies resulted in secondary colony formation, indicating the presence of progenitor cells with self‐renewal potential. Forty‐seven primary colonies were picked and their cells were suspended in liquid culture medium containing PWSCM. Of these, twenty‐one could be expanded to establish permanently growing sublines. Sixteen of these sublines were found to be composed of both erythroid progenitors and mast cells. In five sublines only mast cells could be seen; none of the sublines appeared to be purely erythroid. Karyotypic analysis of mast cells and of erythroid cells of seven sublines derived from individual colonies which arose in cocultures of male and female cells revealed that the mast cells and erythroid cells were both of the same sex in each of the seven sublines; this demonstrates the single cell origin of each colony and of the two lineages derived from it. We conclude that these nonadherent, factor‐dependent cell lines are maintained by self‐renewal and differentiation of bipotential progenitor cells apparently restricted to the erythroid and mast cell lineages.
SV40‐transformed human fibroblasts: Evidence for cellular aging in precrisis cells
Authors: Stein Gretchen H. 
Published: February 2005

AbstractPrecrisis SV40‐transformed human diploid fibroblast (HDF) cultures have a finite proliferative lifespan, but they do not enter a viable senescent state at end of lifespan. Little is known about either the mechanism for this finite lifespan in SV40‐transformed HDF or its relationship to finite lifespan in normal HDF. Recently we proposed that in normal HDF the phenomena of finite lifespan and arrest in a viable senescent state depend on two separate processes: (1) an age‐related decrease in the ability of the cells to recognize or respond to serum and/or other mitogens such that the cells become functionally mitogen‐deprived at the end of lifespan; and (2) the ability of the cells to enter a viable, G1‐arrested state whenever they experience mitogen deprivation. In this paper, data are presented that suggest that precrisis SV40‐transformed HDF retain the first process described above, but lack the second process. It is shown that SV40‐transformed HDF have a progressively decreasing ability to respond to serum as they age, but they continue to traverse the cell cycle at the end of lifespan. Concomitantly, the rate of cell death increases steadily toward the end of lifespan, thereby causing the total population to cease growing and ultimately to decline. Previous studies have shown that when SV40‐transformed HDF are environmentally serum deprived, they likewise exhibit continued cell cycle traverse coupled with increased cell death. Thus, these results support the hypothesis that precrisis SV40‐transformed HDF still undergo the same aging process as do normal HDF, but they end their lifespan in crisis rather than in the normal G1‐arrested senescent state because they have lost their ability to enter a viable, G1‐arrested state in response to mitogen deprivation.
Norepinephrine and epidermal growth factor: Dynamics of their interaction in the stimulation of hepatocyte DNA synthesis
Authors: Cruise Jennifer L. , Michalopoulos George 
Published: February 2005

AbstractPrimary cultures of adult rat hepatocytes are stimulated to enter DNA synthesis by norepinephrine (NE). This stimulation is maximal if the hepatocytes are incubated with NE for more than 12 hr, beginning no later than 2–4 hr after the cells are first plated. After 24 hr in culture, hepatocytes are unresponsive to NE stimulation. A strong synerergistic interaction between NE and epidermal growth factor (EGF) may be observed in cultures incubated with both EGF and NE, or pretreated with NE, then exposed to EGF. This interaction may be related to the finding that NE, in similarity with other factors that enhance EGF stimulation, reduces binding at the EGF receptor during the first 24 hr in culture.
The selective induction of a small number of proteins during G1 transit results from the mitogenic action of pp60v‐src in tsASV‐infected rat cells
Authors: Durkin Jon P. , Whitfield J. F. 
Published: February 2005

AbstractSince the mitogenic/oncogenic pp60v‐src product of the avian sarcoma virus (ASV) mutant, tsLA23, is abnormally thermolabile, tsLA23‐NRK cells were phenotypically nontransformed at 40°C and were consequently rendered quiescent by serum deprivation at this temperature. These serum‐deprived cells were stimulated to transit G1 either as transformed cells by simply dropping the temperature to a pp60v‐src‐activating 36°C, or as nontransformed cells by adding serum at 40°C. Serum stimulation rapidly increased total protein synthesis in these cells and over 100 changes in cellular proteins (resolved by two‐dimensional gel electrophoresis) occurred during G1 transit. By contrast, pp60v‐src‐activation did not increase total protein synthesis and only six proteins (18.5–44 kD) were clearly seen to appear or increase when quiescent cells were stimulated to transit G1 by activating pp60v‐src. Three of these six pp60v‐src‐ induced proteins also appeared or accumulated during the G1transit of serum‐stimulated cells. The appearance and/or accumulation of the six proteins and the subsequent initiation of DNA replication may have resulted from pp60v‐src stimulating only a small number of critical cellular genes because both the protein changes and DNA replication were completely suppressed by the transcription inhibitor actinomycin D.
Two classes of primitive pluripotent hemopoietic progenitor cells: Separation by adherence
Authors: Kerk David K. , Henry Elizabeth A. , Eaves Allen C. , Eaves Connie J. 
Published: February 2005

AbstractMethylcellulose cultures containing mouse marrow cells at low densities and partially purified preparations of erythropoietin and Interleukin‐3 were scored after 2 weeks for the presence of macroscopic multilineage colonies (from “primary” CFU‐macro GEMM). Whole cultures were then harvested and replated to assess the number of “secondary” CFU‐macro GEMM produced, but not detected, during the primary culture period. In such experiments adherent marrow cells yielded significantly higher numbers of secondary CFU‐macro GEMM than did either fresh or nonadherent marrow cells. Removal of macroscopic colonies prior to replating showed that most secondary CFU‐macro GEMM were not derived from primary CFU‐macro GEMM. In vivo studies also revealed a differential effect of adherence separation on the frequency of day 10 CFU‐S, which decreased, by comparison to cells capable of long‐term repopulation, which increased. Primary adherent CFU‐macro GEMM from 5‐fluorouracil (5‐FU) treated mice showed an 18‐fold higher self‐renewal capacity than their counterparts in normal marrow. Nevertheless the majority of secondary CFU‐macro GEMM obtained from primary cultures of adherent 5‐FU cells were again not derived from primary CFU‐macro GEMM. Cells capable of immediately generating large multilineage colonies thus appear to represent an intermediate compartment of pluripotent progenitors whose self‐renewal properties, may, however, vary over a considerable range. Our results further suggest that these progenitors are derived ultimately from a more primitive adherent cell whose tendency to begin to divide in vitro is low and whose presence correlates with cells capable of long‐term myeloid repopulation in vivo.
Differential response of normal rat mammary epithelial cells to mammogenic hormones and EGF
Authors: McGrath Michael , Palmer Sarah , Nandi Satyabrata 
Published: February 2005

AbstractA simple dissociation procedure and the collagen gel culture system have been utilized to determine the effects of mammogenic hormones and epidermal growth factor (EGF) on the proliferation of normal rat mammary epithelial (RME) cells in serum‐free culture. Epithelial fragments, isolated from normal virgin F344 rat mammary glands by enzyme digestion followed by Percoll density gradient centrifugation, were embedded within a rat tail collagen matrix. A three‐to four‐fold increase in cell number was observed when ovine prolactin (PRL) and progesterone (P) were present in the basal medium during 7 days of culture. Mouse EGF stimulated one cell doubling during the same culture period.Isolated mammary organoids produced a ‘stellate’ type colony when PRL + P were present in the culture medium. These colonies were composed of small, tightly packed cuboidal cells. The addition of EGF to the basal medium produced a diffuse ‘basket’ type colony which was composed of large, elongate cells. When the complete hormonal and growth factor combination (PRL + P + EGF) was present, a ‘mixed’ type colony was observed which contained both the large and small epithelial cell types.Immunocytochemical analysis revealed that both the cuboidal and elongate cells present in the two colony types stained with antibodies to keratin indicating that these cells were epithelial in nature. The small cuboidal cells also expressed thioesterase II and alpha‐lactalbumin, both specific for secretory mammary epithelial cells. The large, elongate cell type, however, was positive for actin but did not stain for either secretory epithelial specific marker. The results reported here suggest that normal rat mammary tissue may contain two epithelial populations, one which responds to PRL + P and the other which responds to EGF.
Cell cycle phase‐dependent induction of ornithine decarboxylase‐antizyme
Authors: Linden Margareta , Anehus Siw , Långström Eva , Baldetorp Bo , Heby Olle 
Published: February 2005

AbstractThe activites of ornithine decarboxylase (ODC) and ODC inhibitory protein (ODC‐antizyme) were studied in Ehrlich ascites tumor cells, separated according to their position in the cell cycle by centrifugal elutriation. Release and/or synthesis of ODC‐antizyme was induced by putrescine treatment. Each mouse received an intraperitoneal injection of 25 μmoles of putrescine at 0, 1, 2, and 3 hr after tumor transplantation. Tumor cells obtained from putrescine‐treated and control mice at 4 hr after transplantation were separated into fractions representing all phases of the cell cycle. The cell cycle distribution of the tumor cells in each fraction was determined by flow cytometry.In control tumor cells the ODC activity exhibited two maxima; inlate‐G1/early‐S and in late‐S/G2. A marked decrease in ODC activity was observed in mid‐S phase. This decrease coincided with maximum ODC‐antizyme activity (revealed by putrescine treatment), suggesting that ODC‐antizyme is involved in the regulation of ODC activity during the cell cycle.
Hormonal responses to 1,25‐dihydroxyvitamin D3 in cultured mouse osteoblast‐like cells—modulation by changes in receptor level
Authors: Chen Theresa L. , Li Janet M. , Van Ye Todd , Cone Charlotte M. , Feldman David 
Published: February 2005

AbstractThe level of 1,25(OH)2D3 receptors in cultured mouse osteoblast‐like (OB) cells is modulated by the rate of cell proliferation. We have studied two 1,25(OH)2D3‐induced bioresponses to ascertain whether the changes in receptor levels during growth in culture alter cell responsiveness. Nuclear receptor levels were high (127 fmol/100 μgDNA) in rapidly dividing (log) cells and low (25 fmol/100 μg DNA) in quiescent (confluent) cells. The bioresponses we studied were induction of 25(OH)D3‐24‐hydroxylase. activity (24‐hydroxylase) and inhibition of collagen synthesis. The basal levels of 24‐hydroxylase were low and similar in cells at log growth phase and confluence. At a maximal induction dose of 13 nM, 1,25(OH)2D3 induced a three‐fold rise in enzyme activity at long growth phase, but only caused less than two‐fold rise at confluence. The half‐maximal dose (ED50) was slightly shifted from 0.6 nM to 0.8 nM. Daily measurement of 1,25(OH)2D3 receptor levels and maximal induction of 24‐hydroxylase activity throughout the culture cycle showed a strong correlation between receptor abundance and enzyme induction. The basal level of collagen synthesized by cells in log growth phase was ∼ 5% and increased to ∼ 8% at confluence. Maximal inhibition of collagen synthesis by 1,25(OH)2D3 reached 80% of control levels in log cells, but was only 40% of control in confluent cells. The ED50 was ∼ 0.1 nM in the log cells and increased to ∼ 1 nM at confluence. Daily assay of 1,25(OH)2D3 receptor levels and 1,25(OH)2D3 responses during the culture cycle indicated a correlation between changes in receptor level and the extent of inhibition of collagen synthesis. These changes in biore‐sponse at various growth phases did not occur in rat OB cells where the 1,25(OH)2D3 receptor levels were independent of cell proliferation. The results indicate that cell proliferation rate, via change in receptor levels, determines the magnitude and sensitivity of the cellular responses to 1,25(OH)2D3.
Endocytosis in sickle erythrocytes: A mechanism for elevated intracellular Ca2+ levels
Authors: Rubin Elizabeth , Schlegel Robert A. , Williamson Patrick 
Published: February 2005

AbstractStaining of sickle cells with the fluorescent probes chlortetracycline (a Ca2+probe) and diindocarbocyanine (a general membrane probe) revealed the presence of Ca2+‐containing vesicles which are not found in normal erythrocytes. These vesicles increase in number upon deoxygenation, and are apparently formed by endocytosis, as judged by the use of the extracellular fluorescent probe lucifer yellow. The presence of vesicles is not restricted to any particular morphological or density class of cells in the general population.
Utilization of pyruvate and pyruvate precursors by normal and carcinogen‐altered rat tracheal epithelial cells in culture
Authors: Wasilenko William J. , Marchok Ann C. 
Published: February 2005

AbstractThe metabolism of [14C]pyruvate, [14C]glucose, [14C]glutamine and [14C]alanine was compared between normal rat tracheal epithelial cells and carcinogen‐altered cells derived from dimethylbenz(a)anthracene‐exposed tracheal implants. Normal primary cultures (NPC) of tracheal cells are distinguished by their need for pyruvate‐supplemented medium for growth and survival. The altered cells were selected out by their survival in the unsupplemented medium. Compared to the selected primary cultures (SPC), the NPC showed a three‐ to four‐fold higher incorporation of radioactivity from [2‐14C]pyruvate in all the macromolecular fractions, as well as in all the metabolites isolated from the acid soluble fraction and from lactic acid isolated from the medium. [U‐14C]glucose was also incorporated at higher levels into lactic acid isolated from the acid soluble fraction and the medium of NPC. These data indicate a higher rate of glycolysis in the normal tracheal cells. This was supported by the findings of a two‐fold greater glucose consumption and two‐fold higher production of lactic acid isolated from the NPC medium. Lactate dehydrogenase activity was also two‐fold higher in NPC. Thus, despite the apparently higher level of pyruvate production in the NPC, exogenous pyruvate is necessary to satisfy the metabolic needs of NPC. The utilization of [U‐14C]glutamine or [U‐14C]alanine was not markedly different between NPC and SPC. Furthermore, radioactivity from both of the amino acids was recovered in lactic acid in the medium, indicating that both cell types can derive pyruvic acid from either glutamine or alanine. SPC apparently do not use these routes to supply higher levels of pyruvic acid for survival in culture. The oxidation of none of the radioactive metabolites into CO2 was distinctly different between NPC and SPC except for the 1.7‐fold higher utilization of [1‐14C]glucose along the oxidative arm of the pentose cycle in the normal cells.