bluuebunny/crossref_metadata_2025_split huggingface dataset:
Biological properties of a hepatocyte growth factor from rat platelets
Authors: Russell William E. , McGowan Joan A. , Bucher Nancy L. R. 
Published: February 2005

AbstractIn an accompanying communication we demonstrated that about half of the potency of rat serum to stimulate DNA synthesis in cultured adult rat hepatocytes resides in a polypetidelike substance from the platelets. A lysate of rat platelets was able to restore the potency of platelet‐poor rat serum, whereas a lysate of human platelets inhibited thymidine incorporation by the hepatocytes. Moreover, addition to these cultures of either highly purified human platelet‐derived growth factor (PDGF) or human platelet factor 4 (PF‐4) failed to influence DNA synthesis either alone or in the presence of rat or human platelet‐poor serum, which is required for expression of PDGF activity. Unlike the human platelet factors, rat platelet lysate (RPL) was moderately active by itself and was augmented equally well by platelet‐poor serum from either source. At concentrations below 5%, platelet‐poor serum from hypophysectomized rats was as potent as that from normal rats in augmenting RPL activity. This suggests that, unlike PDGF, which is not activated by hypophysectomized rat serum, the hepatotrophic component of RPL does not require the presence of exogenous somatomedins for activity, but interacts instead with other plasma constituents or with somatomedins produced by the hepatocytes in vitro. Rat platelets do, however, appear to contain PDGF or its rat equivalent in addition to the hepatocyte growth factor, since if they are heated to 100°C for 10 min, their ability to stimulate nuclear labeling in confluent BALB/c 3T3 cells is not impaired, while their ability to stimulate DNA synthesis in rat hepatocytes is destroyed. These studies indicate that the hepatocyte growth factor from rat platelets differs from PDGF in its biological as well as physical characteristics, but that rat platelets also contain PDGF or an equivalent substance.
Maintenance of granulocyte‐monocyte progenitor cells in liquid cultures of human foetal liver
Authors: Toksoz Deniz , Brown Geoffrey 
Published: February 2005

AbstractThese studies describe a liquid suspension culture system for normal myeloid cells derived from human foetal liver. A simple one‐step fractionation procedure was employed to obtain a cell population capable of expanding into all stages of myeloid differentiation, including committed myeloid progenitor cells (GM‐CFC). Cell proliferation in these cultures resulted in the maintenance of early myeloid populations for up to a month. In order to extend myeloid cell maintenance, a specific factor in the form of media conditioned by human endothelial cells (endo C.M.) was used. Addition of endo C.M. to foetal liver cultures resulted in increased myeloid proliferation coupled to extensive myeloid differentiation. Clonally derived foetal liver culture cells proliferated for up to 2 months in the presence of endo C.M. before maturing into macrophages. These results show that endo C.M. exert an extensive proliferative effect on early myeloid cells as well as inducing their differentiation. The large quantity of cells in early stages of myeloid differentiation provided by foetal liver cultures may be useful for biochemical and molecular biology studies of myelopoiesis. In addition, these cultures are a potential source from which to derive normal myeloid lines. The separation of the potent proliferative activity present in endo C.M. may yield an effector which maintains human myeloid cell proliferation.
Insulin inhibits the glucocorticoid‐mediated increase in hepatocyte EGF binding
Authors: Lin Qixiong , Blaisdell Joyce , O'Keefe Edward , Earp H. Shelton 
Published: February 2005

AbstractHydrocortisone and dexamethasone produced a time‐dependent increase [125I]epidermal growth factor ([125I]EGF) binding in primary cultures of isolated rat hepatocytes. Maximally effective doses of glucocorticoids resulted in a 70–100% increase in binding. The effect was similar when hepatocytes were maintained on collagen‐coated plates or directly on culture dishes. The glucocorticoid‐mediated increase in [125I]EGF binding could be detected after 4 h exposure to glucocorticoid and was substantial by 8 h. The major effect of glucocorticoid appeared to be to increase the number of EGF receptors. While insulin (100 nM) had no effect on basal [125I]EGF binding, it significantly inhibited the increase produced by the glucocorticoid. Since the inhibitory effect of insulin was only observed when insulin was added with the inducing glucocorticoid, insulin appears to inhibit an early hydrocortisone‐mediated event.
Rb+ influx in response to changes in energy generation: Effect of the regulation of the ATP content of HELa cells
Authors: Ikehara Toshitaka , Yamaguchi Hisao , Hosokawa Keiko , Sakai Tetsuhiro , Miyamoto Hiroshi 
Published: February 2005

AbstractThe effect of changes in energy metabolism on Rb+ influx was studied in HeLa cells. Irrespective of whether ATP production was controlled by varying the activity of glycolysis or of oxidative metabolism on addition of certain combinations of glucose, carbonylcyanide m‐chlorophenylhydrazone, monoiodoacetic acid, and quercetin, Rb+ influx changed as a linear function of the ATP content, which varied in a wide range up to the normal level (15–20 nmol/mg protein or 3–4 mM). The difference between results obtained by these procedures was not significant. As the intracellular Na+ content varied at different ATP contents, the Na+ content was adjusted to similar levels by chilling the cells with varying ATP contents. However, a linear relation was still observed. A similar dependence was also obtained for cytoplasmic ATP, which would be more closely connected with the Na, K‐pump than total ATP. The ratio of ouabain‐sensitive Rb+ influx to the corresponding part of lactate production was nearly 2 in the presence of 2 mM glucose. From these results it is concluded that (1) active Rb+ influx, which is chiefly maintained by energy generated through glycolysis, can also be supported by oxidative metabolism; (2) Rb+ influx is regulated linearly as a function of the cellular ATP content up to the control level; but does not increase if ATP is raised still further; and (3) 2 Rb+ ions move concomitantly at the expense of one ATP molecule.
Analysis of cell division by time‐lapse cinematographic studies of hydrocortisone‐treated embryonic lung fibroblasts
Authors: Absher Marlene , Cristofalo Vincent J. 
Published: February 2005

AbstractHydrocortisone is a modulator of cell division and has been shown to prolong the replicative in vitro life span of human embryonic lung fibroblasts. Time lapse cinematography was used to analyze the proliferative behavior of individual cells in populations of fibroblasts exposed to hydrocortisone in young cultures during a single growth cycle and in aged cultures that had been continuousiy exposed to hydrocortisone. Results indicate that hydrocortisone causes a decrease in the interdivision time (IDT) of a portion of the cells in the population and this effect is augmented after continuous exposure to hydrocortisone. Hydrocortisone does not appear to increase the number of initial dividers in the population but increases growth rate in the early stages of the culture period. Analysis of mother‐daughter IDT pairs further suggests that hydrocortisone exerts its effects on IDT independently for a given cell.
Mitogenic and comitogenic properties of the indole alkaloid class of tumor promoters
Authors: Novogrodsky Abraham , Patya Miriam , Fujiki Hirota , Rubin Albert L. , Stenzel Kurt H. 
Published: February 2005

AbstractWe determined the mitogenic and comitogenic properties of tumor promoters in the indole alkaloid series; agents that differ structurally from 12–0‐tetradecanoylphorbol‐13‐acetate (TPA), which is known to be a lymphocyte mitogen. Teleocidin and dihydroteleocidin B were mitogenic for human lymphocytes, and lyngbyatoxin A elicited little or no mitogenesis. Catalase enhanced the mitogenicity of teleocidin and dihydroteleocidin B, and lyngbyatoxin A was also mitogenic in the presence of catalase. The potency of the agents was TPA = teleocidin > dihydroteleocidin B > lyngbyatoxin A. Dimethylsulfoxide and butyric acid markedly inhibited proliferation induced by these agents in human lymphocytes. The indole alkaloid tumor promoters were all comitogenic for murine thymocytes and induced production of interleukin‐2. While the comitogenic effect of teleocidin was similar to that of TPA in its susceptibility to inhibition by dimethylsulfoxide and butyric acid, the comitogenic effect of dihydroteleocidin B and lyngbyatoxin A were less susceptible to these agents. These findings may facilitate the identification of cellular sites responsible for the inhibitory effect of differentiating agents on proliferative responses of lymphocytes.
Effect of retinoic acid on the proliferation and phagocytic capability of murine macrophage‐like cell lines
Authors: Goldman Rachel 
Published: February 2005

AbstractRetinoic acid (RA) exerted a variable degree of growth inhibitory activity on the macrophage‐like cell lines P388D1, J774.2, WEHI‐265, WEHI‐3, and PU‐5. Comparison of cell proliferation and clonal growth suggests that at concentrations of 10−9‐10−6 M the inhibitory activity stems from processes leading to elongation of cell cycle time and not from terminal differentiation processes. RA was shown to be a potent inducer of the development of high‐phagocytic phenotypes (assessed by phagocytosis of heat‐killed yeast cells) in the P388D1, J774.2, and WEHI‐265 cell lines which differ substantially in their proliferative and adherence characteristics. The PU‐5 and WEHI‐3 cell lines were not induced by RA to express an enhanced phagocytic activity toward heat‐killed yeast cells. The augmented phagocytic capability was dose dependent over a wide range of RA concentrations. In P388D1 cells, 2 × 10−12 M RA already exerted significant phagocytosis augmentation effects, which progressively increased up to 2 × 10−5 M RA, the highest concentration tested. Retinal, retinyl acetate, and retinol had similar effects to those of RA on both cell adherence and phagocytosis in P388D1 cells, albeit at concentrations four to six orders of magnitude higher. Optimal development of the high‐phagocytic phenotype in P388D1, J774.2, and WEHI‐265 cells required at least 96 hr of culture in the presence of RA; at 48 hr and 23 hr the effects were already substantial, whereas at 4 hr of exposure to RA no significant enhancement of phagocytosis could be detected. Thus both extended periods of culture in the presence of RA (more than two to three cell cycles) and high concentrations were needed for induction, in more than 90% of the cells, of the expression of a high‐phagocytic phenotype. The reversion to a low‐phagocytic phenotype upon removal of RA was also rather slow and required several cell cycles. In P388D1 cells RA also enhanced the phagocytosis of latex beads but had no effect on the phagocytosis of starch particles, or the extent of binding of immunoglobulin G‐coated sheep red blood cells (SRBC). The expression of receptors for concanavalin A and for nonopsonized SRBC remarkably increased in RA‐treated cells, as was the ability to perform Fc‐receptor mediated erythrophagocytosis. Both P388D1 cells and WEHI‐265 cells were induced by RA to express nitroblue tetrazolium reducing activity. The data suggest that RA induces profound changes in the functional capabilities of macrophage‐like cell lines which are apparently not dependent on cessation of growth and terminal differentiation processes.
Mutants of chinese hamster cells deficient in thymidylate synthetase
Authors: Li I‐Chian , Chu Ernest H. Y. 
Published: February 2005

AbstractStable mutants of Chinese hamster V79 cells deficient in thymidylate synthetase (TS; E.C. 2.1.1.45) have been selected from cultures grown in medium supplemented with folinic acid, aminopterin, and thymidine (FAT). After chemical mutagenesis, the frequency of colonies resistant to the “FAT” medium increased more than 100‐fold over the spontaneous frequency. The optimal expression time of the mutant phenotype was 5–7 days after mutagen treatment. The recovery of FAT‐resistant colonies in the selective medium was not affected by the presence of wild‐type cells at a density below 9,000 cells per cm2. All 21 mutants tested exhibited thymidine auxotrophy; neither folinic acid nor deoxyuridine could support mutant cell growth. There was no detectable TS activity in all 11 mutants so far examined and only about 50% of wild‐type activity in three prototrophic revertants, as measured by whole‐cell and cell‐free enzyme assays. The apparent Michaelis‐Menten constant (Km) for deoxyuridine‐5′‐monophosphate and inhibition constant (Ki) for 5‐fluorodeoxyuridine‐5′‐monophosphate, measured by whole‐cell enzyme assay, appear to be similar for the wild‐type and revertant cell lines. Using 5‐fluoro‐[63H]‐2′‐deoxyuridine 5′‐monophosphate as active site titrant, the relative amounts of TS in crude cell extract from the parental, revertant, and mutant cells were shown to exist in a 1:0.5:0 ratio. Furthermore, the enzymes from two revertants were more heat labile than that of V79 cells. These properties, taken together, suggest that the FAT‐resistant, thymidine auxotrophic phenotype may be the result of a structural gene mutation at the TS locus. The availability of such a mutant facilitates studies on thymidylate stress in relation to DNA metabolism, cell growth, and mutagenesis.
Endocytosis and exocytosis of protein in capillary endothelium
Authors: Williams Stuart K. , Greener Deborah A. , Solenski Nina J. 
Published: February 2005

AbstractThe transport of proteins across continuous capillary endothelium is believed to be mediated by micropinocytic vesicles which shuttle molecules between the lumenal and abluminal plasma membrane. We have studied the ability of capillary endothelial cells isolated from rat epididymal fat to endocytose fluorescently labelled ovalbumin within micropinocytic vesicles. Net association of fluorescent ovalbumin with endothelial cells reaches an equilibrium after 40 minutes of incubation. This equilibrium is presumably due to a balance between endocytosis and subsequent exocytosis of this protein. Capillaries equilibrated with fluorescent ovalbumin exhibited rapid exocytosis of this protein when it was removed from the external medium. The rate of endocytosis was concentration dependent and obeyed the kinetics expected for adsorptive phase endocytosis. High concentrations of ovalbumin stimulated the ingestion of 14C‐sucrose, a marker of fluid endocytosis, suggesting that protein can affect the movement of vesicles within the endothelial cytoplasm. These results imply that capillary endothelium isolated from rat epididymal fat exhibits the ability to endocytose and subsequently exocytose protein. This demonstrates that the two components of endothelial vesicular transport or transcytosis can be observed and studied in a system of isolated capillary endothelium.
Dependence of lethality induced by a direct DNA perturbation of synchronized human diploid fibroblasts on different periods of the DNA synthetic period (S Phase)
Authors: Tsutsui Takeki , Ohmori Manabu , Suzuki Nobuko , Maizumi Heiji 
Published: February 2005

AbstractThe cytotoxic effect of a direct perturbation of DNA during various portions of the DNA synthetic period (S phase) of cultured human diploid fibroblasts was examined. The cells were synchronized by a period of growth in low serum with a subsequent blockage of the cells at the G1/S boundary by hydroxyurea. This method resulted in over 90% synchrony, although approximately 20% of the cells were noncycling. Synchronized cells were treated for each of four 2‐hour periods during the S phase with 5‐bromodeoxyuridine (0.1–10 μM), followed by irradiation with near‐UV (5–10 min). The 5‐bromodeoxyuridine‐plus‐irradiation treatment was cytotoxic, while treatment with 5‐bromodeoxyuridine alone or irradiation alone was not cytotoxic. The cytotoxicity was dependent upon the periods of S phase during which treatment was administered. The highest lethality was observed for treatment in early to middle S phase, particularly in the first 2 hours of S phase, whereas scarce lethality was observed in late S phase. The extent of substitution of 5‐bromodeoxyuridine for thymidine in newly synthesized DNA was similar in every period of the S phase. Furthermore, no specific period during S phase was significantly more sensitive to treatment with respect to DNA damage, as determined by an induction of unscheduled DNA synthesis. These results suggest that a certain region or regions in the DNA of human diploid fibroblasts, as designated by their specific temporal relationship in the S phase, may be more sensitive to the DNA perturbation by 5‐bromodeoxyuridine treatment plus near‐UV irradiation for cell survival.
The role of polyamines in somatomedin‐stimulated differentiation of L6 myoblasts
Authors: Ewton Daina Z. , Erwin Bradley G. , Pegg Anthony E. , Florini James R. 
Published: February 2005

AbstractThe somatomedins are potent stimulators of proliferation and differentiation of cultured myoblasts. In studies on the mechanism(s) of these actions, we have measured the activities of ornithine decarboxylase (ODC), an enzyme associated with rapid cell proliferation, and creatine kinase (CK), a biochemical marker for muscle differentiation, after treatment of L6 myoblast cultures with Multiplication Stimulating Activity (MSA), a member of the somatomedin family of insulinlike growth factors. ODC levels reached a peak 24 hours after MSA addition (before any detectable differentiation of the myoblasts) and then decreased as differentiation commenced and CK activity increased. Addition of alpha‐difluoromethylornithine (DFMO), an irreversible inhibitor of ODC, caused a dramatic decrease in differentiation. Measurement of 3H‐thymidine incorporation, DNA content, and cell number established that the effect of DFMO on differentiation was not a simple consequence of its antiproliferative actions. Cellular levels of putrescine and spermidine (but not spermine) decreased substantially following addition of DFMO to the cultures. The inhibitory effects of DFMO were abolished upon addition of exogenous polyamines to the medium. However, addition of polyamines in the absence of MSA or DFMO did not mimic the stimulation of differentiation by MSA. We conclude that polyamines play an essential role in the stimulation of L6 myoblast differentiation by somatomedins, but they are not sufficient to effect this stimulation.
Verapamil enhances the toxicity of conjugates of epidermal growth factor with Pseudomonas exotoxin and antitransferrin receptor with pseudomonas exotoxin
Authors: Akiyama Shin‐Ichi , Gottesman Michael M. , Hanover John A. , Fitzgerald David J. P. , Willingham Mark C. , Pastan Ira 
Published: February 2005

AbstractVerapamil, a clinically important calcium channel blocker, has been found to cause a 40‐fold enhancement of killing of the human KB cell line by a cytotoxic conjugate of epidermal growth factor with Pseudomonas exotoxin (EGF‐PE). Synergistic effects of verapamil and EGF‐PE are also seen on HeLa D98 cells and a human epidermal carcinoma cell line, A431. Verapamil also potentiates the effect of a toxic conjugate formed between Pseudomonas exotoxin and a monoclonal antibody to the human transferrin receptor (anti‐TFR‐PE) and enhances the effect of Pseudomonas exotoxin (PE) alone. Two other calcium antagonists were tested. Diltiazem enhances the cytotoxic effect of EGF‐PE, but nifedipine does not. Verapamil does not affect the binding and uptake of 125I‐EGF by KB cells, but it significantly delays the disappearance of internalized 125I‐EGF from the cells. Density gradient fractionation studies using cell homogenates suggest that 125I‐EGF accumulates in an undegraded form in lysosomes when cells are treated with verapamil. By immunofluorescence microscopy using an antibody to PE, EGF‐PE was found to accumulate in lysosomes; by electron microscopy the lysosomes had an abnormal appearance. The effects of verapamil on toxicity of EGF‐PE and lysosomal function appear to be related. However, it is not known whether the enhanced toxicity of EGF‐PE in the presence of verapamil is due to its delayed degradation in lysosomes or some more general effect of verapamil on membrane permeability.
Characterization of the metabolic turnover of epidermal growth factor receptor protein in A‐431 cells
Authors: Stoscheck Christa M. , Carpenter Graham 
Published: February 2005

AbstractThe metabolism of the receptor for epidermal growth factor (EGF) in A‐431 cells has been measured by labeling the receptor in vivo with radioactive amino acid precursors and then determining, by immunoprecipitation with specific anti‐EGF receptor antisera, the rate of degradation of the receptor when the cells are placed in a nonradioactive medium. The rate of EGF receptor degradation (t½12 = 20 hr) was faster than the rate of degradation of total cell protein (t½12 = 52 hr). When EGF was added at the beginning of the chase, the half‐life of prelabeled receptor decreased to 8.9 hr. This decrease was specific, as the level of total cellular protein and another plasma membrane protein, the transferrin receptor, were relatively unaffected by EGF. The carbohydrate portion of the receptor is degraded, in the presence or absence of EGF, at approximately the same rate as the protein moiety. The amount of EGF receptor protein in A‐431 cells has been quantitated by radiolabeling total cellular protein and quantitating the immunoprecipitable receptor. The EGF receptor constitutes approximately 0.15% of the total cell protein in A‐431 cells. These cells, therefore, have approximately 30 times more EGF receptor protein than fibroblasts. The EGF receptor constitutes an even higher proportion of 3H‐glucosamine‐ or 3H‐mannose‐labeled macromolecules in A‐431 cells, 1.5% or 5.2%, respectively. The EGF receptor from A‐431 cells can easily be identified by submitting carbohydrate‐labeled, solubilized cells to electrophoresis as described by Laemmli (1970).
Growth requirements of low‐density rabbit costal chondrocyte cultures maintained in serum‐free medium
Authors: Kato Yukio , Gospodarowicz Denis 
Published: February 2005

AbstractThe factors required for the active proliferation of low‐density rabbit costal chondrocytes exposed to 9:1 (v/v) mixture of Dulbecco's modified Eagle's medium and Ham's F12 medium have been defined. Low‐density primary cultures of rabbit costal chondrocytes proliferated actively when the medium was supplemented with high‐density lipoprotein (300 μg/ml), transferrin (60 μg/ml), fibroblast growth factor (FGF) (1 ng/ml), hydrocortisone (10−6 M), and epidermal growth factor (EGF) (30 ng/ml). Insulin, although it slightly decreased the final cell density, was required for reexpression of the cartilage phenotype at confluence. Optimal proliferation of low‐density chondrocyte cultures was only observed when dishes were coated with an extracellular matrix (ECM) produced by cultured corneal endothelial cells, but not on plastic. Furthermore, serum‐free chondrocyte cultures seeded at low density and maintained on ECM‐coated dishes gave rise to a homogeneous cartilage‐like tissue composed of spherical cells. These chondrocytes therefore seem to provide a good experimental system for analyzing factors involved in supporting proliferation of chondrocytes and their phenotypic expression.