bluuebunny/crossref_metadata_2025_split huggingface dataset:
Abortive transformation of temperature‐sensitive mutants of rat 3Y1 cells by simian virus 40: Effect of cellular arrest states on entry into S phase and cellular proliferation
Authors: Mitsudomi Tetsuya , Kimura Genki 
Published: February 2005

AbstractFour temperature‐sensitive (ts) mutants of rat 3Y1 fibroblasts, representing independent complementation groups, cease to proliferate predominantly with a 2n DNA content, at the restrictive temperature (39.8°C)(temperature arrest) or at the permissive temperature (33.8°C) at a confluent cell density (density arrest) (Ohno et al., 1984). We studied the temperature‐ or the density‐arrested cells of these mutants infected with simian virus 40 (SV40) or its mutants affecting large T or small t antigen with respect to kinetics at 39.8°C of entry into S phase and cellular proliferation. Three mutants, 3Y1tsD123, 3Y1tsF121 and 3Y1tsG125, expressed T antigen and entered S phase at 39.8°C from both the arrested states after infection with either wild‐type, tsA mutants, or a .54/.59 deletion mutant of SV40, whereas in the density‐arrested 3Y1tsH203, expression of T antigen and entry into S phase were inefficient and ts. Following the WT‐SV40 induced entry into S phase, the temperature‐arrested 3Y1tsD123 detached from the substratum with no detectable increase in cell number, whereas the density‐arrested ones completed a round of the cell cycle and then detached. 3Y1tsF121 and 3Y1tsG125 in the both arrested states proliferated through more than one generation. 3Y1tsF121 and 3Y1tsG125 in the density‐arrested state infected with tsA mutants once proliferated and then ceased to increase in number as the percentage of T‐antigen positive population decreased. These results suggest that wild‐type and tsA‐mutated large T antigens are able to overcome the cellular ts blocks of entry into S phase in the 3 ts mutants of 3Y1 cells in both the arrested states, and that small t antigen is not required to overcome the blocks. It is also suggested that cellular behaviors subsequent to S phase (viability, mitosis, and proliferation in the following generations) depend on cellular arrest states, on traits of cellular ts defects, and on the duration of large T antigen expression.
Defective endocytosis of low‐density lipoprotein in monensin‐resistant mutants of the mouse Balb/3T3 cell line
Authors: Tomita Kichinobu , Ono Mayumi , Masuda Akinori , Akiyama Shin‐Ichi , Kuwano Michihiko 
Published: February 2005

AbstractTwo monensin‐resistant clones show similar low‐density lipoprotein binding activity but less internalization or degradation of low‐density lipoerotein than the parental Balb/3T3 or other resistant clone. Sterol synthesis from radioactive acetate in the resistant mutant, MO‐5, is inhibited by more than 70% of control in the presence of tenfold higher amounts of low‐density lipoprotein than the dose that inhibits the parental Balb/3T3 to similar level. 3‐Hydroxy‐3‐methylglutaryl coenzyme A reductase activity of Balb/3T3 and MO‐5 is inhibited by 48% and 27% of control, respectively, in the presence of 10 μg/ml of low‐density lipoprotein. Colloidal silica gradient centrifugation shows that transport of low‐density lipoprotein from the surface membrane to the lysosome is much slower in MO‐5 cells than in Balb/3T3 cells. Down regulation of low‐density lipoprotein receptors on the cell surface in Balb/3T3 is observed by exposing the cells to 5–15 μg/ml low‐density lipoprotein, whereas only slight if any down regulation is observed when MO‐5 cells are treated with low‐density lipoprotein. The altered endocytosis of low‐density lipoprotein behaves as a dominant trait in hybrids of MO‐5 and THO2‐2, a derivative of Balb/3T3 resistant to both ouabain and 6‐thioguanine.
Lipoproteins increase growth of mitogen‐stimulated arterial smooth muscle cells
Authors: Libby Peter , Miao Paul , Ordovas Jose M. , Schaefer Ernst J. 
Published: February 2005

AbstractThe relationship between lipoproteins and growth of aortic smooth muscle cells has been a matter of controversy. We therefore reexamined this issue using serum‐free defined media methodology. By themselves, LDL or HDL (50–500 μg/ml) from normolipemic human or bovine plasma produced little or no growth of homologous aortic smooth muscle cells incubated in serumfree medium that was supplemented with insulin and transferrin to maintain cell viability. In fact, LDL prepared in the absence of an antioxidant (BHT) was toxic to these cells. However, in the presence of maximally effective concentrations of platelet‐derived growth factor (PDGF), LDL or HDL consistently increased the growth of homologous smooth muscle cells (up to twofold increases in DNA accumulation in 48 hr). Lipoproteins also augmented the growth response of arterial smooth muscle cells to fibroblast growth factor or epidermal growth factor. The mechanism of this effect was investigated further with HDL, because, in contrast to LDL, HDL apoproteins are water‐soluble. Neither HDL delipidated by solvent extraction (apoHDL), purified bovine apoA‐I, nor cholesterol added in the form of phospholipid vesicles appreciably increased PDGF‐induced growth of bovine smooth muscle cells. However, HDL‐like particles reconstituted by sonication of apoHDL with cholesterol and phospholipids did increase the growth of cultures of bovine smooth muscle cells treated with PDGF. Uptake of tritiated thymidine by cultures incubated with partially purified PDGF alone (10 μg/ml) was 5,693 ± 235 dpm/24 hr compared to 10,381 ± 645 dpm/24 hr (p < 0.01) in the presence of both PDGF and reconstituted HDL‐like particles (250 μg protein/ml). Thus both the lipid and protein components of HDL may be necessary for optimal potentiation of growth of mitogen‐stimulated cells. These results indicate that lipoproteins from normolipemic sera are not bona fide growth factors but can potentiate the growth of mitogen‐stimulated cells, perhaps by supplying exogenous cholesterol required for membrane biogenesis. This finding might be important in arterial injury when the release of PDGF and exposure to plasma lipoproteins could act in concert to stimulate the proliferation of smooth muscle cells.
The fate of DNA originally existing in the zygote nucleus during achromosomal cleavage of fertilized echinoderm eggs in the presence of aphidicolin: Microscopic studies with anti‐DNA antibody
Authors: Yamada Hisafumi , Hirai Setsuro , Ikegami Susumu , Kawarada Yoshihiko , Okuhara Eiji , Nagano Hiroshi 
Published: February 2005

AbstractIn fertilized echinoderm eggs, the male and female pronuclei fuse to form the diploid nucleus even in the presence of aphidicolin, a specific inhibitor of eukaryotic DNA polymerase‐α. The subsequent first cleavage is independent of chromosomes but dependent on spindle and amphiaster. The fate of DNA originally existing in the fused nucleus during achromosomal cleavage of fertilized sea urchin and starfish eggs induced by aphidicolin was determined using antidenatured DNA antibody. The nucleus is not formed in the divided daughter cells at the two‐cell stage but the nuclear‐envelope‐free chromatin mass which is unassociated with mitotic apparatus remains in the center region of embryos, especially near the first cleavage furrow. These results indicate that the condensed and nonreplicated chromatin can not be associated with spindle and asters in the presence of aphidicolin.
Effect of heparin on vascular smooth muscle cells. II. Specific protein synthesis
Authors: Cochran David L. , Castellot John J. , Karnovsky Morris J. 
Published: February 2005

AbstractHeparin suppresses the proliferation of vascular smooth muscle cells both in vivo and in vitro. The mechanism of action of the antiproliferative activity of heparin is not known. We have detected differences in the synthesis of specific proteins when vascular smooth muscle cells are exposed to heparin and report here that many characteristics of these protein alterations parallel the properties of the antiproliferative activity. The induction into the culture medium of a pair of proteins of approximately 35,000 dalton mw in heparintreated smooth muscle cell cultures and the antiproliferative effect of heparin share the following characteristics: 1) the effect is reversible, 2) the effect is specific for smooth muscle cells, 3) anticoagulant and non‐anticoagulant heparin are equally effective, 4) the effect is lost with time in culture and, 5) thermore, heparin causes a transient suppression of a 48,000 dalton substrateattached protein, whereas chondroitin sulfate A and C and dermatan sulfate had much less effect. Dextran sulfate was almost as effective as heparin in suppressing the synthesis of the substrate‐attached protein. These proteins appear to be noncollagenous and the induced synthesis of the 35,000 dalton proteins is inhibited by actinomycin D. Although a direct relationship between these specific protein changes and the antiproliferative effect of heparin has not been proven, these protein alterations may play a crucial role in the effect of heparin on smooth muscle cell growth.
Mitogen and co‐mitogen stimulation of lymphocytes inhibited by three Ca++ antagonists
Authors: Grier Charles E. , Mastro Andrea M. 
Published: February 2005

AbstractThe Ca++ requirement for in vitro lymphocyte stimulation by lectins is well known and can be demonstrated by the use of Ca++ chelators. In this study, three Ca++ antagonists were examined for their effects on lymphocyte proliferation. [3H]‐thymidine incorporation was employed to measure DNA synthesis in several systems. Stimulation and proliferation were achieved by the addition of one of the following: the mitogenic lectin concanavalin A (ConA); the combination of two co‐mitogens, the calcium ionophore A23187 and the phorbol ester, 12‐0‐tetradecanoylphorbol‐13‐acetate (TPA), neither of which is mitogenic alone; or the non‐mitogenic lectin, wheat germ agglutinin (WGA) with TPA. These mitogenic systems were tested for their sensitivity to the Ca++ channel blockers verapamil and nicardipine and the intracellular Ca++ antagonist TMB‐8. We found that the ConA and WGA plus TPA treated cells were inhibited approximately 50% by 10 μM verapamil, nicardipine or TMB‐8. The stimulation caused by A23187 and TPA was only inhibited by TMB‐8 and nicardipine. The inhibitory effects caused by the Ca++ antagonists could not be reversed by the addition of exogenous Ca++ (0.1–1.5 mM), but were reversed by repeated washings in antagonist free media. Using TMB‐8 we saw an apparent intracellular Ca++ dependence throughout the G1 phase. Previous studies using Ca++ chelators or Ca++ antagonists suggested an endpoint at about halfway through this period.
Regulation of thymidine kinase activity in the cell cycle by a labile protein
Authors: Coppock Donald L. , Pardee Arthur B. 
Published: February 2005

AbstractPrevious studies have shown that the onset of DNA synthesis in Balb/c 3T3 cells appears to be regulated by a labile protein. We have found that induction of thymidine kinase (TK) activity, after quiescent cells are stimulated by the addition of serum, is similarly regulated by a labile protein. Eight hours after serum stimulation, a 6‐h pulse of cycloheximide (CHM) caused an excess delay of 2h in TK induction. A similar delay also was found in the induction of thymidylate synthase (TS). In contrast, the bonzo(a)pyrene transformed 3T3 cell line, BP‐A31, which had previously been shown to have to excess delay for the onset of DNA synthesis also had no excess delay for the induction of TK activity after a pulse of CHM. The indudction of TK was inhibited by actinomycin D and dicholoribofuranosylbenzimidizole (DRB) suggesting a requirement for new RNA synthesis. It did not appear to depend on DNA synthesis as it was not blocked by aphidicolin. In conclusion, the induction of TK activity appears to be regulated by the same labile cellular signal as the onset of DNA synthesis, and to depend on an increase in the level of TK mRNA in late G1 or early S phase.
The specific phosphorylation of a 40S ribosomal protein in growth‐arrested tetrahymena is induced by sodium
Authors: Cuny Marguerite , Sripati Conjeevaram E. , Hayes Donal H. 
Published: February 2005

AbstractIn the past few years, in vivo phosphorylation of ribosomal proteins has been the subject of extensive studies and the results have shown that reversible phosphorylation of small subunit ribosomal protein S6, ubiquitous in eukaryotic cells, is apparently related to regulation of protein synthesis initiation. Thus the level of protein synthesis under various conditions is correlated with the level of S6 phosphorylation. In exponentially growing Tetrahymena, however, such phosphorylation does not occur, but when these cells are transferred to starvation buffers, the rate of protein synthesis is drastically reduced and a 40S ribosomal protein analogous to S6 of higher eukaryotic cells is fully and rapidly phosphorylated in all the ribosomes. We have studied the conditions which lead to this phosphorylation in growth‐arrested Tetrahymena, in order to understand the physiological significance of this process. Our results show that there is no obvious correlation between this phosphorylation and starvation. Moreover, it is not a developmentally regulated process related to the conjugation cycle, but a modification induced by the presence of sodium ions or high concentration of Tris in the starvation buffer. The physiological significance of this process is discussed in terms of accumulation of negative charge density probably required for initiation of protein synthesis in the growth‐arrested cells starving in Na+‐containing buffers.
Amphiphilic cationic peptides mediate cell adhesion to plastic surfaces
Authors: Rideout Darryl C. , Lambert Michael , Kendall Debra A. , Moe Gregory R. , Osterman David G. , Tao H. P. , Weinstein I. Bernard , Kaiser E. T. 
Published: February 2005

AbstractFour amphiphilic peptides, each with net charges of +2 or more at neutrality and molecular weights under 4 kilodaltons, were found to mediate the adhesion of normal rat kidney fibroblasts to polystyrene surfaces. Two of these peptides, a model for calcitonin (peptide 1, MCT) and melittin (peptide 2, MEL), form amphiphilic α‐helical structures at aqueous/nonpolar interfaces. The other two, a luteinizing hormone‐releasing hormone model (peptide 3, LHM) and a platelet factor model (peptide 4, MPF) form β‐strand structures in amphiphilic environments. Although it contains only 10 residues, LHM mediated adhesion to surfaces coated with solutions containing as little as 10 pmoles/ml of peptide. All four of these peptides were capable of forming monolayers at air‐buffer interfaces with collapse pressures greater than 20 dynes/cm. None of these four peptides contains the tetrapeptide sequence Arg‐Gly‐Asp‐Ser, which has been associated with fibronectin‐mediated cell adhesion. Ten polypeptides that also lacked the sequence Arg‐Gly‐Asp‐Ser but were nonamphiphilic and/or had net charges less than +2 at neutrality were all incapable of mediating cell adhesion (Pierschbacher and Ruoslahti, 1984). The morphologies of NRK cells spread on polystyrene coated with peptide LHM resemble the morphologies on fibronectin‐coated surfaces, whereas cells spread on surfaces coated with MCT or MEL exhibit strikingly different morphologies. The adhesiveness of MCT, MEL, LHM, and MPF implies that many amphiphilic cationic peptides could prove useful as well defined adhesive substrata for cell culture and for studies of the mechanism of cell adhesion.
Keratinocyte growth‐promoting activity from human placenta
Authors: O'Keefe E. J. , Payne R. E. , Russell N. 
Published: February 2005

AbstractExtracts of term human placenta were tested for enhancement of proliferative growth of primary cultures of human keratinocytes. Saline extracts or supernatants from homogenates were dialyzed extensively, lyophilized, and tested in subcultures of keratinocytes in MCDB 153 medium with 0.1 mM Ca++ containing only defined supplements (insulin, hydrocortisone, transferrin, ethanolamine, phosphoethanolamine). Cells plated in the absence of EGF at moderately high densities (1000–3000 cells per cm2) formed colonies and grew in the presence of placental extract at 25–500 μg/ml. Extracts of cord serum or maternal serum were inactive, suggesting that the activity is derived from placental tissue. The activity is not EGF, since the activity in the placental extract, unlike EGF, did not promote growth at low cell density, was synergistic with EGF under some conditions, and did not produce changes in colonial morphology which occurred in the presence of EGF. Unlike keratinocyte growth‐promoting activity in bovine hypothalamic extract, the activity is nondialyzable and is destroyed at 100°C. Placental extract could not replace any of the defined components of the medium and is therefore distinct from them. The presence of activity in the placenta with distinctive properties suggests that this is a previously undescribed material with growth‐promoting properties for epithelium.
Opposing effects of dexamethasone on the clonal growth of granulocyte and macrophage progenitor cells and on the phagocytic capability of mononuclear phagocytes at different stages of differentiation
Authors: Shezen Elias , Shirman Michael , Goldman Rachel 
Published: February 2005

AbstractDexamethasone, a synthetic glucocorticosteroid, was shown to modulate the colony‐stimulating factor‐dependent clonal growth of myeloid progenitor cells in semisolid agar cultures, enhancing the formation of granulocyte colonies (50–100%) and suppressing the formation of macrophage colonies (75–97%). Modulation of the pattern of myeloid colony formation by dexamethasone (12–125 nM) was brought about when the steroid was administered to 6‐day cultures at the time of culture initiation and up to 72 hr later. Dexamethasone inhibited myeloid cell proliferation when administered to 5‐day liquid cultures at culture initiation and up to 96 hr later. Dexamethasone (12–250 nM) also enhanced the phagocytic activity of bone marrow‐derived mononuclear phagocytes toward heat‐killed (HK) yeast cells (up to 100%) and IgG‐coated sheep red blood cells (up to 60%). Enhancement of the phagocytic capability depended critically on the stage in culture at which dexamethasone was administered. Exposure to dexamethasone for 28 hr up to 96 hr of 96‐hr cultures of bone marrow cells did not lead to a modulation of phagocytic activity of the developing mononuclear phagocytes. The presence of dexamethasone during the critical period of 96 hr to 120 hr after culture initiation led to an enhanced phagocytic capability, which was statistically significant already 12 hr after the administration of the glucocorticoid. Dexamethasone induced an enhanced phagocytic activity when administered at any time after culture initiation provided that it was in culture during this critical period. When added at 120 hr of culture, dexamethasone no longer enhanced the phagocytic capability of mononuclear phagocytes and when added later than 156 hr of culture suppressed it. Dexamethasone also suppressed (up to 68%) the phagocytic capability of resident and elicited peritoneal macrophages. The results suggest that glucocorticoids shift the balance of granulocyte vs. macrophage formation at early stages of precursor cell differentiation. Reduction in mononuclear phagocyte growth and enhancement of its phagocytic capability might reflect accelerated differentiation/maturation steps. The inhibitory effect of dexamethasone on macrophage formation and on the phagocytic capability of mature mononuclear phagocytes and peritoneal macrophages might be a relevant aspect of the in vivo immune suppression encountered after glucocorticoid administration.
DNA synthesis in rat hepatocytes: Inhibition by a platelet factor and stimulation by an endogenous factor
Authors: Hayashi Izumi , Carr Brian I. 
Published: February 2005

AbstractPrimary monolyer cultures of adult rat hepatocytes can be induced to undergo DNA synthesis in serum‐free medium in the presence of insulin, glucagon, and epidermal growth factor (three factors). We have found that hepatocyte DNA synthesis is affected not only by an endogenous stimulant produced by the hepatocytes and released into the culture medium. Serum has a strong inhibitory effect on hepatocyte DNA synthesis. Partially purified human platelet extract (“platelet inhibitor”) inhibits the three‐factor‐induced DNA synthesis in a concenration‐dependent manner. Pure βTGF at 0.5 ng/ml as well as HPLC‐purified PDGF at 10 ng/ml completely inhibit the three‐factor‐induced DNA synthesis. Determination of the time required for the presence of the three factors and the platelet inhibitor to exert their effects indicated that the inhibition of DNA synthesis is caused not by competition of the platelet inhibitor with any of the three factors but through an independent pathway. Hepatocyte DNA synthesis is density‐dependent and is greater if medium is not changed during the course of an experiment than if medium is changed daily. Hepatocyte‐conditioned medium is also affective in stimulating DNA synthesis beyond the level induced by the three factors. These results suggest that an endogenous stimulant for hepatocyte DNA synthesis is produced by the hepatocytes themselves. Our studies demonstrate that hepatocyte DNA synthesis is subject to both stimulatory and inhibitory controls. Unlike the three factors, the endogenous stimulant can overcome the inhibition by the platelet inhibitor, suggesting the importance of these factors in the physiological control of hepatocyte DNA synthesis.
Regulation of purine biosynthesis in G1 phase‐arrested mammalian cells
Authors: Emmett Kim , Patrick Joe , Aronow Bruce , Ullman Buddy 
Published: February 2005

AbstractThe effects of G1 phase growth arrest on purine biosynthesis were studied in cultured S49 T lymphoma cells. Incubations of wildtype S49 cells for 18 hr with dibutyryl cyclic AMP or forskolin, two agents which induced G1 arrest, reduced the rates of purine biosynthesis by 95%. Time course and concentration dependence studies indicated that the decrease in rates of purine biosynthesis correlated with the extent of G1 phase arrest. Similar studies with somatic cell mutants deficient in some component of cyclic AMP action or metabolism indicated that the depression in purine synthetic rates required G1 arrest and did not result from cell death. Rates of RNA and DNA synthesis were also markedly diminished in the growth arrested cells. Measurements of purine rates in the presence of azaserine indicated that the block in purine biosynthesis was prior to the formation of phosphoribosylformylglycinamide. Additionally, the activities of adenylosuccinate synthetase and IMP dehydrogenase were diminished in G1 arrested cells. The levels of all controlling enzymes, substrates, and cofactors, however, were not diminished in G1 arrested cells. Despite diminished rates of purine biosynthesis, the amounts of intracellular nucleotides in G1 cells were equivalent to those in exponentially growing cells. However, the concentrations of intracellular nucleotides were 30–50% higher in the growth arrested cells. These results suggested that perturbations in the consumption of nucleotides via inhibition of nucleic acid synthesis have profound effects on the purine pathway and indicated the importance of feedback inhibition by nucleotides in the regulation of purine synthesis in situ.