bluuebunny/crossref_metadata_2025_split huggingface dataset:
Induction and/or selective retention of proteins in mammalian cells exposed to cycloheximide
Authors: Sorrentino V. , Battistini A. , Curatola A. M. , Francesco P. Di , Rossi G. B. 
Published: February 2005

AbstractExposure of a number of murine and human cell lines to low graded doses of cycloheximide (CXM) results in a pattern of protein synthesis consisting of enhanced and induced species. These can be divided into two main classes according to molecular weight (20–40 and 70–120 Kd), similar to what has been described for other agents that modify the physiological conditions of growth. In addition, the pronounced synthesis of a hitherto unreported 50‐Kd protein species has been consistently observed in all lines tested. Simultaneous exposure of cells to CXM and actinomycin D results in suppressing synthesis of some but not all protein species observed, indicating that control mechanisms at both transcriptional and posttranscriptional levels may be operative in this system.
Incorporation of [3H]thymidine by isolated fetal myoblasts and fibroblasts in response to human placental lactogen (HPL): Possible mediation of HPL action by release of immunoreactive SM‐C
Authors: Hill D. J. , Crace C. J. , Milner R. D. G. 
Published: February 2005

AbstractWe investigated the actions of human placental lactogen (HPL) and human growth hormone (HGH) on [3H]thymidine incorporation and the release of immunoassyable somatomedin‐C (SM‐C) by isolated myoblasts, dermal fibroblasts, and costal cartilage explants taken from human fetuses, at 11–21 weeks of gestation. The incorporation of [3H] thymidine by myoblasts and fibroblasts was significantly increased after incubation for 20 hr or 44 hr, and cell number after incubation for 7 days, in the presence of 50–250 ng/ml HPL. Incubation with HPL did not increase [3H]thymidine incorporation into cartilage explants, whereas incubation with HGH failed to enhance the uptake of this isotope by any of the tissues. Following extraction with acid‐ethanol, culture medium conditioned by exposure to myoblasts or fibroblasts for 44 hr, and to cartilage explants for 7 days, contained radioimmunoassayable SMC. Myoblast‐conditioned medium contained significantly more SM‐C [1,609 ± 953 mU/mg cell protein (mean ± SD) n = 10] than did that conditioned by fibroblasts (637 ± 323; n = 5; P < 0.02). In 1 week of culture, cartilage explants released 4.1 ± 1.1 mU/mg wet weight (n = 7). The release of immunoassayable SM‐C from cultured cells was significantly increased in the presence of 250 ng/ml HPL in five of eight experiments with myoblasts and two of four experiments with fibroblasts. Neither fibroblasts or myoblasts showed increased SM‐C release following exposure to HGH.The results suggest that HPL, but not HGH, is growth‐promoting for some human fetal tissues in vitro and that this action is mediated, at least in part, by an increased release of somatomedins.
Serum factors that stimulate fatty acid oxidation: Properties of factors
Authors: Wice Burton M. , Stanisz Jolanta , Kennell David E. 
Published: February 2005

AbstractCultured heart muscle cells, but not HeLa cells, oxidize long‐chain fatty acids in medium containing dialyzed serum. Addition of chicken serum dialysate (or non‐dialized serum) stimulated palmitic acid oxidation by HeLa cells 10 to 20 fold. This serum activity was not eliminated by lipid extraction, ethanol or acid precipitation, alkaline phosphatase treatment, or autoclaving. About 80% was lost after any one of the following treatments: 6N HCl at 110°C for 16 hr, pepsin, Dowex cation exchange at pH 3, or 1N KOH at 100°C for 1 hr. Serum activity was separated into five or more peaks by gel filtration with Sephadex G‐10. Each of these peak fractions was further purified by HPLC using a cyanopropyl‐bonded resin. Carnitine, which is important for the transport of long‐chain fatty acids into mitochondria for oxidation, also stimulated the oxidation of palmitate. However, these serum factors are not known precursors to carnitine since its immediate precursor 4‐n‐trimethylaminobutyrate, did not stimulate palmitate oxidation. Total carnitine, including that in acylcarnitine compounds, was approxoimately 15 μM in the chicken sera to give ∼ 0.7 μM in the medium. Based on the fraction of total activity accountable by carnitine and fractional stability to acid, alkali, and pepsin, about 75% of the activity is from non‐carnitine compounds. Only one of the factors appears to be carnitine or an acylcarnitine derivative. Several lines of evidence suggest that the other factors are peptide compounds.
Cell kinetic study on GM colonies using autoradiography and collagen gel culture with special reference to unique proliferation of eosinophils
Authors: Izumi Tomoyuki , Suda Tetsuo , Shinonome Shougo , Maekawa Tadashi 
Published: February 2005

AbstractUsing a newly developed technique of autoradiography and collagen gel culture, a kinetic study on human GM colonies was attempted. Colonies of immature cells appeared first on day 5. The number of mixed colonies (mixture of immature cells, neutrophils, and/or monocyte‐macrophages) and neutrophil colonies attained a maximum on days 8 to 10 and a broad peak of monocyte‐macrophage colonies was observed on days 11 to 16. Eosinophil colonies appeared first on day 12, reached a maximum on day 18, and then gradually decreased. A detailed analysis of the order of appearance of the colonies suggests that mixed, neutrophil and monocyte‐macrophage colonies originate from immature cell colonies or clusters, while eosinophil colonies do not. An autoradiographic study was designed to study the proliferation characteristics of each colony. Labeling indices (LI) with 3H‐TdR of the cells in immature cell colonies were always high. LI of the cells in differentiated colonies such as neutrophil, monocyte‐macrophage, and mixed colonies were low throughout the observation period. In contrast, LI of the cells in eosinophil colonies were constantly high regardless of the size of cell aggregates and the duration of the culture period. Both mitotic indices and mean grain counts on the nuclei of eosinophils were similar to those of immature cells. These results suggest that eosinophil colonies develop from their own small clusters and that eosinophils retain a fairly good proliferative capacity even when differentiated to the level in which specific granules appear in the cytoplasm.
A direct correlation between hyperthermia‐induced membrane blebbing and survival in synchronous G1 CHO cells
Authors: Borrelli M. J. , Wong R. S. L. , Dewey W. C. 
Published: February 2005

AbstractHeating synchronous G1 cells at 45.5°C for 3–20 min induced varying degrees of membrane blebbing ranging from nonblebbed cells indistinguishable from control cells to those with blebs larger than the cell itself. Both the proportion of cells exhibiting blebbing and the mean diameter of the blebs increased with heating duration. Scoring individual cells for both blebbing and colony formation demonstrated that cells with blebs larger than 50% of the cell diameter did not survive to form colonies. Electron microscopy showed that all subcellular organelles, save the ribosomes, were absent from the membrane blebs. Freeze fracture replicas revealed no changes in membrane ultra‐structure, except on some 15% of the blebs that contained bald patches devoid of membrane particles.
Substratum influence on collagen and fibronectin biosynthesis by arterial smooth muscle cells in vitro
Authors: Holderbaum Daniel , Ehrhart L. Allen 
Published: February 2005

AbstractCollagen, fibronectin, and nonfibrous protein biosynthesis were examined in cultures of rabbit arterial smooth muscle cells grown on tissue culture plastic precoated either with rabbit plasma fibronectin or bovine serum albumin. Cells seeded into fibronectin‐coated wells appeared to reach confluence more quickly than counterparts grown on albumin‐coated surfaces. Measurement of 3H‐thymidine incorporation into DNA by these cultures suggested that this was probably a consequence of more rapid and efficient cell attachment rather than an increased rate of proliferation of smooth muscle cells grown on fibronectin. In preconfluent cultures, the rates of collagen and fibronectin biosynthesis were reduced to 34 and 57%, respectively, on a per‐cell basis in cultures grown on fibronectin‐coated surfaces compared with cells grown on albumin‐coated plasticware. In preconfluent cultures grown on fibronectin‐coated surfaces, a greater percentage of the total fibronectin synthesized was incorporated into the cell layer. The distribution of newly synthesized collagen between culture medium and cell layer, however, was not affected by alteration of substratum composition. There was no difference in the rate of synthesis of noncollagen proteins between the two groups of preconfluent cells. In postconfluent cultures the rates of collagen and fibronectin biosynthesis were equivalent in both albumin‐ and fibronectin‐treated cultureware. In preconfluent cultures, analyses of procollagens showed that the overall amounts of both types I and III procollagens were reduced in fibronectin‐treated wells, indicating the reduction in collagen synthesis to be general and not type‐specific. Although type V procollagen biosynthesis was not detected in either preconfluent group, it was found in postconfluent cultures. The reduction of fibronectin synthesis in cells grown in fibronectin‐coated wells was significant as early as 4 hours after plating. Together, these findings suggest that cultured arterial smooth muscle cells are capable of deriving information from their substratum and regulating the biosynthetic rates of extracellular matrix components in response to the immediate needs of the cell.
Nonhaem complexes of FeIII stimulate cell attachment and growth by a mechanism different from that of serum, 2‐oxocarboxylates, and haemproteins
Authors: Kay G. F. , Ellem K. A. O. 
Published: February 2005

AbstractMost cell lines, even those producing their own growth factors, need a serum supplement when growing in several commonly used media. The requirement for serum to sustain attachment and growth in RPMI 1640 and MEM has been found to be met by a range of 2‐oxocarboxylates, by diverse coordination complexes of Felll, and by a variety of haem‐containing proteins including catalase. The latter directly implicates H2O2 in the serum shift‐down effects. H2O2 was found to accumulate in low serum media under normal laboratory lighting conditions to levels that were shown to be sufficient, when added to freshly prepared media, to explain the depressed cell performance. With the exception of some of the nonhaem Felll coordination complexes, substances found to stimulate cell attachment and growth were capable of scavenging H2O2. This suggests that an important function of serum and the 2‐oxocarboxylates (α‐keto acids) frequently used as “nonessential” medium additives is to remove H2O2 produced photodynamically during the storage and manipulation of media containing a high content of riboflavin. However, the nonhaem Felll complexes with saturated coordination shells, although capable of reducing photodynamic generation of H2O2 to a greater or lesser extent, have their prime effect by an unknown, intriguing mechanism, probably based on a common redox function.
Regulation of the A system of amino acid transport in Chinese hamster ovary cells, CHO‐K1: The difference in specificity between the apo‐repressor inactivator (apo‐ri) and the transporter and the characterization of the proposed apo‐ri
Authors: Moffett John , Englesberg Ellis 
Published: February 2005

AbstractWhen amino acids that are generally transported through the A system are added to derepressed cultures of CHO‐K1 cells or to cultures that are undergoing starvation‐derepression, as in the co‐repressor (co‐r), co‐inactivator (co‐i), (co‐ri) assay, the A system undergoes trans‐inhibition, inactivation, and repression. The effect of inactivation and repression is not related to the ability of amino acids to bind to the A system transporter but supports a model in which these amino acids act as co‐r's/co‐i's, and by binding to a aporepressor/inactivator (apo‐ri), the product of gene R1, convert it into a repressor/inactivator (ri). For example, β‐alanine acts as a strong co‐r but does not inhibit proline transport through the A system. Hydroxyproline and histidine, although poor inhibitors of proline transport, are very effective as co‐ri's. Diaminobutyrate, phenylalanine α‐keto glutarate, pyro‐glutamate, isoleucine, and valine, compounds that inhibit A system transport, listed in decreasing order of effectiveness, are all equally poor as co‐ri's. Also the Km for the transport of 2‐(methylamino)isobutyric acid (MeAIB) through the A system is two times the concentration of MeAIB required to produce one‐half inactivation. Amino acid effectors and mutation can modify the conversion of the apo‐ri to repressor (r) and inactivator (i). The apo‐ri is converted by alanine, serine, proline, and MeAIB to ri, by β‐alanine and tryptophane to r, and by hydroxyproline to r and reduced i. The full constitutive and partial constitutive mutants alar4 and alar2, respectively, are in the same complementation group. Alar4 has no active apo‐ri while the rate of derepression of alar2 is twice and the inactivation rate is equal to that of the parent culture.
DNA‐mediated transfer of cAMP resistance in CHO cells
Authors: Abraham Irene , Brill Steven , Chapman Margaret , Hyde Jennifer , Gottesman Michael 
Published: February 2005

AbstractChinese hamster ovary (CHO) strain 10215 carries a dominant mutation which confers resistant to cAMP by virtue of an altered catalytic subunit of the cAMP‐dependent protein kinase (Evain et al., 1979). This mutation was transferred to wild‐type CHO cells by DNA‐mediated gene transfer. Based on the absence of cAMP growth inhibition, seven transformant colonies were isolated. One of these, 11586, was studied in detail. This transformant showed the same phenotype as the mutant, including resistance to the morphological changes and growth inhibitory effects of 1 mM 8‐Br‐cAMP, reduced total cAMP dependent protein kinase activity and lowered sensitivity of the kinase to cAMP activation. When the cAMP‐dependent protein kinase was fractionated on a DEAE‐cellulose column, the transformant was lacking in type II cAMP dependent protein activity, to the same degree as the mutant. The transformant and mutant, but not wild‐type cells, also failed to phosphorylate a 52,000‐dalton protein in a cAMP‐dependent manner. These characteristics support the conclusion that the gene for the mutant cAMP‐dependent protein kinase has been transferred. The ability to transfer this gene by DNA‐mediated transfer suggests that this methodology may be useful for the molecular isolation of the gene encoding the catalytic subunit of cAMP‐dependent protein kinase.
Multilineage, non‐species specific hematopoietic growth factor(s) elaborated by a feline fibroblast cell line: Enhancement by virus infection
Authors: Abkowitz Janis L. , Holly Richard D. , Segal Gerald M. , Adamson John W. 
Published: February 2005

AbstractIn studies designed to determine the role of feline leukemia virus (FeLV) in the pathogenesis of marrow failure in the cat, we tested medium conditioned by uninfected and FeLV‐infected feline embryonic fibroblasts (FEA) for its effect on hematopoietic colony growth in culture. As opposed to an inhibitory effect, we found that the conditioned medium (CM) from FEA or FEA/FeLV increased the in vitro growth of multiple hematopoietic progenitor cell types including erythroid burst‐forming cells (BFU‐E), granulocyte/macrophage colony‐forming cells, megakaryocytic colony‐forming cells, and mixed‐cell colony‐forming cells. Furthermore, CM enhanced the growth of progenitors in cultures of mouse or human marrow cells, as well as cat marrow cells. Stimulation of feline BFU‐E was most marked with an increment in growth of 400% over control. The human burst promoting activity (BPA) of the CM was equivalent or better than other CM available in our laboratory. The evidence suggests that the growth‐promoting activity is a constitutive product(s) released by FEA which was enhanced eightfold with virus infection. Studies with non‐adherent and T‐lymphocyte‐depleted human marrow cells and human peripheral blood cells suggest that the growth factor(s) acts directly on progenitor cells and not through readily identified accessory cells. These findings are consistent with the concept that mesenchymal cells such as fibroblasts have the capacity to release hematopoietic growth factor(s) capable of acting on primitive hematopoietic progenitors. The results provide an example of how injury of such cells, through virus infection, may enhance growth factor(s) release and influence the hematopoietic microenvironment.
Selection and characterization of mutant S49 T‐lymphoma cell lines resistant to phosphonoformic acid: Evidence for inhibition of ribonucleotide reductase
Authors: Albert Daniel A. , Gudas Lorraine J. 
Published: February 2005

AbstractPhosphonoformic acid (PFA) and its congener phosphonoacetic acid (PAA) are inhibitors of viral replication whose mechanism of action appears to be the inhibition of viral DNA polymerase. These drugs inhibit mammalian DNA polymerase to a lesser extent. We sought to characterize the effects of phosphonoformic acid on mammalian cells by examining mutants of S49 cells (a mouse T‐lymphoma line), which were selected by virtue of their resistance to phosphonoformic acid. The 11 mutant lines that were resistant to growth inhibition by 3 mM PFA had a range of growth rates, cell cycle distribution abnormalities, and resistance to the inhibitory effects of thymidine, acycloguanosine (acyclovir), aphidicolin, deoxyadenosine, and novobiocin. Most mutant lines had pools of ribonucleoside triphosphates and deoxyribonucleoside triphosphates similar to those of wild‐type S49 cells. However, one line (PFA 3–9) had a greatly elevated dCTP pool. When this mutant line was further characterized, no apparent defect in DNA polymerase α activity was seen, but an increased ribonucleotide reductase activity, as assayed by CDP reduction in permeabilized cells, was observed. The CDP reductase activity in the PFA 3–9 cells decreased to wild‐type control levels, and the CDP reductase activity of wild‐type cells was also greatly reduced when PFA (2–3 mM) was added to permeabilized cells during the enzyme assay. These results demonstrate that PFA can directly inhibit ribonucleotide reductase activity in permeabilized cells. In addition, when PFA was added to exponentially growing cultures of either wild‐type or PFA 3–9 mutant cells, the drug caused an arrest in S phase of the cell cycle and a decrease in all four deoxyribonucleotide pools, with the most dramatic decrease in the dCTP pools. The reduction in the dCTP pool level could be reversed by addition of exogenous deoxycytidine, but this reversed PFA toxicity only marginally. These observations suggest that PFA is an inhibitor of mammalian ribonucleotide reductase and that partial resistance to PFA can be effected by mutation to increased CDP reductase activity resulting in a large dCTP pool. This mutation results in less than twofold resistance to PFA, suggesting that other sites of inhibition coexist.
Transient stimulation of glucose metabolism by insulin in the 1‐day chick embryo
Authors: Baroffio Anne , Raddatz Eric , Markert Michèle , Kučera Pavel 
Published: February 2005

AbstractEffects of insulin upon glucose metabolism were investigated in chick embryos explanted in vitro during the first 30 h of incubation. Insulin stimulated the glucose consumption of the chick gastrula (18 h) and neurula (24 h), but had no effect on the late blastula (0 h:laying) and on the stage of six to eight somites (30 h). The increase in glucose consumption concerned both the embryonic area pellucida (AP) and extraembryonic area opaca (AO). AP responded to a greater extent (50%) and at a lower range of concentrations (0.1–1.0 ng/ml) than AO (30%; 1–100 ng/ml). Insulin had no effect on the oxygen consumption of blastoderms, whereas it stimulated the aerobic lactate production (approximately 70% of the additional glucose consumption was converted to lactate).The nanomolar range of stimulating concentrations suggests that insulin has a specific effect in the chick embryo, and that it could modulate glucose metabolism in ovo as well. The transient sensitivity of the embryo to insulin is discussed in relation to behavior of mesodermal cells.
Retinoic acid effect on cyclic AMP‐dependent protein kinases in embryonal carcinoma cells: Studies with differentiation‐defective sublines
Authors: Plet Ariane , Gerbaud Pascale , Sherman Michael I. , Anderson Wayne B. , Brion Daniele Evain 
Published: February 2005

AbstractRetinoic acid induces the differentiation of PCC4.aza 1R and Nulli‐SCC1 embryonal carcinoma (EC) cells. In response to retinoic acid treatment, the levels of cyclic AMP (cAMP)‐dependent protein kinases are enhanced in the plasma membrane within 17 hours and in the cytosol fractions of these cells within 2 to 3 days, as determined by phosphotransferase activity and by 8‐azido‐cyclic [32P]AMP binding to the RI and RII regulatory subunits. PCC4 (RA)−1 and Nulli (RA)−1 are mutant EC lines that fail to differentiate in response to retinoic acid. The former line, but not the latter, lacks cellular retinoic acid‐binding protein (cRABP). Basal levels of cAMP‐dependent protein kinase activities are elevated in PCC4 (RA)−1 cells. When these cells are treated with retinoic acid, neither cAMP‐dependent protein kinase activities nor cAMP binding activities are enhanced; rather, there is a decrease in cytosolic kinase activity and RI subunit. On the other hand, Nulli (RA)−1 cells exhibit increases both in cAMP‐dependent protein kinase activities and cAMP binding in response to retinoic acid. These results raise the possibility that cRABP mediates the enhancement of regulatory and catalytic subunits of cAMP‐dependent protein kinases in both the membrane and the cytosolic fractions of the teratocarcinoma cells. There also might be some effects of retinoic acid on the cAMP‐dependent protein kinase that are unrelated to differentiation and to the presence of cRABP.
Collagen synthesis by murine bone marrow cell culture
Authors: Waterhouse Elizabeth J. , Quesenberry Peter J. , Balian Gary 
Published: February 2005

AbstractCollagen types synthesized by murine bone marrow cells were studied and the effect of lithium chloride on collagen biosynthesis in vitro was investigated. In the liquid culture system used, an adherent, mixed cell population supports hemopoiesis. Radioactive labeling of cell cultures and subsequent fractionation with ammonium sulfate, enzyme digestion, immune precipitation, and gel electrophoresis indicated that the bone marrow cells synthesized precursors to collagen types I, III, and IV, and fibronectin. A previously undescribed molecule or fragment with an apparent molecular weight of 17,000 daltons that was susceptible to bacterial collagenase and containing no interchain disulfide bonds was also identified in the culture media of both control and lithium‐treated cells. Lithium treatment did not affect the types of collagen synthesized, although the relative proportions of collagen types may differ from controls. However, lithium does have an effect on the appearance of some, as yet unidentified, non‐collagenous components in the cell culture media.
Characterization of RNA polymerase type II from human term placenta
Authors: Freund Erwin , McGuire Peter M. 
Published: February 2005

AbstractRNA polymerase type II from human term placenta has been isolated and characterized with respect to its template, ammonium sulfate, divalent cation, and buffer preferences. In addition, the apparent Michaelis constants for AMP amd UMP incorporation have been determined. The enzyme was also analyzed by native and denaturing polyacrylamide gel electrophoresis, and evidence is presented that a single polypeptide is radiolabeled with azido purine nucleoside triphosphate photoprobes.