bluuebunny/crossref_metadata_2025_split huggingface dataset:
A study of binding‐solubilization of some glycolytic enzymes in striated muscle in situ
Authors: Ross Robert E. , Hultin Herbert O. 
Published: February 2005

AbstractThe solubilization of lactate dehydrogenase (LDH), glyceralde‐hyde‐3‐phosphate dehydrogenase (GAPDH), and alpha‐glycerophosphate dehydrogenase (aGPDH) was studied in pressed muscle as a function of ionic strength and NADH concentration.The results indicate that these factors affect the binding‐solubilization of LDH and GAPDH in a similar way to their effect in dilute homogenized tissue. Alpha‐glycerophosphate dehydrogenase was included as a typical soluble enzyme, since we have been unable to demonstrate its binding to subcellular fractions under any conditions. As with homogenized tissue, LDH was less susceptible to solubilization by ionic strength than GAPDH. It was demonstrated that LDH isozymes richer in heart‐type subunits were more easily removed from muscle by centrifugation‐imbibition than isozymes richer in the muscle‐type subunits. This was interpreted as indicating that binding of the enzyme to subcellular structures was a major factor in the restricted removal of these enzymes from muscle, since only the muscle‐type subunit is capable of binding to subcellular particles.It was further demonstrated that LDH could be taken up into muscle tissue, depleted in the enzyme, against an apparent concentration gradient. This was also interpreted as binding of the enzyme to the particulate structure of the muscle. Furthermore, this uptake of LDH occurred under conditions of ionic strength (0.25) and pH (7.5) that would prevent binding of the enzyme to the particulate fraction of a dilute suspension of homogenized muscle tissue. Thus, physiological conditions of pH and ionic strength do not necessarily induce solubilization of chicken breast muscle LDH in situ. Data obtained with dilute tissue homogenates, therefore, may not necessarily be easily and safely extrapolated to conditions in situ.
Measurement of intracellular pH in sea urchin eggs by 31P NMR
Authors: Inoue Hiroko , Yoshioka Tohru 
Published: February 2005

AbstractThe intracellular pH (pHi) of sea urchin eggs was measured using 31P nuclear magnetic resonance (NMR) spectroscopy. The mean value of pHi of unfertilized eggs was about 6.2 (H. pulcherrimus and A. crassispina) to 6.3 (P. depressus). In contrast to results of other studies, pHi of sea urchin eggs was not changed after fertilization. During exposure of the eggs to NH4Cl or procaine‐containing natural sea water (NSW), however, pHi rose about 0.4‐0.8 pH units; the pHi fell to its initial value upon washing the eggs with weak basefree NSW. These changes of pHi by weak base treatment agreed well with the data obtained by other workers. In order to understand the discrepancy of pHi changes in fertilized eggs between NMR data and other measuring procedures, we measured acid production and O2 uptake, so that CO2 accumulation and proton release did not result in alkalinization. The invariance of the fertilized eggs under anaerobic conditions; otherwise NMR showed a different answer from other measuring procedures, because of its particular characteristics such as non‐destructivity and compartmentation of pH.
Regulation of metallothionein synthesis in HeLa cells by heavy metals and glucocorticoids
Authors: Karin Michael , Slater Emily P. , Herschman Harvey R. 
Published: February 2005

AbstractMetallothioneins (MTs) are low molecular weight, cysteine‐rich proteins that bind heavy metals. MT induction occurs in liver in response to either heavy metal (Zn++ or Cd++) administration or stress. The synthesis of MT can also be induced by either heavy metals or glucocorticoid hormones in HeLa cells cultured in serum‐free medium. Induction of MT by zinc is subject to “desensitization.” In contrast, dexamethasone (dex) induction results in a continued elevation in the rate of MT synthesis. The stability of MT is dependent on the availability of metal; consequently, MT induced by dex is degraded much more rapidly (half‐life of 11 to 12 hours) than MT induced by elevated zinc levels (half‐life of 36 to 38 hours). Removal of either inducer results in biphasic degradation curves, as apothionein and zinc come into balance. In contrast, deinduction kinetics for MT synthesis following removal of the two inducers (zinc and dex) are the same, with a half‐life of two and one‐half hours. Inhibition of RNA synthesis blocks deinduction after removal of inducer. Induction of MT occurs in a wide variety of species, from blue‐green algae to man. This system should provide an excellent model for the comparative biochemistry of regulation of gene expression.
External cell surface protein phosphorylation in normal and Rous sarcoma virus transformed chick embryo fibroblasts
Authors: Yee Siu‐Pok , Branton Philip E. 
Published: February 2005

AbstractNormal and Rous sarcoma virus (RSV)‐transformed chick embryo fibroblasts growing on plastic dishes were incubated with ATP (γ32P) in situ to detect external cell surface protein kinase activity. Under the conditions employed, 32P was incorporated exclusively into proteins, specifically those at the external cell surface, as radioactivity was removed by tryspin treatment of labeled whole cells. In addition, exogenous histones were phosphorylated when added to the reaction mixture. Cyclic nucleotides had virtually no effect on 32P incorporation, suggesting that little or no cyclic nucleotide‐dependent protein kinase activity was present at the external cell surface. Cell surface protein kinase activity was higher in transformed than in normal cells, and, using a temperature‐sensitive RSV src mutant, this difference was shown to be transformation‐specific. Several differences were observed in the cell surface proteins phosphoryllated in normal and transformed cells and at least two of these were transformation‐specific. These data suggest that changes in external cell surface protein physphorylation are associated with RSV transformation and thus could play a role in the formation of the transformed cell phenotype.
Enhancement of melanotic expression in cultured mouse smelanoma cells by retinoids
Authors: Lotan Reuben , Lotan Dafna 
Published: February 2005

AbstractRetinoic acid (RA), which reduces the rate of cell proliferation in S91 mouse melanoma clone C2 cells, was found to stimulate the expression of their melanotic phenotype. RA treatment also induced the extension of long cellular processes. The RA effects on melanogenesis included stimulation of tyrosinase activity and augmentation of cellular melanin content to levels 3‐ to 4‐fold higher than in untreated cultures at similar cell densities. These effects became apparent after 48 hours of exposure to 10−5 M RA and increased thereafter. Halfmaximal stimulation in cells treated for 6 days occurred at 5 × 10−7 M RA. Although the degrees of melanogenesis enhancement by RA (10−5 M) and by α‐melanocyte stimulatory hormone (2 × 10−7 M) were similar, the former did not alter the intracellular cAMP level, whereas the latter induced a transient 4‐fold increase. In high‐passage (p28) cells, as well as in low‐passage cells (< p10) treated with tyrosinase inhibitor phenylthiocarbamate, melanin synthesis was suppressed in the absence and presence of RA, yet the ability of RA to inhibit cell proliferation was not compromised. In the presence of the tumor promotor phorbol myristate acetate (> 5 × 10−9 M) melanin synthesis in control as well as in cells exposed to RA was dramatically inhibited. Phorbol which is not active in tumor promotion had no effect on melanogenesis. In addition to RA, other retinoids, such as 13‐cis‐retinoic acid, retinyl acetate, the TMMP analog of RA and the phenyl analog of RA, but not the pyridyl analog of RA or retinyl palmitate, also inhibited cell growth and enhanced melanin synthesis.
Effects of parathyroid hormone and cyclic AMP analogues on the activity of ornithine decarboxylase and expression of the differentiated phenotype of chondrocytes in culture
Authors: Takigawa M. , Takano T. , Suzuki F. 
Published: February 2005

AbstractParathyroid hormone (PTH) greatly increased the level of adenosine 3′,5′ cyclic monophosphate (cAMP) in rabbit costal chondrocytes in culture 2 minutes after its addition. PTH, as well as N6 O2' dibutyryl adenosine 3′,5′ cyclic monophosphate (DBcAMP) and 8 Bromo adenosine 3′,5′ cyclic monophosphate (8 Br‐cAMP) induced ornithine decarboxylase (ODC; L‐ornithine carboxylyase; EC 4.1.1.17), which reached a maximum 4 hours after their addition. Neither cAMP, N6 O2′ dibutyryl guanosine 3′,5′ cyclic monophosphate (DBcGMP), nor sodium butyrate increased the activity of the enzyme. PTH had no effect on DNA synthesis, while DBcAMP and 8 Br‐cAMP decreased DNA synthesis. Expression of the differentiated phenotype of chondrocytes in culture was also induced by PTH, DBcAMP, and 8 Br‐cAMP, but not by cAMP, DBcGMP, or sodium butyrate, as judged by morphological change. Glycosaminoglycan synthesis, a characteristic of the cartilage phenotype, began to increase 8 hours after addition of PTH or DBcAMP, reaching a plateau 32 hours after their addition.These findings suggest that PTH induces increase of ODC activity and expression of the differentiated phenotype of chondrocytes through increase of cAMP and that induction of ODC is closely related to expression of the differentiated phenotype of chondrocytes.
Somatomedin‐C and platelet‐derived growth factor stimulate human fibroblast replication
Authors: Clemmons David R. , Van Wyk Judson J. 
Published: February 2005

AbstractPrevious studies have demonstrated that serum contains mitogens, such as platelet‐derived growth factor (PDGF), which may alter fibroblast responsiveness to growth factors contained in plasma. Somatomedin‐C (SM‐C) has been identified as one of the plasma growth factors required for mouse Balb/c 3T3 fibroblasts to initiate DNA synthesis. The present experiments were undertaken to explore the interaction between PDGF, human growth hormone (hGH), SM‐C, and other growth‐promoting agents in stimulating the growth of human fibroblasts. Proliferation of human dermal fibroblasts plated at low density (3,000 cells/cm2) was found to be equally stimulated by continuous exposure to either normal or somatomedin‐C‐deficient serum. In contrast, when confluent monolayers were sequentially exposed to PDGF, followed either by normal platelet poor plasma (PPP) or hypopituitary PPP, the cells exposed to normal PPP entered the “S” phase of the cell cycle 50% faster. This difference could be abolished by a 6‐hour incubation with growth hormone (10 ng/ml) or somatomedin‐C (5 ng/ml) preceding the addition of plasma. When medium containing either hGH or Sm‐C was changed frequently so as to remove factors secreted by fibroblasts, only those cells exposed to exogenous somatomedin‐C entered DNA synthesis. This finding is in agreement with previous findings that human fibroblasts are capable of making Sm‐C in response to hGH. These findings support the hypothesis that somatomedin is required for fibroblast replication in vitro, and that growth hormone appears to stimulate replication indirectly through somatomedin production.
Gluconeogenesis in rat liver parenchymal cells in primary culture: Permissive effect of the glucocorticoids on glucagon stimulation of gluconeogenesis
Authors: Stumpo Deborah J. , Kletzien Rolf F. 
Published: February 2005

AbstractPrimary cultures of parenchymal cells isolated from adult rat liver by a collagenase perfusion procedure and maintained as a monolayer in a serum‐free culture medium were used to study glucoeogenesis and the role that the glucocorticoids play in the control of this pathway. These cells carried out gluconeogenesis from three‐carbon precursors (alanine and lactate) in response to glucagon and dexamethasone added alone or in combination. Maximum glucose production was observed with cells pretreated for several hours with dexamethasone and glucagon prior to addition of substrate and glucagon (8‐ to 12‐fold increase over basal glucose production). Half‐maximum stimulation of gluconeogenesis was seen with 3.6 × 10−10 M glucagon and 3.6 × 10−8 M dexamethasone. Maximum stimulation was oberved with 10−7 M glucagon and 10−6 M dexamethasone. The length of time of dexamethasone pretreatment was found to be important in demonstrating the effect of glucocorticoids on glucagon‐stimulated gluconeogenesis. Treeatment of cells with dexamethasone for 2 hours did not result in an increase in glucose production over identical experimental conditions in the absence of dexamethasone, wherease pretreatment for 5 hours (1.2‐fold increase) or 15 hours (1.7‐fold increase) did result in an increase in glucose production. The results establish that the adult rat liver parenchymal cells in primary culture are a valid model system to study hepatic gluconeogenesis. In addition, we have established directly that the glucocorticoids amplify the glucagon stimulation of gluconeogenesis.
Proliferative characteristics of clonal endothelial cell strains
Authors: Rosen Eliot M. , Mueller Stephen N. , Noveral James P. , Levine Elliot M. 
Published: February 2005

AbstractWe have utilized clonal strains of bovine fetal aortic endothelial cells to study cellular senescence in a differentiated cell type of physiological significance. Serial subcultivation of nine endothelial clones derived from three fetal calf aortas revealed proliferative life‐spans in vitro of 53–125 population doublings (PDs), compared with 60 and 143 PDs for two lines of bovine fetal lung cells and 85 and 147 PDs for two lines of bovine vascular smooth muscle cells. Serial growth curves showed marked reductions associated with endothelial cellular senescence both in cellular growth rate and culture plateau density. Studies of the 24‐hour [3H]‐thymidine labeling index versus percentage of proliferative life‐span completed indicated that clonal endothelial cultures contained a large proportion (greater than 90%) of rapidly cycling cells until about 75% of the life‐spans were completed. Senescent endothelial cells showed evidence of large increases in cell area, cell volume, and protein content. In those clones examined, one specialized endothelial function, Factor VIII antigen expression, was retained qualitatively throughout the life‐spans.
Cellular control of ornithine decarboxylase activity by its antizyme
Authors: Heller John S. , Canellakis Evangelos S. 
Published: February 2005

AbstractConditions have been established under which the antizyme of ornithine decarboxylase (E.C. 4.1.1.17, L‐ornithine carboxy‐lyase, ODC) a noncompetitive protein inhibitor of ODC, can be detected in cells in response to as little as 10−7 M putrescine. The maintenance of intracellular antizyme activity depends upon the continued presence of putrescine in the medium. Removal of putrescine results in a rapid decline of antizyme activity. These phenomena are unaffected by the presence of cycloheximide and are comparable to the requirement of L‐asparagine for the maintenance of ODC activity.The extent to which the antizyme level is increased is inversely related to the preexisting level of intracellular ODC at the time of addition of putrescine. The time of appearance of free antizyme is delayed in cells that have high levels of ODC; the amount of free antizyme that can be assayed for in these cells, at any particular time is correspondingly less. The converse is also true. In cells that have high levels of antizyme, the delay in appearance of ODC is greater and the amount of ODC that can be assayed for is correspondingly less than in cells with low levels of antizyme.These experiments, as well as others, indicate that the ODC antizyme and ODC interact in vitro with each other to modify their respective activities.
Patients with myotonic dystrophy, a possible segmental progeroid syndrome, and duchenne muscular dystrophy have fibroblasts with normal limits for in vitro lifespan and growth characteristics
Authors: Wertz Richard L. , Hartwig Geoffrey B. , Frost Arlo P. , Brophy Jere J. , Atwater Susan K. , Roses Allen D. 
Published: February 2005

AbstractMyotonic dystrophy (MyD) has been suggested to be a segmental progeroid syndrome in man, as this syndrome has some clinical manifestations of premature aging. Fibroblasts from patients with other progeroid syndromes have been shown to have diminished in vitro lifespans or growth characteristics; therefore, it was of interest to study cellular senescence in fibroblasts from patients with MyD. Fibroblast cultures from patients with Duchenne muscular dystrophy (DMD) were used as additional controls, as premature aging is not associated with this genetic disorder. Primary skin fibroblast cultures obtained from patients with MyD or DMD and from age‐sex matched controls were grown in DMEM plus 10% FBS. The in vitro lifespan was determined by either a 1:4 split ratio or with a constant initial inoculum of 1 × 104 cells/cm2, followed by determination of the final density at weekly intervals. Our results demonstrate that there is no difference in the limits of the in vitro lifespan for either the MyD or DMD fibroblast strains compared to the controls. Likewise, no difference could be detected in the growth characteristics of these cells. The only observable difference was that the pooled age‐matched controls and MyD cultures had a shorter in vitro lifespan than the DMD group and their pooled controls, a finding expected because of the age of the patients in each group. Unlike the other progeroid syndromes, MyD fibroblasts have normal limits for in vitro lifespan. MyD is probably not closely related to the other premature aging syndromes, although there is an increasing phenotypic expression as a function of age.
The tumor promoter 12‐0‐tetradecanoylphorbol‐13‐acetate induces ornithine decarboxylase in proliferating basal cells but not in differentiating cells from mouse epidermis
Authors: Lichti Ulrike , Patterson Elroy , Hennings Henry , Yuspa Stuart H. 
Published: February 2005

AbstractThe cultivation of mouse epidermal cells in medium of reduced calcium concentration (0.02–0.1 mM) selects for basal cell growth. Elevation of medium calcium levels above 0.1 mM results in rapid and well defined differentiative changes. This model was utilized to determine which cell type in mouse epidermis responds to the phorbol ester tumor promoter, 12‐0‐tetradecanoyl‐phorbol‐13‐acetate (TPA), by an induction of the enzyme ornithine decarboxylase (ODC). Previous data had shown that TPA induces ODC in primary mouse epidermal cells only during the first 36 hr after plating in medium containing 1.44 mM Ca2+. In contrast, the induction in cells grown in low calcium medium was 2–10‐fold greater, and inducibility persisted for at least 4 weeks. The greater inducibility of ODC in low calcium cells is not paralleled by increased thymidine incorporation after TPA treatment, probably because these cells are already proliferating at a maximum rate. When low calcium cells grown in 0.07 mM Ca2+ medium were switched to 1.2 mM Ca2+, there was a rapid loss of ODC inducibility. These results strongly suggest that the basal cells of the epidermis constitute the major target cells for the induction of ODC by TPA. The induction of ODC by ultraviolet light was not enhanced by growth of cells in low calcium medium, indicating that extracellular calcium concentration per se does not determine ODC inducibility. When epidermal cells grown in 1.2 mM or 0.07 mM Ca2+ medium were exposed to both UV light and TPA, there was a significant synergistic effect of combined treatment over the sum of each individual response, suggesting that factors in addition to differentiation determine the extent of ODC induction.
Demonstration of a hematopoietic stem cell antigen (SC‐1) on a murine lymphoma and isolation of variants lacking the antigen
Authors: Basch Ross S. , Panagiotatos Thomas , Berman Joan W. , Buxbaum Joel N. 
Published: February 2005

AbstractMurine multipotential hematopoietic stem cells (CFU‐s) bear an antigen (SC‐1) which is recognized by heterologous antisera to mouse brain. We have found that cloned Thy‐1 negative variants of the T‐cell lymphoma RL ♂1 are sensitive to complement‐mediated cytolysis by anti‐brain serum and can absorb the anti‐stem cell activity from the antiserum. We have isolated several subclones derived from a primary Thy‐1 negative variant which are not susceptible to anti‐brain serum. The surface of the resistant lines has little or no antigen capable of binding anti‐mouse brain antibodies as measured by either immunofluorescence or a radioimmunoassay. These lines are also unable to absorb the anti‐bodies responsible for the cytotoxic effect of rabbit anti‐mouse brain serum against CFU‐s. We conclude that the predominant antigen, serologically detectable on Thy‐1 negative variants of RL ♂1, is SC‐1.
Insulin and growth factors stimulate rapid posttranslational changes in glucose transport in ovarian granulosa cells
Authors: Allen W. Ross , Nilsen‐Hamilton Marit , Hamilton Richard T. 
Published: February 2005

AbstractThe glucose analogues, 3‐0‐methyl‐D‐glucose and 2‐deoxy‐D‐glucose, have been used to characterize glucose transport and its regulation by serum and growth factors in monolayer cultures of granulosa cells obtained from bovine ovaries. Uptake of 3‐0‐methylglucose was shown to be independent of the Na+‐gradient, independent of energy, did not show accelerated exchange, and was stereospecific. Serum withdrawal resulted in a biphasic decrease in initial rates of glucose uptake with half‐times for the two phases of 50 minutes and 3 hours. Insulin could prevent the decrease in uptake rates with a half‐maximum concentration of 10.5 1/8 3 nM. Insulin was shown to stimulate DNA synthesis with a concentration of half‐maximum response of 28 nM.Insulin or serum stimulation of 3‐0‐methylglucose uptake in serum‐starved cells resulted in a two to threefold increase in initial rates, with a time for halfmaximum stimulation of 3 minutes. The insulin‐stimulated increase was insensitive to cycloheximide and cyanide during the first 30 minutes, and this early, rapid stimulation was also produced by brain FGF (fibroblast growth factor), pituitary FGF, epidermal growth factor, calf serum, and some but not all samples of follicular fluid. Insulin also stimulated 2‐deoxyglucose and a‐aminoisobutyric acid uptake during the first 5 minutes of addition, and these early stimulations were shown to be posttranslational changes.
Growth inhibitory protein(s) in the 3T3 cell plasma membrane. Partial purification and dissociation of growth inhibitory events from inhibition of amino acid transport
Authors: Raben Dan , Lieberman Michael A. , Glaser Luis 
Published: February 2005

AbstractIt has been reported previously that a plasma membrane‐enriched fraction from 3T3 cells arrests the growth of sparse 3T3 cells early in the G1 phase of growth (Whittenberger et al., ′78, ′79). Addition of membranes to sparse 3T3 cells also results in a decrease of the rate of transport of αaminoisobutyric acid (Lieberman et al., ′79). We have partially purified the growth‐inhibitory proteins from plasma membrane of 3T3 cells. These growth‐inhibitory proteins behave as intrinsic membrane proteins and do not require membrane lipids for activity. The partially purified proteins arrest cell growth without affecting the rate of α‐aminoisobutyric transport; thus, inhibition of both transport and cell growth are not obligatorily coupled events.