bluuebunny/crossref_metadata_2025_split huggingface dataset:
Evidence that a critical threshold of DNA polymerase‐alpha activity may be required for the initiation of DNA synthesis in mammalian cell heterokaryons
Authors: Pendergrass William R. , Saulewicz Anthony C. , Burmer Glenna C. , Rabinovitch Peter S. , Norwood Thomas H. , Martin George M. 
Published: February 2005

AbstractThe specific activity of DNA polymerase (90% alpha) was determined in nine “neoplastoid” cell lines (Martin and Sprague, 1973) and in three different strains of HDF (human diploid fibroblast‐like cells), all examined in logarithmic phases of growth. This was compared to the ability of each cell type to “rescue” (reinitiate DNA synthesis in) senescent HDF cells subsequent to polyethylene glycolmediated cell fusions. A sharp “threshold” value of DNA polymerase activity was observed below which reinitiation of DNA synthesis in heterokaryons with senescent HDF does not occur. This threshold was especially obvious when the specific activity of DNA polymerase (p moles dTTP incorporated per mg protein or per cell) was divided by the percent of S‐phase cells present in each culture as determined by flow microfluorometry. Our results indicate that the specific activity of DNA polymerase‐alpha (or some other factor tightly coregulated with it) in “recessive” cell types (those unable to rescue senescent cells) is only about two times this theoretical “threshold” value, and that fusion of recessive cell types to senescent HDF cells reduces the specific activity in the heterokaryon to below this minimum, thus preventing the cells from entering S phase.
Cells transformed by rous sarcoma virus release transforming growth factors
Authors: Krycève‐Martinerie Cécile , Lawrence David A. , Crochet Janine , Jullien Pierre , Vigier Philippe 
Published: February 2005

AbstractChicken embryo fibroblasts and hamster BHK cells transformed by Rous sarcoma virus (RSV) release in their culture media growth factors which enhance markedly anchorage‐independent colony formation in gelified medium, at the restrictive temperature (41°5 C), of chicken embryo fibroblasts (CEF) infected by RSV mutants with a ts mutation of the src gene. This action is not observed with uninfected CEF, and, therefore, appears to require some expression of the viral src gene in the target cells. The enhancing factors are proteins related to the family of the transforming growth factors (TGFs) by their molecular weight (about 20 kd), their heat and acid resistance, and their sensitivity to dithiothreitol. They do not compete with 125I EGF for binding on the EGF receptors of the membrane of A431 cells. As chicken embryo fibroblasts are devoid of EGF receptors, their activity is not potentiated by EGF.
Growth stimulation of Rous sarcoma virus‐transformed BHK cells by biotin and serum lipids
Authors: Cheng David K. S. , Moskowitz Merwin 
Published: February 2005

AbstractBiotin or a serum lipid extract stimulated proliferation of G1 arrested Rous sarcoma virus‐transformed BHK cells in modified Eagle's MEM (BM). The cells could be maintained continuously in BM plus biotin (BMB), but not in BM plus serum lipid extract (BM · L). Avidin inhibited growth stimulation when added to BMB, but did not inhibit growth when added to BM · L.14C‐acetate incorporation into total cellular lipids was stimulated in BMB, but not in BM. Thin‐layer chromatography of the labeled cellular lipid extract indicated that relatively large amounts of 14C‐acetate were incorporated into phosphatidylserine and little into the other major phospholipids. In the neutral lipids, the largest amount of incorporation was in cholesterol.G1 arrested cells multiplied rapidly in BM supplemented with dialyzed serum (BM · DS), but they did not multiply in BM with delipidized serum (BM · DLS). The addition of biotin or serum lipid extract to BM · DLS stimulated growth. Growth stimulation in BM · DLS by biotin was inhibited by avidin, but avidin had no effect on growth stimulation by serum lipid extract. Biotin stimulated additional multiplication in BM · DS and avidin inhibited this additional growth stimulation.These results suggest that growth stimulation requires lipids supplied by serum lipids or by de novo synthesis stimulated by biotin. In the absence of serum, the stimulation of the synthesis of growth factor(s) by biotin are also required for continuous multiplication.
Are factors originating from serum, plasma, or cultured cells involved in the growth‐promoting effect of the extracellular matrix produced by cultured bovine corneal endothelial cells?
Authors: Gospodarowicz D. , Gonzalez R. , Fujii D. K. 
Published: February 2005

AbstractThe possibilities that the growth‐promoting effect of the extracellular matrix (ECM) produced by cultured bovine corneal endothelial (BCE) cells could be due to: (1) adsorbed cellular factors released during the cell lysis process leading to the denudation of the ECM; (2) adsorbed serum or plasma factors: or (3) adsorbed exogenous growth factors have been examined. Exposure of confluent BCE cultures to 2 M urea in medium supplemented with 0.5% calf serum denudes the ECM without cell lysis. The ECM prepared by this procedure supports cell growth just as well as ECM prepared by denudation involving cell lysis. Thus, it is unlikely that the growth‐promoting properties of ECM are due to adsorbed cellular factors. When the ECM produced by BCE cells grown in defined medium supplemented with high‐density lipoprotein, transferrin, and insulin was compared to the ECMs produced by cells grown in the presence of serum‐ or plasma‐supplemented medium, all were found to be equally potent in stimulating cell growth. It is therefore unlikely that the growth‐promoting ability of the ECM is due to adsorbed plasma or serum components. When fibroblast growth factor (FGF)‐coated and ECM‐coated plastic dishes were submitted to a heat treatment (70°C, 30 min) which results in the inactivation of FGF, the growth‐supporting ability of FGF‐coated dishes was lost, while the comparable ability of ECM‐coated dishes was not affected significantly. This observation tends to demonstrate that the active factor present in the ECM is not FGF. Nor is it platelet derived growth factor (PDGF), since treatment known to destroy the activity of PDGF, such as exposure to dithiothreitol (0.1 M, 30 min, 22°C) or to β‐mercaptoethanol (10%) in the presence or absence of 6 M urea for 30 min at 227°C, does not affect the growth‐promoting activity of ECM. It is therefore unlikely that the growth‐promoting effect of ECM is due to cellular growth‐promoting agents or to plasma or serum factors adsorbed onto the ECM.
Reactivation of NAD(H) biosynthetic pathway by exogenous NAD+ in nil cells severely depleted of NAD(H)
Authors: Mandel Kenneth G. , Lively Mary K. , Lombardi Donald , Amos Harold 
Published: February 2005

AbstractThe culture of Nil hamster fibroblasts in MEM lacking nicotinamide (NAm− MEM) leads to: (1) the rapid loss of intracellular total nicotinamide adenine dinucleotide (NAD(H)) content in these cells from a level of 150–200 pmoles/105 cells to less than 20 pmoles/105 cells; (2) the cessation of cell division and inhibition of DNA synthesis; and (3) a reduction of glucose consumption and lactic acid production. In most situations, following nicotinamide starvation, the restoration of intracellular NAD(H) follows rapidly the readdition of NAD+ (oxidized), nicotinamide mononucleotide (NMN), nicotinamide, or nicotinic acid. Resumption of cell division occurs after only a lag of about 24 hours. Nil cells subcultured for three consecutive times in the absence of nicotinamide (3° NAm‐ cells) exhibit different behavior. These severely starved cells are incapable of quickly restoring their intracellular NAD(H) content to normal levels when provided with any pyridine ring compound except NAD+. One‐hour exposure of such cells to NAD+ allows utilization of nicotinamide to rapidly restore intracellular NAD(H). This short incubation with NAD+ does not result in any significant restoration of intracellular NAD(H) or lead to the accumulation of an intracellular pool of some precursor. This function of NAD+ as a stimulatory signal to the NAD(H)‐biosynthetic pathway in severely starved Nil cells is a previously unreported role of NAD+, and does not require protein synthesis.
Cell hybrids between SV40‐transformed macrophage cell lines and a Chinese hamster cell line: Growth responsiveness and induction of colony‐stimulating factor
Authors: Ohki Kazunori , Nagayama Ariki 
Published: February 2005

AbstractThree cell lines from resident macrophages of BALB/c mice and four from activated macrophages of the same strain were isolated by infection with simian virus 40 (SV40). A majority of these cells showed dependency on L cell‐conditioned medium (LCM), which is necessary for proliferation of normal macrophages in vitro. Somatic cell hybridization was applied in the study of macrophage growth responsiveness. A macrophage cell line (BR 15) with strict dependency on LCM for growth was fused to a Chinese hamster cell line (hs222‐16); it was found that dependency on LCM was a dominant trait in the hybrids. Following fusion fo a macrophage cell line (BAM3) which grew without LCM to hs222‐16, a large number of colonies appeared in the selection medium containing LCM . Four hybrids not requiring LCM for growth were selected in an LCM‐free culture, and their hybrid properties were examined. Three out of the four hybrids secreted colony‐stimulating factor (CSF) constitutively, whereas the fourth secreted no CSF. The level of acid phosphatase activity in the hybrids was higher than in the parent cells. Two peaks of CSF activity were observed after gel filtration chromatography of conditioned medium: One was eluted at molecular weight of 36,000 and the other at 17,000.
Modulation of microsomal glucose‐6‐phosphate translocase activity by free fatty acids: Implications for lipid domain structure in microsomal membranes
Authors: Hill David J. , Dawidowicz Eliezar A. , Andrews Mark L. , Karnovsky Morris J. 
Published: February 2005

AbstractRecent studies from this laboratory have demonstrated differential effects of cis‐unsaturated (type A) free fatty acids (FFA) and trans‐unsaturated or saturated (type B) FFA on protein‐mediated surface phenomena, namely, (1) capping on surface immunoglobulin in lymphocytes, (2) receptor‐mediated aggregation of platelets, and (3) adhesion of BHK cells. These results were explained in terms of the FFA‐perturbing specific lipid domains of the plasma membrane with subsequent modulation of the function of proteins occupying those domains. We wanted to determine if a differential effect of type A and type B FFA could be measured in an isolated membrane system, and chose to study the glucose‐6‐phosphate (Glc‐6‐P) translocase:hexose phosphate phosphohydrolase complex of rat liver endoplasmic reticulum. It was found that in intact microsomes, hydrolysis of Glc‐6‐P was inhibited by linoleic acid and linolenic acid. When the permeability barrier of the microsome was disrupted inhibition of hydrolysis was abolished. These results suggested that the Glc‐6‐P translocase was effected by the type A FFA. Importantly, palmitic acid, stearic acid, and elaidic acid had no significant effect on either translocation or hydrolysis of Glc‐6‐P. In addition, other microsomal enzymes, including the serine ethanolamine base exchange protein, diacylglycerol CDP choline phosphotransferase, diacylglycerol CDP ethanolamine phosphotransferase, NAD(P)H cytochrome C reductase, and NADH ferricyanide reductase were not significantly effected by the FFA used in these experiments. The FFA used, although bound to microsomes, were apparently not incorporated into phospholipids, or cyclooxygenated into prostaglandins during the time course of these experiments. Based on previous results showing that cis‐unsaturated FFA exert their greatest perturbing effects in gel‐like lipid, we postulate that the transport protein occupies such a gel‐like lipid domain in the endoplasmic reticulum bilayer.
Microtubule‐disrupting agents can independently affect the prereplicative period and the entry into S phase stimulated by prostaglandin F2α and fibroblastic growth factor
Authors: Otto Angela M. , De Asua Luis Jimenez 
Published: February 2005

AbstractFibroblastic growth factor (FGF) and prostaglandin F2α (PGF2α) are two different growth factors in Swiss 3T3 cells: each stimulates the initiation of DNA synthesis after a lag phase of 14–15 hr. Colchicine, colcemid, and nocodazole have no mitogenic effect on their own, but each has a synergistic effect on the rate of initiation of DNA synthesis stimulated by either FGF or PGF2α. Independent of the growth factor, the microtubule‐disrupting drug had to be added before 8 hr of the lag phase to enhance the stimulation. Colchicine could be removed at 5 hr of the lag phase with no impairment of the synergistic effect. However, both colcemid and nocodazole had to remain present up to 10 hr of the lag phase to achieve the full synergy. This result can be accounted for by the fact that colchicine, in contrast to colcemid and nocodazole, binds almost irreversibly to microtubules, and it strengthens the interpretation that the state of microtubular organization plays a role in regulating the rate of entry into S phase. Preincubating quiescent Swiss 3T3 cells with colchicine, colcemid, or nocodazole before stimulation by FGF or PGF2α shortened the lag phase by about 3 hr. When the microtubule‐disrupting drug was removed prior to stimulation, the lag phase was also shortened, but there was no enhancement of the rate of initiation of DNA synthesis, except upon preincubation with colchicine. This suggests that microtubule disruption can independently affect the length of the lag phase and the rate of initiation of DNA synthesis, and that these two phenomena can be uncoupled.
Hyperthermia‐induced increase in potassium transport in Chinese hamster cells
Authors: Stevenson Anita P. , Galey William R. , Tobey Robert A. , Stevenson Michael G. , Jett James H. 
Published: February 2005

AbstractThe effect of hyperthermia (42°C) on K+ transport has been examined by monitoring the uptake of 42KCl by Chinese hamster (line CHO) cells at both 37° and 42°C as well as 37°C following a 30‐min treatment at 42°C. Early studies showed that CHO cells exhibit two components of uptake at both 37° and 42°C. Consequently, it was necessary to determine rate equations describing a three‐compartment system (i.e., the bathing medium and two cell‐associated compartments) and apply them to this system. Using the equations describing the behavior of the tracer in each compartment as well as the general solutions given by Robertson et al (1957), a method was developed for obtaining the two components of influx. The two transport rates were determined from equations arising from mathematical analysis and requiring only measurable quantities for their solutions. The results of this study showed that (1) both components of influx at 42°C are increased over influx at 37°C; (2) the increase is significant (by an average 68% for the rapid component and 55% for the slow component); and (3) the increase is due entirely to an increase in active transport. The increased active transport is both immediate and quickly reversible when cells are returned to 37°C; resumption of cell division is delayed for several additional hours, indicating that changes in potassium transport are not directly responsible for the prolonged delay in cell division. Rather, the results suggest that increased activity of the membrane pump may be the initial event in a series of events resulting from heat treatment.
Induction of thermotolerance and enhanced heat shock protein synthesis in chinese hamster fibroblasts by sodium arsenite and by ethanol
Authors: Li Gloria C. 
Published: February 2005

AbstractSynthesis of a family of proteins called “heat shock” proteins is enhanced in cells in response to a wide variety of environmental stresses. This suggests that these proteins may have functions essential to cell survival under stressful conditions. A causative relationship between heat shock protein synthesis and development of thermotolerance would imply that agents known to induce heat shock protein synthesis, such as sodium arsenite, also induce thermotolerance. Conversely, agents known to induce thermotolerance, such as ethanol, would also enhance heat shock protein synthesis. To test this hypothesis, I have examined the effect of sodium arsenite or ethanol treatment on protein synthesis and cell survival in Chinese hamster ovary HA‐1 cells. After either sodium arsenite or ethanol treatment, the synthesis of heat shock proteins was greatly enhanced over that of untreated cells. In parallel, cell survival was increased as much as 104‐fold when cells exposed to either agent were challenged by a subsequent heat treatment. The synthesis of heat shock proteins correlated well with the development of thermotolerance. A qualitative analysis of individual proteins suggests that the synthesis of 70,000 and 87,000 molecular weight proteins most closely mirrored the development of thermotolerance. The results, therefore, strongly reinforce the hypothesis that a causal relationship exists between the enhanced synthesis of heat shock protein and cell survival under specific stresses.
The nature of 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA)‐stimulated hemopoiesis, colony stimulating factor (CSF) requirement for colony formation, and the effect of TPA on [125I]CSF‐1 binding to macrophages
Authors: Guilbert L. J. , Nelson D. J. , Hamilton J. A. , Williams N. 
Published: February 2005

AbstractThe tumor‐promoting phorbol diester, 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA) was found to act both independently of and synergistically with the mononuclear phagocyte specific colony stimulating factor (CSF‐1) to stimulate the formation of macrophage colonies in cultures of mouse bone marrow cells. In contrast, TPA did not synergize with other CSF subclasses that stimulate the formation of eosinophil, eosinophil‐neutrophil, neutrophil, neutrophil‐macrophage, and macrophage colonies, nor with either of the two factors required for megakaryocyte colony formation, megakaryocyte CSF, and megakaryocyte colony potentiator. In serum‐free mouse bone marrow cell cultures TPA retained the ability to independently stimulate macrophage colony formation. However, TPA‐stimulated colony formation was suboptimal and delayed in serum‐free cultures that could support optimal colony formation in the presence of CSF‐1. In addition, TPA did not directly compete with [125I]CSF‐1 at 4°C for its specific, high‐affinity receptor on mouse peritoneal exudate macrophages. However, a 2‐hour preincubation of the cells with TPA at 37° caused almost complete loss of the receptor. Thus, TPA is able to mimic CSF‐1 in its effects on CSF‐1 responsive cells in some aspects (the spectrum of target cells, the morphology of resulting colonies, and the ability to down‐regulate the CSF‐1 receptor) but it is not able to mimic CSF‐1 in other ways (TPA alone cannot stimulate the full CSF‐1 response, TPA does not stimulate the most primitive CSF‐1 responsive cells, and TPA does not bind to the CSF‐1 receptor).
The effects of thyroparathyroidectomy and 1,25 dihydroxyvitamin D3 on changes in the activities of some cytoplasmic and nuclear protein kinases during liver regeneration
Authors: Sikorska M. , Whitfield J. F. , Rixon R. H. 
Published: February 2005

AbstractPartial hepatectomy (HPX), which proliferatively activates the remaining liver cells, triggered two transient prereplicative surges in the total activities of cytoplasmic types I and II cyclic AMP‐dependent protein kinase holoenzymes, and of nuclear catalytic subunits from cyclic AMP‐dependent protein kinases. It also induced a transient prereplicative increase in the activities of a nuclear Ca2+‐calmodulin‐stimulable, protamine‐phosphorylating protein kinase, and a nuclear poly(L‐lysine)‐phosphorylating, 105 kDa protein kinase. Thyroparathyroidectomy (TPTX) delayed and reduced the first surge and completely eliminated the second surge of both of the cytoplasmic cyclic AMP‐dependent protein kinases, reduced the rises in the activity of nuclear catalytic subunits, and completely eliminated the surge of the Ca2+‐calmodulin‐stimulable protein kinase, but did not affect the surge of the nuclear 105 kDa protein kinase. The impairment of the responses of the two cyclic AMP‐dependent protein kinases to HPX in TPTX rats was not accompanied by a rise in the level of heat‐stable inhibitor of cyclic AMP‐dependent protein kinase activity. One intraperitoneal injection of 1,25‐dihydroxyvitamin D3 into TPTX rats immediately after HPX completely restored the post‐HPX surges in the activity of type I cyclic AMP‐dependent protein kinase, but the hormone, even in high doses, had little or no effect on the type II isoenzyme or the nuclear Ca2+‐calmodulin‐stimulable, protamine‐phosphorylating enzyme.
Effects of Zn2+ ions on protein phosphorylation in epithelial cell membranes
Authors: Alitalo Kari , Keski‐Oja Jorma , Bornstein Paul 
Published: February 2005

AbstractThe regulation of protein phosphorylation by Zn2+ ions and by other divalent cations was studied in membrane vesicles from a normal mouse epithelial cell line, MMC‐E (Mus musculus castaneous). Four major phosphoacceptor polypeptides were found in these membranes. Micromolar concentrations of Zn2+ ions inhibited the phosphorylation of the epidermal growth factor (EGF) receptor and of threonine residues in a 47,000‐dalton polypeptide. In contrast, two polypeptides with molecular weights of 54,000 and 57,000 showed increased phosphorylation, mainly of serine residues, in the p. esence of Zn2+ ions. These results were not obtained using similar concentrations of other divalent cations and were apparently not due to an effect of Zn2+ ions on phosphoprotein phosphatases. Thus, the effects of Zn2+ ions on protein phosphorylation in membrane vesicles are complex and are not restricted to an inhibition of a single protein phosphatase or kinase.
The role of calmodulin in the proliferation of transformed and phenotypically normal tsASV‐infected rat cells
Authors: Durkin Jon P. , Whitfield James F. , MacManus John P. 
Published: February 2005

AbstractNRK cells infected with a temperature‐sensitive, transformation‐defective mutant of avian sarcoma virus (ASV), tsLA23, behaved as if nontransformed at a nonpermissive 40°C and were rendered quiescent by serum deprivation. These serum‐deprived cells were stimulated to start entering S phase about 7 hours after serum addition at 40°C or about 9 hours after shifting the cultures to 36°C, a temperature allowing the production of active viral pp60src and expression of the transformed phenotype. The transit of both serum‐and temperature‐stimulated tsLA23‐NRK cells through later G1 was inhibited by the unrelated calmodulin antagonists W7 and R24571. The former drug was found to block the cells at a point in the cell cycle no more than 2 hours from the G1/S transition. The weaker calmodulin antagonist, W5, was less effective in impairing progression. Thus, calmodulin is likely required for the transit of both transformed and phenotypically normal tsLA23‐NRK cells through the later stages of their G1 phases. Cells neoplastically transformed by ASV contain more calmodulin than uninfected, non‐neoplastic cells. At the nonpermissive 40°C, the calmodulin content of the tsLA23‐NRK cells dropped to the non‐neoplastic level. When these phenotypically non‐transformed cells were enabled to reenter the cell cycle while still in low‐serum medium by a 40 to 36°C shift, they passed through the G1 and S phases and divided without a concomitant rise in the total calmodulin content. Thus, a calmodulin rise does not appear to be required for the expression of one characteristic of transformed cells, i.e., reduced requirement for exogenous growth factors.