bluuebunny/crossref_metadata_2025_split huggingface dataset:
Altered cellular responses to serum mitogens, including platelet‐derived growth factor, in cultured smooth muscle cells derived from arteries of patients with moyamoya disease
Authors: Aoyagi Masaru , Fukai Naomi , Sakamoto Hiroshi , Shinkai Tadashi , Matsushima Yoshiharu , Yamamoto Mari , Yamamoto Kiyotaka 
Published: February 2005

AbstractProgressive stenosis or occlusion of bi ateral internal carotid arteries by fibrocellular intimal thickening results in cerebral ischemia in moyamoya disease. The etiology is unknown. We examined cultured arterial smooth muscle cells (SMC) from scalp arteries of five patients with moyamoya disease. In this study we investigated the responsiveness of the cells in culture to serum mitogens including platelet‐derived growth factor (PDGF), a major mitogen of SMC, and compared the response to that of cells derived from age‐matched control patients. SMC from patients with moyamoya disease proliferated less rapidly in a medium with 15% serum than did control SMC and responded poorly to the addition of PDGF to 5% serum. PDGF alone did not stimulate SMC in a quiescent state to initiate DNA synthesis in moyamoya disease, without serum factors other than bovine serum albumin, though it significantly stimulated the controls. Simultaneous additions of epidermal growth factor, insulin‐like growth factor‐I, and PDGF stimulated initiation of DNA synthesis in cells from moyamoya disease, but not as much as PDGF alone did in the controls. Although direct correlations with the pathogenesis of the disease remain to be clarified, the results indicate altered interrelations between serum factors and the cellular responses in vessels of moyamoya disease.
Genetic characterization of adenovirus transformed cell revertants resistant to methylglyoxal bis(guanylhydrazone): Evidence for the involvement of three genetic loci
Authors: Alami Nezha , Chopra Arvind , Thirion Jean‐Paul 
Published: February 2005

AbstractFive new independent rat somatic cell mutants resistant to the antileukemic drug methylglyoxal bis(guanylhydrazone) (MGBG) were isolated after mutagen treatment. The mutants were 7‐ to 10‐fold more resistant to MGBG than were the parental wild‐type cells. When the MGBG‐resistant (MGR) mutants were exposed to the drug in the presence of Tween‐80, a nonionic detergent, they became as sensitive (MGS) to MGBG as the wild‐type cells, indicating that they were probably permeability mutants. Genetic analysis of hybrids between MGR mutants and wild‐type cells showed the MGR and the nontransformed alleles to be recessive to the MGS (wild‐type) and transformed phenotype, respectively. Complementation analysis of the seven mutants revealed three functional genetic units or loci responsible not only for the MGR phenotype but also for tumorigenicity as determined in nude mice. Only the MGS hybrids produced tumors in the nude mice, whereas the MGR hybrids and mutants did not. Our results suggest the existence of cellular membrane components that are responsible both for cellular tumorigenicity and resistance to MGBG.
Altered levels of phosphoinositide metabolites and activation of guanine‐nucleotide dependent phospholipase C in rat hepatic tumors
Authors: Lalwani Narendra D. , Hylemon Philip B. , Strom Stephen C. 
Published: February 2005

AbstractThe metabolism of phosphatidylinositol was studied in normal quiescent hepa‐tocytes, hepatocellular carcinomas induced by single dose of diethylnitrosamine, followed by 2‐acetylaminofluorene and partial hepatectomy (Solt‐Farber model), and in an established hepatoma cell line, JB1. The JB1 hepatoma cell line and hepatocellular carcinomas demonstrated a 4‐ to 5‐fold higher rate of turnover of [3H]‐inositol and [3H]‐glycerol than the control hepatocytes. Significantly, elevated levels of second messengers inositol 1,4,5‐trisphosphate and sn‐1,2‐diacylglycerol were noted in hepatic tumor cells within 4 hr of labeling with precursor molecules, whereas no detectable level of 3H‐labelled inositol trisphos‐phate was noted in quiescent hepatocytes, even after incubation with 10 mM LiCl for 30 min. Approximately 2.5‐fold higher specific activities of a guanine nucleotide and Ca+2 dependent phosphatidylinositol 4,5‐bisphosphate specific phospholipase C were detected in the hepatocellular carcinoma cells. The cellular location of the phospholipase C activity was also different, being membrane bound in hepatocytes and equally distributed between cytosolic and membrane factions in the hepatomas. These data are consistent with the hypothesis that the enhanced production of diacylglycerol and inositol 1,4,5‐trisphosphate in hepa‐tocellular carcinomas may be due to the activation of a guanine nucleotide dependent phosphatidylinositol 4,5‐bisphosphate specific phospholipase C. These data are the first to compare phosphoinositide turnover in normal liver and hepatic tumor cells and suggest that the sustained levels of second messengers is closely associated with the transformation and enhanced growth rate in hepatic tumor cells.
Insulin and other regulatory factors modulate the growth and the phosphoenolpyruvate carboxykinase (PEPCK) activity of primary rabbit kidney proximal tubule cells in serum free medium
Authors: Wang Yue , Taub Mary 
Published: February 2005

AbstractInsulin was observed to modulate the growth and the phosphoenolpyruvate carboxykinase(PEPCK) activity of primary cultures of rabbit renal proximal tubule cells in serum free medium. Insulin was stimulatory to primary proximal tubule cell growth at a concentration of 10−8 M. In contrast, insulin was inhibitory to a proximal tubule function, PEPCK activity, following a 5‐minute incubation period. An insulin dosage as low as 10−10 M was inhibitory to PEPCK activity, suggesting the involvement of insulin receptors. Although insulin was required at a significantly higher dosage to stimulate the growth of the primary renal proximal, tubule cells than to inhibit PEPCK activity, the elevated dosage required in order to observe a growth effect may be explained by the degradation of insulin by the primary renal proximal tubule cells. However the possible involvement of receptors for Insulin‐like Growth Factor I (IGF‐I) and Insulin‐like Growth Factor II (IGF‐II) in mediating the effects of insulin cannot be excluded. Other effector molecules were also examined with respect to their effects on PEPCK activity. The possible involvement of cyclic AMP in the control of the PEPCK activity of the primary renal cells was indicated by the stimulatory effects of 8 bromocyclic AMP, isobutyl methylxanthine (a cyclic AMP phosphodiesterase inhibitor), and forskolin (an activator of adenylate cyclase). Phorbol 12‐myristate 13‐acetate (TPA), which activates protein kinase C, was inhibitory. The actions of these effector molecules and insulin on the PEPCK activity of the primary renal cultures are remarkably similar to their effects on hepatic PEPCK. Several growth factors, fibroblast growth factor (FGF), and transforming growth factor beta (TGF beta) were also examined. FGF was observed to be stimulatory, whereas TCF beta was inhibitory to the PEPCK activity of the primary renal proximal tubule cells.
In vitro kinetics of basic fibroblast growth factor diffusion across a reconstituted corneal endothelium
Authors: Dabin Isabelle , Courtois Yves 
Published: February 2005

AbstractWe have studied the ability of bFGF to traverse and be trapped within basement membranes. An extract of EHS tumor (matrigel) coated on culture chamber filters was used as an in vitro model of basement membranes. Our results showed a slow diffusion of bPGF dependent on the amount of low affinity binding sites present within the matrigel. High amounts 01 bFGF and heparin increased the initial rate of diffusion by displacement of bFGF bound to matrigel. An in vitro corneal endothelium model (endothelial cells overlying matrigel) was also developed. This monolayer, with a weak permeability, decreased the kinetic rate of bFGF diffusion compared with matrigel alone. These results indicate that modulation of bFGF distribution in a tissue by a basement membrane is dependent on bFGF concentration, basement membrane composition, permeability of associated cells, and local presence of heparin. This selective control may be a regulating step in bFGF action.
Effects of a putative prostaglandlin E2 antagonist, AH6809, on chondrogenesis in serum‐free cultures of chick limb mesenchyme
Authors: Capehart Anthony A. , Biddulph David M. 
Published: February 2005

AbstractIn the present study, we have examined the effects of a putative antagonist of prostaglandin E2 (PGE2), AH6809, on chondrogenesis in serum‐free cultures of mesenchyme from distal tips of stage 25 chick limb buds in order to test the hypothesis that endogenous PGE2, through receptor‐linked adenylate cyclase (AC), initiates differentiation of cartilage in limb mesenchyme. Daily addition of 10−4 M concentrations of AH6809 produced marked inhibition of chondrogen‐esis over a 5‐day period of cell culture as evaluated by Alcian green binding to cartilage matrix components. Inhibition of chondrogenesis by this compound was further shown to be reversible and treatment of cells with the antagonist limited to periods when chondrocytes had differentiated and were actively secreting cartilage‐specific matrix components had little effect. Preincubation of control cells in 1094 M concentrations of AH6809 inhibited PGE induced activation of AC by greater than 80% without significant (P>.05) inhibition of basal activity by the antagonist. Responses to parathyroid hormone, which increased AC activity by 7‐fold, and forskolin which increased AC activity by 23‐fold in control cells, were also uninhibited by preincubation in AH6809. The results demonstrate that blockade of PGE2‐AC linked receptors in prechondrogenic limb mesenchyme inhibits chondrogenesis supporting the hypothesis that endogenous PGE2 concentrations in undifferertiated limb mesenchyme play aninitiating role in the differentiation of cartilage.
Replacement of media in cell culture alters oxygen toxicity: Possible role of lipid aldehydes and glutathione transferases in oxygen toxicity
Authors: Sullivan S. J. , Roberts R. J. , Spitz D. R. 
Published: February 2005

AbstractReplacement of media in cell cultures during exposure to hyperoxia was found to alter oxygen toxicity. Following 100 hr of exposure to 95% or 80% O2, the surviving fraction (SF) of Chinese hamster fibroblasts, as assayed by clonogenicity, was less than 1 × 10−3 when the culture media was replaced only at the onset of the O2 exposure. Media replacement every 24 hr throughout the hyperoxic exposure resulted in SFs of 1.7 × 10−1 (95% O2) and 1.9 × 10−1 (80% O2) at 95 hr. Cellular resistance to and metabolism of 4‐hydroxy‐2‐nonenal (4HNE), a cytotoxic byproduct of lipid peroxidation, was examined in cells 24 hr following exposure to 80% O2 for 144 hr with media replacement. These O2‐exposed cells were resistant to 4HNE, requiring 2.6 times as long in 80 μM 4HNE to reach 30% survival as compared to density‐matched normoxia control. Furthermore, during 40 and 60 min of exposure to 4HNE, the O2‐preexposed cells metabolized greater quantities of 4HNE (fmole/cell) relative to control. The activity of glutathione S‐transferase (GST), an enzyme believed to be involved with the detoxification of 4HNE, was significantly increased in the O2‐preexposed cells compared with controls. Catalase activity was significantly increased, but no change was found in total glutathione content, glutathione peroxidase, manganese superoxide dismutase, and copper‐zinc superoxide dismutase activities at the time of 4HNE treatment in the O2‐preexposed cells relative to density‐matched control. The results demonstrate that in vitro tolerance to the cytotoxic effects of hyperoxia can be achieved through media replacement during O2 exposure. Tolerance to oxygen toxicity conferred resistance to the cytotoxic effects of 4HNE, possibly through GST‐catalyzed detoxification. These results provide further support for the hypothesis that toxic aldehydic byproducts of lipid peroxidation contribute to hyperoxic injury.
Amplification of the metallothionein‐1 and metallothionein‐2 genes in copper‐resistant hepatoma cells
Authors: Czaja Mark J. , Weiner Francis R. , Freedman Jonathan H. 
Published: February 2005

AbstractThe molecular basis for increased metallothionein concentrations in copperresistant hepatoma cells was examined. The copper‐resistant cell line HAC600, which is maintained in 600 μm copper, had increased steady‐state mRNA levels for both the metallothionein‐1 (MT‐1) and the metallothionein‐2 (MT‐2) genes. Levels of mRNA were increased 11‐fold for MT‐1 and 15‐fold for MT‐2, with no significant change in α‐tubulin mRNA content. HAC600NM cells, which are copper‐resistant cells kept in a normal copper concentration for over 1 year, also had eight‐ and tenfold increases in MT‐1 and MT‐2 mRNA levels. Nuclear run‐on assays showed that MT‐1 and MT‐2 gene transcription was increased nine‐ and eightfold in HAC600 cells and seven‐ and tenfold in HAC600NM cells, respectively. Southern blot analysis showed amplification of both metallothionein genes in HAC600 and HAC600NM cells. Thus the molecular basis of increased metallothionein in these hepatoma cells involved a stable gene amplification of both MT genes. The greater increase in metallothionein mRNA levels in HAC600 cells relative to the changes in transcription suggests that posttranscriptional mechanisms of gene regulation may also be acting in these cells.
Transforming growth factor type beta 1 (TGF‐β1) down‐regulates interleukin‐2 production and up‐regulates interleukin‐2 receptor expression in a thymoma cell line
Authors: D'Angeac Arnaud Dupuy , Dornand Jacques , Emonds‐Alt Xavier , Jullien Pierre , Garcia‐Sanz Jose A. , Erard François 
Published: February 2005

AbstractTransforming growth factor type β1 (TGB‐β1) belongs to a family of polypeptides with regulatory effects on growth and differentiation of a variety of cell types. TGB‐β1 plays an important role in regulation of immune response by acting as a negative control signal for T cell proliferation through still unknown mechanisms. In this study we have analysed the effects of TGB‐β1 on EL 4–6.1, a variant of the murine EL 4 thymoma, which can be induced by phorbol 12‐myristate 13‐acetate (PMA) and/or interleukin 1 (IL‐1) to secrete interleukin 2 (IL‐2) and express IL‐2 receptors (IL‐2R). Using this defined model system, we show that TGB‐β1 simultaneously down‐regulates IL‐2 expression and up‐regulates the number of both high and low affinity IL‐2R. These changes correlate with changes at the mRNA level, suggesting an effect at the pre‐translational level. The specificity of both TGF‐β1 effects was demonstrated using a neutralizing antiserum to TGF‐β1. Our data also suggest that TGF‐β1 does not interfere with early activation signals of PMA and/or IL‐1. This model might be useful for elucidating the complex role of TGF‐β1 in the regulation of T cell responses.
Increased proteoglycan synthesis by the cardiovascular system of coarctation hypertensive rats
Authors: Lipke David W. , Couchman John R. 
Published: February 2005

AbstractProteoglycan (PG) synthesis in the cardiovascular system of coarctation hypertensive rats was examined by in vivo and in vitro labeling of glycosaminoglycans with 35SO4 in rats made hypertensive for short (4 days) and longer (14 days) durations. With in vivo labeling, only tissues directly exposed to elevated pressure (left ventricle, LV and aorta above the clip, AOR ↑) exhibited elevated PG synthesis after 4 days of hypertension. By 14 days, tissues both exposed to (LV and AOR ↑) and protected from elevated pressure (right ventricle and kidney) exhibited elevated PG synthetic rates. Slight elevations in the proportion of galactosamnoglycans were observed with a concurrent proportional decrease in heparan sulfate PGs. Using the in vitro labeling procedure, no significant increases in PG synthesis were observed in any tissue at either 4 days or 14 days of hypertension. These data indicate that: (1) coarctation hypertension stimulates PG production that is dependent initially on increased pressure and later, on additional non‐pressure related factors, (2) these other factors are responsible for enhanced PG production in tissues not directly exposed to pressure overload, (3) pressure and/or these other factors are essential for enhanced PG production in coarctation hypertension, and (4) synthesis of all GAG types appears to be affected.
Serum factor induces selective increase in Na‐channel expression in cultured skeletal muscle
Authors: Brodie Chaya , Sampson S. R. 
Published: February 2005

AbstractWe have examined effects of horse serum (HS) and various fractions (1 million‐1M, 300K, 100K, and 30K nominal molecular weight limit) obtained by ultrafiltration on expression of TTX‐sensitive Na‐channels and on activities of the Na‐K pump and glucose transport systems in cultured myotubes obtained from 1‐2‐day‐old neonatal rat pups. Five‐day‐old cells were transferred to serum‐free medium with no hormone or growth factor supplements (DMEM) for 24 hr and then treated with the various serum fractions for 48 hr. Measurements were made of specific [3H]‐saxitoxin (STX) binding, action potential properties, 86Rb‐uptake and 2‐deoxyglucose (2‐DG) uptake. HS significantly increased all parameters compared to DMEM (increases in STX‐binding, 69%; Rb‐uptake, 65%; 2‐DG uptake, 93%). Results of treatment with the separate fractions showed that the 300K fraction caused a significantly greater increase in STX‐binding than either HS or the other fractions. In contrast, the increases in Rb and 2‐DG uptakes induced by the different fractions were not different from that obtained with HS. We conclude that serum contains a factor that selectively increases expression of TTX‐sensitive Na‐channels in skeletal muscle.
Kidney‐specific enzyme expression by human kidney cell lines generated through oncogene transfection
Authors: Robinson Polly S. , Goochee Charles F. 
Published: February 2005

AbstractThe human kidney cell line 293 was generated by transfection of adenovirus DNA into normal human embryonic kidney (HEK) cells (Graham et al., 1977), whereas the human kidney cell lines ST‐1i and STt‐4i were generated by transfection of HEK cells with plasmids encoding SV40 viral oncogenes (Abcouwer et al., 1989). In this study, we examined kidney‐specific enzyme activity levels in 293, ST‐1i, and STt‐4i cells to determine their ability to exhibit kidney‐specific gene expression. Enzymes examined were leucine aminopeptidase (LAP), γ‐glutamyl transpeptidase (γ‐GTP), and the disaccharidases trehalase and maltase. Enzymatic activity levels were compared to three other kidney cell lines (MDCK, OK, and LLC‐PK1) as well as to normal human embryonic kidney (HEK) cells and the human hepatoma cell line, Hep G2. Modulation of kidney‐specific enzyme activities was assessed in response to several differentiation‐inducing agents (adenosine, n‐butyric acid, hexamethylene bisacetamide (HMBA), dimethyl sulfoxide (DMSO), N, N′‐dimethylformamide (DMF), isobutyl methyl xanthine (IBMX), di butyryl cAMP, and retinoic acid). ST‐1i and STt‐4i exhibit elevated levels of LAP, γ‐GTP, trehalase, and maltase, consistent with their kidney cell origin, whereas 293 cells exhibit elevated levels of just γ‐GTP and maltase. Maltase and γ‐GTP enzyme activities in ST‐1i and STt‐4i cells were very responsive to the various inducing agents; 293 cells were less responsive at the inducer concentrations examined. None of the three human cell lines formed domes under any of the experimental conditions. In summary, ST‐1i and STt‐4i are comparable to normal HEK cells in expression of kidney‐specific enzymes and in responsiveness to differentiation‐inducing agents, in spite of continued expression of SV40 oncogenes.
Analysis of the c‐myc P2 promoter
Authors: Moberg Kenneth H. , Andrew Tyndall W. , Pyrc Janina , Hall David J. 
Published: February 2005

AbstractThe cis‐acting elements governing transcription from the murine c‐myc P2 promoter have not been well defined. To gain a better understanding of the nature of the protein‐DNA interactions that take place on the P2 promoter, protein binding assays were performed. The ME1a2 and E2F factors appear to be the predominant proteins bound to a region spanning positions −140 to −24 relative to the P2 transcription start site. By a number of criteria, these factors appear to be distinct. When c‐myc promoter sequences were coupled to the chloramphenicol acetyltransferase gene (CAT) and transiently transfected into tissue culture cells it was found that optimal transcription from P2 was heavily dependent on the ME1a2 element.
Characterization of sarcoplasmic reticulum in skinned heart muscle cultures
Authors: Weinstein Leah , Brik Hadassa , Rotmensch Heschi H. , Shainberg Asher 
Published: February 2005

AbstractThe plasma membranes of rat heart muscle, grown in cell culture, were made permeable with saponin in a Ca‐free solution. The cells were then supplied with a medium resembling the cytosol, and the adenosine triphosphate (ATP)‐dependent Ca2+ sequestration was measured in the presence of oxalate. The nonmitochondrial component accounts for about 50% of the total Ca2+ uptake. The nonmitochondrial accumulation of Ca2+ within myocardial cells was found to be reversible by addition of the Ca2+ ionophore A23187. On the other hand, the Ca2+ antagonist D‐600 (50 μM) had almost no effect on Ca2+ accumulation. Caffeine reduced Ca2+ accumulation in the skinned cardiomyocytes in a concentration‐dependent manner. In addition, the anticalmodulin drug trifluo‐perazine (TFP) reduced Ca2+ accumulation in the skinned cells. Because of the analogy between nonmitochondrial ATP‐dependent Ca2+ accumulation and the sarcoplasmic reticulum (SR) function with regard to the influence of various agents, it is assumed that we actually measure Ca2+ accumulation in the SR. The rate of Ca2+ accumulation into the SR measured during the development of the cardiomyocytes in culture shows an almost linear increase as a function of culture age. Amiodarone, a potent antiarrhythmic agent, and its metabolite, desethyl‐amiodarone, inhibited Ca2+ accumulation into SR, which may explain their therapeutic effect.
Relationship of IgE receptor topography to secretion in RBL‐2H3 mast cells
Authors: Seagrave JeanClare , Pfeiffer Janet R. , Wofsy Carla , Oliver Janet M. 
Published: February 2005

AbstractIn RBL‐2H3 rat leukemic mast cells, cross‐linking IgE‐receptor complexes with anti‐lgE antibody leads to degranulation. Receptor cross‐linking also stimulates the redistribution of receptors on the cell surface, a process observed here by labeling the anti‐lgE with 15 nm protein A‐gold particles that are visible by back‐scattered electron imaging in the scanning electron microscope. We report that anti‐lgE binding stimulates the redistribution of IgE‐receptor complexes at 37°C from a dispersed topography to distributions dominated sequentially by short chains, small clusters, and large aggregates of cross‐linked receptors. Cells incubated with 1 μg/ml anti‐IgE, a concentration that stimulates maximum net secretion, redistribute receptors into chains and small clusters during a 15 min incubation period. At 3 and 10 μg/ml anti‐IgE, net secretion is reduced and the majority of receptors redistribute rapidly into clusters and large aggregates. The addition of Fab fragments with the high anti‐IgE concentrations, to reduce cross‐linking, delays receptor aggregation and enhances secretion. The progression of receptors from small clusters to large aggregates is prevented in cells treated with dihydrocytochalasin B to prevent F‐actin assembly. These results establish that characteristic patterns of receptor topography are correlated with receptor activity. In particular, they link the formation of large receptor aggregates to reduced signalling activity. Cytoskeleton‐membrane interaction is implicated in the formation or stabilization of the large receptor clusters.