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R Bartrons

Publications and source records attributed to R Bartrons.

At least 55 records · Page 3Linked to original sources
PubMed1996

Hepatocyte growth factor and transforming growth factor beta regulate 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase gene expression in rat hepatocyte primary cultures.

Hepatocyte growth factor (HGF) and transforming growth factor beta (TGF-beta) are believed to be of major importance for hepatic regeneration after liver damage. We have studied the effect of these growth factors on fructose 2,6-bisphosphate (Fru-2,6-P2) levels and the expression of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (6PF2K/Fru-2,6-BPase) in rat hepatocyte primary cultures. Our results demonstrate that HGF activates the expression of the 6PF2K/Fru-2,6-BPase gene by increasing the levels of its mRNA. As a consequence of this activation, the amount of 6PF2K/Fru-2,6-BPase protein and 6-phosphofructo-2-kinase activity increased, which was reflected by a rise in Fru-2,6-P2 levels. In contrast, TGF-beta decreased the levels of 6PF2K/Fru-2,6-BPase mRNA, which led to a decrease in the amount of 6PF2K/Fru-2,6-BPase protein and Fru-2,6-P2. The different actions of HGF and TGF-beta on 6PF2K/Fru-2,6-BPase gene expression are concomitant with their effect on cell proliferation. Here we show that, in the absence of hormones, primary cultures of hepatocytes express the F-type isoenzyme. In addition, HGF increases the expression of this isoenzyme, and dexamethasone activates the L-type isoform. HGF and TGF-beta were able to inhibit this activation.

Animals↗
PubMed1996

MPP+ toxicity in rat striatal slices: relationship between non-selective effects and free radical production.

Incubations of rat striatal slices have been used to assay MPP+ neurotoxicity. MPP+, at concentrations of 1 mM or higher, caused a marked increase in hydroxyl radicals, measured as malondialdehyde (MDA) accumulation, but not in nitric oxide production. At these doses, MPP+ showed an effect on dopamine terminals, causing a massive dopamine decrease, and on non-neuronal glial cells, where a marked reduction in glutamine synthetase activity was detected. At lower concentrations (25 mu M), the toxic effect on dopaminergic endings was maintained without increasing malondialdehyde concentrations or inhibiting glutamine synthetase activity. The effect on glutamine synthetase was prevented by the addition to the medium of 0.5% dimethyl sulfoxide, a hydroxyl-radical scavenger, but this did not protect the effect of dopamine depletion. We propose that non-selective effects of MPP+, at doses of 1 mM or higher, are mediated by extracellular overproduction of hydroxyl radicals. The main factor responsible for this overproduction would not be the released dopamine but rather the MPP+ itself through non selective inhibition of the mitochondrial respiratory chain or through a redox cycling that can trigger oxygen radical production.

1-Methyl-4-phenylpyridinium↗
PubMed1996

Activity of phosphoglycerate mutase and its isoenzymes in serum after acute myocardial infarction.

Aims/background-In humans there are three phosphoglycerate mutase (PGM, EC 5.4.12.1) isoenzymes (MM, MB and BB) which have similar distribution and developmental pathways to creatine kinase (CK, EC 2.7.3.2) isoenzymes. Total serum PGM activity increases in acute myocardial infarction with the same time course as creatine kinase activity. The present study was undertaken to determine changes in the activity of PGM and its isoenzymes after acute myocardial infarction.Methods-PGM activity was measured spectrophotometrically, by coupling the formation of 2-phosphoglycerate from 3-phosphoglycerate with enolase, pyruvate kinase and lactate dehydrogenase catalysed reactions. Inter- and intra-assay reproducibility was assessed. PGM isoenzyme activities were measured using cellulose acetate electrophoresis.Results-Total PGM activity in serum was increased in patients with a confirmed diagnosis of acute myocardial infarction. PGM activity peaked 12 to 24 hours after the onset of symptoms and returned to normal values within 48 hours. Electrophoretic analysis of serum from healthy subjects showed a band corresponding to BB-PGM and two other artefactual bands that did not correspond to adenylate kinase. After myocardial infarction, BB-PGM activity increased and MB-PGM and MM-PGM could be detected. On immunoblot analysis, normal serum contained an inactive form of MM-PGM with a smaller molecular weight than that of PGM tissue isoenzymes.Conclusions-Total serum PGM activity increased in patients with acute myocardial infarction, following the same temporal course as creatine kinase activity. The increase in MM-PGM and MB-PGM activities in these patients was not as high as expected. It is suggested that PGM isoenzymes, after release into the blood, undergo postsynthetic, probably proteolytic, transformation.

Journal Article↗
PubMed1996

Effects of taxol on isolated rat hepatocyte metabolism.

Taxol is a natural product, isolated from Taxus brevifolia, with increasing clinical applications because of its potent antitumor activity. Although it is mainly metabolized in the liver, its effect on hepatocyte metabolism has been scarcely investigated. In this study, the response of isolated rat hepatocytes to taxol was evaluated by correlating the changes observed in global indexes such as viability and heat dissipation with those produced in oxygen consumption and intracellular metabolites (ATP, fructose 2,6-bisphosphate, and lactate). The results indicate that taxol reduces aerobic metabolism, which induces an insufficient increase of the ATP via anaerobic glycolysis. Moreover, incubations of isolated mitochondria with taxol indicate that the respiratory chain is directly affected by this drug.

Animals↗
PubMed1995

Chronic effects of single intrastriatal injections of 6-hydroxydopamine or 1-methyl-4-phenylpyridinium studied by microdialysis in freely moving rats.

Extracellular dopamine (DA) and its main cerebral metabolites, dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), were measured by bilateral striatal microdialysis in rats at different times (2, 7, 15 and 60 days) after unilateral administration into the right striatum of 1-methyl-4-phenylpyridinium ion (MPP+) or 6-hydroxydopamine (6-OHDA). In both cases the decrease in extracellular dopamine did not exceed 40% of control values. The response of DOPAC and HVA depended on the treatment: MPP+ caused a marked acute decrease in the dopamine metabolites but allowed a progressive recovery that was very evident after 60 days; 6-OHDA caused a progressive decrease in the dopamine metabolites throughout the two months of the study. Tyrosine hydroxylase immunostaining revealed severe neuronal loss in substantia nigra two months after striatal administration of 6-OHDA, whereas no significant neuronal loss was found at the same time after MPP+ administration. A bilateral challenge infusion of MPP+ through the microdialysis probe was used to assess the dopaminergic capacity of both striata: at all the times studied there was a sharp depletion of DA on the non-lesioned side; both MPP(+)- and 6-OHDA-treated striata were unresponsive after a short time (2 days); after 2 months the response in MPP(+)-lesioned rats was similar on both sides, whereas 6-OHDA-lesioned striata were still unresponsive to MPP+. In rats, then, the effects of MPP+ could be partly reversed whereas the effects of 6-OHDA were not. These results suggest that neurotoxins causing striatal dopamine loss may act through different mechanisms, which could be significant for the etiopathogenic development of Parkinson's disease.

1-Methyl-4-phenylpyridinium↗
PubMed1995

Role of the N-terminal region in covalent modification of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase: comparison of phosphorylation and ADP-ribosylation.

The effect of cyclic AMP (cAMP)-dependent phosphorylation and ADP-ribosylation on the activities of the rat liver bifunctional enzyme, 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (PFK-2/FBPase-2), was investigated in order to determine the role of the N-terminus in covalent modification of the enzyme. The bifunctional enzyme was demonstrated to be a substrate in vitro for arginine-specific ADP-ribosyltransferase: 2 mol of ADP-ribose was incorporated per mol of subunit. The Km values for NAD+ and PFK-2/FBPase-2 were 14 microM and 0.4 microM respectively. A synthetic peptide (Val-Leu-Gln-Arg-Arg-Arg-Gly-Ser-Ser-Ile-Pro-Gln) corresponding to the site phosphorylated by cAMP-dependent protein kinase was ADP-ribosylated on all three arginine residues. Analysis of ADP-ribosylation of analogue peptides containing only two arginine residues, with the third replaced by alanine, revealed that ADP-ribosylation occurred predominantly on the two most C-terminal arginine residues. Sequencing of the ADP-ribosylated native enzyme also demonstrated that the preferred sites were at Arg-29 and Arg-30, which are just N-terminal to Ser-32, whose phosphorylation is catalysed by cAMP-dependent protein kinase (PKA). ADP-ribosylation was independent of the phosphorylation state of the enzyme. Furthermore, ADP-ribosylation of the enzyme decreased its recognition by liver-specific anti-bifunctional-enzyme antibodies directed to its unique N-terminal region. ADP-ribosylation of PFK-2/FBPase-2 blocked its phosphorylation by PKA, and decreased its PFK-2 activity, but did not alter FBPase-2 activity. In contrast, cAMP-dependent phosphorylation inhibited the kinase and activated the bisphosphatase. These results demonstrate that ADP-ribosylation of arginine residues just N-terminal to the site phosphorylated by PKA modulate PFK-2 activity by an electrostatic and/or steric mechanism which does not involved uncoupling of N- and C-terminal interactions as seen with cAMP-dependent phosphorylation.

Adenosine Diphosphate Ribose↗
PubMed1995

Cloning and expression of a catalytic core bovine brain 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase.

A cDNA encoding the catalytic core of a novel brain 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase isoenzyme was isolated from a lambda gt10 bovine brain library. This brain cDNA begins and ends in an open reading frame encoding a peptide of 476 amino acids. This peptide contains both the catalytic kinase and bisphosphatase domains and has an overall 65% and 67% indentity with the bovine heart and liver isozymes, respectively, whereas the NH2 and COOH-termini are divergent. An active catalytic core brain bifunctional enzyme was expressed in E. coli using a T7 RNA polymerase-based expression system. These results support the presence of a distinct gene coding for the protein in bovine brain.

Amino Acid Sequence↗
PubMed1995

Cerebrospinal dopamine metabolites in rats after intrastriatal administration of 6-hydroxydopamine or 1-methyl-4-phenylpyridinium ion.

Dopamine (DA) and its main cerebral metabolites, dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) were measured in striatum and cerebrospinal fluid (CSF) from cisterna magna in rats bilaterally lesioned by intrastriatal administration of 6-hydroxydopamine (6-OHDA) or 1-methyl-4-phenylpyridinium ion (MPP+). 6-OHDA caused a progressive lesion in striatum that is only moderately reflected in the decrease in dopamine metabolite concentration in CSF. MPP+ caused an acute but less selective lesion in the dopamine striatal system, as indicated by a significant reduction in striatal GABA content, followed by a slow recovery in dopamine striatal metabolism and content. The locomotor activity was dramatically reduced in both groups 48 hours after the treatment but remained significantly decreased after two months only in 6-OHDA lesioned animals. A positive correlation was found between HVA CSF concentration and striatal DA content in MPP+ lesioned rats, but not in 6-OHDA lesioned rats. It is concluded that the concentration of dopamine metabolites in CSF can be altered only after a severe striatal lesion: reduction of striatal dopamine content below 50% of normal values and involvement of neuronal or non-neuronal elements other than the dopaminergic system, similarly to the lesions caused by MPP+. These results may partly explain why CSF dopamine metabolites concentrations were significantly decreased both in advanced stages of parkinsonism and in other neurodegenerative disorders.

1-Methyl-4-phenylpyridinium↗
PubMed1994

Gene expression of enzymes regulating ketogenesis and fatty acid metabolism in regenerating rat liver.

Levels of mRNA for mitochondrial 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) synthase, carnitine palmitoyltransferase I (CPT I) and carnitine palmitoyltransferase II (CPT II), fatty acid synthase (FAS) and actin were analysed during liver regeneration. mRNA levels for mitochondrial HMG-CoA synthase decreased rapidly, reaching a minimum 12 h after partial hepatectomy and returning to normal at 24-36 h. In contrast, CPT I, CPT II and FAS mRNAs increased throughout the period examined. Expression of actin increased significantly during regeneration. Levels of mRNA for mitochondrial HMG-CoA synthase also decreased as a result of surgical stress, although the effect of hepatectomy was much greater. We determined the levels of mitochondrial HMG-CoA synthase using specific antibodies. The amount of protein rapidly decreased, although less markedly than the corresponding mRNA levels. These results show that the decrease described in ketogenesis in partially hepatectomized rats correlated with the decrease in the expression of mitochondrial HMG-CoA synthase, suggesting that this enzyme may also be a control point in ketogenesis in the regenerating liver, as it is in normal and diabetic rats.

Actins↗
PubMed1994

Control of fructose 2,6-bisphosphate metabolism by different mitogenic signals in Swiss 3T3 fibroblasts.

The mitogenic signals that control fructose 2,6-bisphosphate metabolism in murine Swiss 3T3 fibroblasts have been studied. Bombesin, vasopressin, insulin, protein kinase C activation by phorbol esters, or increase in the intracellular cAMP concentration by forskolin induced an increase in fructose 2,6-bisphosphate levels. When the cells were incubated in the presence of insulin or phorbol esters, an increase in the Vmax of 6-phosphofructo-2-kinase activity was observed. However, forskolin did not produce this effect. The increase in 6-phosphofructo-2-kinase activity elicited by phorbol 12,13-dibutyrate was blocked by cycloheximide. In contrast, the effect of insulin did not require protein synthesis. This study demonstrates that different mitogenic signal transduction pathways control the levels of fructose 2,6-bisphosphate. The high rate of glycolysis in proliferating Swiss 3T3 cells may be explained by an increase in the levels of this regulatory metabolite.

3T3 Cells↗
PubMed1994

Effect of different preservation solutions on adenine nucleotide content and metabolism in human kidney transplantation.

Differences in purine metabolism produced by three preservation solutions were studied by determining the adenine nucleotide (ATP, ADP, AMP, and IMP) and nucleoside (adenosine, inosine, and hypoxanthine) levels in human kidney cortical biopsies. Forty kidney allografts were studied using University of Wisconsin (UW) solution (n = 20), Euro-Collins (EC) solution (n = 12), and modified EC solution with mannitol (M; n = 8). No significant differences were found between the three solutions studied with regard to ATP, ADP, or AMP changes. The mean ATP level (nmol/mg prot +/- SEM) at the end of preservation in the UW group was 2.7 +/- 0.3 nmol/mg, in the EC group 3.8 +/- 0.7 nmol/mg, and in the M group 2.3 +/- 0.4 nmol/mg. ATP 30 min after reperfusion in the UW, EC, and M groups was 5.7 +/- 0.8 nmol/mg, 6.4 +/- 1.0 nmol/mg, and 4.6 +/- 0.5 nmol/mg, respectively. However, an important difference appeared in the catabolic products determined. Kidneys perfused with UW solution had a significantly higher level of adenosine (2.6 +/- 0.6 nmol/mg), inosine (11.8 +/- 2.2 nmol/mg), and hypoxanthine (18.1 +/- 2.1 nmol/mg) at the end of cold storage than those perfused with EC (0.4 +/- 0.1 nmol/mg, 2.0 +/- 0.8 nmol/mg, and 7.1 +/- 1.4 nmol/mg) and M solutions (0.2 +/- 0.05 nmol/mg, 0.5 +/- 0.1 nmol/mg, and 5.2 +/- 0.6 nmol/mg; P < 0.05). These levels returned to initial values 30 min postreperfusion and there were no differences with the EC or M solution groups at that time.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenine Nucleotides↗
PubMed1994

Cerebrospinal monoamine metabolites and amino acid content in patients with parkinsonian syndrome and rats lesioned with MPP+.

Monoamine metabolites and amino acid concentration in cerebrospinal fluid (CSF) of 33 untreated patients with parkinsonian syndrome, and 20 control patients without specific neurological symptoms have been compared with those obtained in cerebrospinal fluid of rats intrastriatally lesioned with 1-methyl-4-phenylpyridinium ion (MPP+) and sham operated animals. Homovanillic acid content was found to be significantly lower in patients with severe parkinsonism (motor score of UPDRS > 24), but not in patients with mild symptoms (motor score < or = 24). A correlation between the loss of striatal dopamine and the decrease in cerebrospinal homovanillic acid has been established in rats treated with MPP+. The extrapolation of these results to those obtained from human patients could be important in assessing the degree of striatal dopamine loss shown by humans with parkinsonian syndrome at the moment of clinical diagnosis. No significant differences were found between the other monoamine metabolites analyzed and free amino acid content in human and rat CSF.

1-Methyl-4-phenylpyridinium↗
PubMed1994

Stereotaxic administration of 1-methyl-4-phenylpyridinium ion (MPP+) decreases striatal fructose 2,6-bisphosphate in rats.

The stereotaxic administration of 1-methyl-4-phenylpyridinium ion (MPP+) into the neostriatum of male rats caused a lesion that resulted in a large dose-dependent loss of striatal fructose 2,6-bisphosphate; initial values were restored 5 days after the treatment. This effect was not protected by systemic administration of MK-801 or by nitroarginine. The content of hexose 6-phosphates and ATP was also reduced by MPP+ treatment, whereas lactate was increased. Biochemical and histological results suggested that MPP+ caused a nonselective cell death, followed by a pronounced astroglial response, parallel to fructose 2,6-bisphosphate recovery. The stereotaxic administration of rotenone showed a different time effect on fructose 2,6-bisphosphate cerebral content, with a significantly faster recovery. These results indicate that cerebral fructose 2,6-bisphosphate may be a sensitive metabolite related to brain damage caused by potent neurotoxins such as MPP+. On the other hand, they show that MPP+ acts in the brain through a quick, strong cytotoxic mechanism, which probably involves mechanisms other than mitochondrial chain blockage.

1-Methyl-4-phenylpyridinium↗
PubMed1994

Fructose 1,6-bisphosphate protects against D-galactosamine toxicity in isolated rat hepatocytes.

Incubation of hepatocytes with D-galactosamine (GalN) produced a dose-dependent alteration in cell viability and a fall in ATP and fructose 2,6-bisphosphate (Fru-2,6-P2) levels. The reduction in Fru-2,6-P2 can be explained by changes in the substrates or modulators of 6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase, because neither the adenosine 3',5'-cyclic monophosphate level nor the activity ratio of the enzyme was modified. Microcalorimetric measurements showed that GalN produced an exothermic peak followed by a progressive decrease in heat dissipation. Simultaneous administration of GalN and fructose 1,6-bisphosphate (Fru-1,6-P2) significantly increased cell viability, and concentrations of ATP and Fru-2,6-P2 and led to stable heat production. In the presence of Fru-1,6-P2 alone, hepatocytes kept ATP and Fru-2,6-P2 levels constant, whereas they increased the oxygen uptake-to-heat output ratio. Our results suggest that GalN initiates the hepatotoxic effect by means of an energy-dissipating interaction, produced before its metabolism and presumably at the membrane level, whereas Fru-1,6-P2 protects the cells against this injury in a way that prevents the initial interaction and increases the metabolic efficiency of the cell.

Adenosine Triphosphate↗
PubMed1994

c-met mRNA overexpression in human hepatocellular carcinoma.

This study was aimed at assessing the presence of c-met overexpression in human hepatocellular carcinoma and at determining whether this feature is associated with a definite clinical or pathological characteristic. Expression of c-met was determined by Northern-blot hybridization of a specific probe (human met proto-oncogene) in 18 tumoral and nontumoral liver samples obtained in 18 cirrhotic patients with hepatocellular carcinoma submitted to surgical treatment. Eight of the 18 hepatocellular carcinomas exhibited c-met overexpression, with an increase ranging between 2-fold and 10-fold when compared by densitometry with the surrounding liver. By contrast, in the remaining 10 cases c-met expression was almost identical to that of the surrounding nontumoral liver tissue. Overexpression of c-met was not related to either the age, sex, etiology or functional status of the underlying liver disease, or to the size of the tumor, to its differentiation degree or to the presence of pseudocapsule invasion and existence of additional neoplastic nodules. These data indicate that almost half of the human hepatocellular carcinomas exhibit c-met overexpression. Nevertheless, the biological relevance of this characteristic is not known.

Blotting, Northern↗
PubMed1993

Regulation of hepatic 6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase gene expression by glucagon.

The control of hepatic 6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase gene expression by glucagon was studied. Intraperitoneal administration of glucagon rapidly decreased the fructose 2,6-bisphosphate content by phosphorylation of 6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase and diminution of its Vmax. Immunologic studies using a specific liver antibody showed that the amount of enzyme rapidly decreased. Northern blot analysis showed that the isozyme expressed is the adult liver form. The 6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase mRNA content decreased, whereas that of phosphoenolpyruvate carboxykinase increased, and that of albumin did not change. Run-on transcription assays with isolated nuclei showed inhibition in the relative transcription rate of the 6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase gene and a stimulation of phosphoenolpyruvate carboxykinase gene. The regulation of mRNA stability of 6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase by glucagon was also studied. The half-life of mRNA decreased in the presence of glucagon, suggesting that proteins modulated by a glucagon-dependent process are regulating its stability. The time course of mRNA levels correlated with the transcription inhibition of gene and destabilization of mRNA, indicating that glucagon modulates 6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase gene expression at both transcriptional and posttranscriptional levels.

Amino Acid Sequence↗
PubMed1993

Protective effect of fructose 1,6-bisphosphate against carrageenan-induced inflammation.

Administration of carrageenan (0.5 mg) to the plantar tissue of rats resulted in reversible inflammatory injury. This damage was monitored as changes in foot volume, using a plethysmometer. Administration of fructose 1,6-bisphosphate at different doses, orally or intraperitoneally, prevented the inflammatory action induced by the simultaneous injection of carrageenan in the rat paw. The effect was dose and time dependent. In contrast, fructose or fructose 6-phosphate afforded no significant protection. In order to extend the average half-life of the drug, we prepared liposomes of fructose 1,6-bisphosphate which, administered orally or intraperitoneally, showed a greater and more prolonged antiinflammatory action. The significance of these findings with respect to the mechanism of the antiinflammatory action of fructose 1,6-bisphosphate is discussed.

Administration, Oral↗