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Biomedical subjects

J Girard

Publications and source records attributed to J Girard.

At least 91 records · Page 5Linked to original sources
PubMed1997

Is leptin the link between obesity and insulin resistance?

Leptin is the product of OB gene. This 16 kDa protein is produced by mature adipocytes and is secreted in plasma. Its plasma levels are strongly correlated with adipose mass in rodents as well as in humans. Leptin inhibits food intake, reduces body weight and stimulates energy expenditure. It has been suggested that leptin could be the link between obesity and diabetes. Recent experiments in rodents have shown that leptin expression in adipocytes is also regulated at short-term by hormones and nutrients. Leptin expression increases after food intake and decreases during fasting and diabetes. Insulin and glucocorticoids increase leptin expression, whereas catecholamines, via beta-adrenergic receptors and cAMP, and long-chain fatty acids (and thiazolidinediones), via PPARy, inhibit leptin expression. Leptin is a cytokine that binds to transmembrane receptors similar to the receptors of cytokine family (type IL-6), and transmit their information inside the cell, after dimerisation. A short-form of leptin receptor (with a cytoplasmic domain of 34 amino residues) has been identified in the choroid plexus. This type of receptor should be used for leptin transport across the blood-brain barrier. Then leptin binds to a long-form of leptin receptor in the hypothalamus (with a cytoplasmic domain of 302 amino residues) and decreases the production of neuropeptide Y, a neuromediator of food intake. The long-form of leptin receptor, transmits its information via the Janus Kinases (JAK) who subsequently phosphorylate transcription factors of the STAT family. Intermediary forms of leptin receptor have been identified in other tissues: liver, heart, skeletal muscles, endocrine pancreas. The role of leptin receptors in these tissues remains obscure, but is of considerable interest. Recent studies have shown that leptin inhibits insulin secretion and have anti-insulin effects on liver and adipose tissue. If these effects are confirmed, leptin could play a role similar to TNF alpha and could participate in the insulin-resistance of obesity and type II diabetes.

Adipose Tissue↗
PubMed1996

Peptide nucleic acid characterization by MALDI-TOF mass spectrometry.

Peptide nucleic acids (PNAs) are a new class of DNA mimics in which the regular nucleobases of adenine, thymine, cytosine, and guanine are connected via a peptide-like backbone. PNA molecules retain the same Watson-Crick base pairing as regular oligonucleotides, with the added benefits of greater specificity and resistance to enzymatic digestion. While the use of PNAs has grown rapidly because of their potential applications in biotechnology, little work has been done on developing analytical procedures for characterizing them. We have found matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry to be an effective tool for PNA analysis. PNA molecules survive the MALDI process intact and are easily ionized with almost no multiply-charged species. These features allow mixtures to be easily characterized. Traditional protein matrices (e.g., sinapinic acid,2,5-dihydroxybenzoic acid, alpha-cyano-4-hydroxycinnamic acid) were found to be superior to DNA matrices (e.g., trihydroxy-acetophenone, 3-hydroxypicolinic acid, picolinic acid). In addition, the new DNA matrix 6-aza-2-thiothymine worked well. The ability of MALDI-TOF-MS to ascertain PNA purity and sequence information at low picomole levels will be important as greater quality control of PNA synthesis is needed (e.g., when PNAs are used as antisense or antigene drugs).

Base Sequence↗
PubMed1996

Azelastine protects against CA1 traumatic neuronal injury in the hippocampal slice.

Activation of NMDA receptors appears to play a important role in traumatic neuronal injury. Additionally, N-methyl-D-aspartate (NMDA) excitotoxicity may involve leukotriene production. Therefore, we investigated whether azelastine, an anti-allergic agent inhibiting the synthesis and release of leukotrienes, could protect against CA1 traumatic neuronal injury in the hippocampal slice. Fluid percussion trauma produced evidence of severe neuronal injury with CA1 antidromic population spike amplitude recovering after 95 min to only a mean 16 +/- 1 % S.E. of initial amplitude. With 15 microM azelastine treatment given after trauma for 35 min this recovery improved to 112 +/- 17%. The azelastine EC50 for this protection was 10 microM. Significant protection was also seen with azelastine application begun 15 min after trauma. Azelastine also protected the ability to induce long-term potentiation after trauma. The specific leukotriene inhibitors, MK-571 and MK-886, similarly provided significant neuroprotection. These findings suggest that CA1 traumatic neuronal injury may be mediated by leukotriene production.

Animals↗
PubMed1996

Requirement of glucose metabolism for regulation of glucose transporter type 2 (GLUT2) gene expression in liver.

Previous studies have shown that glucose increases the glucose transporter (GLUT2) mRNA expression in the liver in vivo and in vitro. Here we report an analysis of the effects of glucose metabolism on GLUT2 gene expression. GLUT2 mRNA accumulation by glucose was not due to stabilization of its transcript but rather was a direct effect on gene transcription. A proximal fragment of the 5' regulatory region of the mouse GLUT2 gene linked to a reporter gene was transiently transfected into liver GLUT2-expressing cells. Glucose stimulated reporter gene expression in these cells, suggesting that glucose-responsive elements were included within the proximal region of the promoter. A dose-dependent effect of glucose on GLUT2 expression was observed over 10 mM glucose irrespective of the hexokinase isozyme (glucokinase K(m) 16 mM; hexokinase I K(m) 0.01 mM) present in the cell type used. This suggests that the correlation between extracellular glucose and GLUT2 mRNA concentrations is simply a reflection of an activation of glucose metabolism. The mediators and the mechanism responsible for this response remain to be determined. In conclusion, glucose metabolism is required for the proper induction of the GLUT2 gene in the liver and this effect is transcriptionally regulated.

Acetylglucosamine↗
PubMed1996

Induction of the glucokinase gene by insulin in cultured neonatal rat hepatocytes. Relationship with DNase-I hypersensitive sites and functional analysis of a putative insulin-response element.

Previous, in vivo experiments have shown that an appropriate hormonal environment (high plasma insulin, low plasma glucagon) was unable to induce the accumulation of glucokinase mRNA in term fetal rat liver, whereas it was very efficient in the newly born rat. We have confirmed in the present study that insulin induced the accumulation of glucokinase mRNA in cultured hepatocytes from 1-day-old newborn rats, but not in cultured hepatocytes from 21-day-old fetuses. To identify regulatory regions of the glucokinase gene involved in the insulin response, we have scanned the glucokinase locus for DNase I hypersensitive sites in its in vivo conformation. We confirmed the presence of four liver-specific DNase I hypersensitive sites located in the 5' flanking region of the gene. Moreover, two additional hypersensitive sites, located at 2.5 kb and 3.5 kb upstream of the cap site were found but none of these new sites displayed inducibility by insulin. Finally, an increase of the sensitivity of hypersensitive site-1 and hypersensitive site-2 to DNase I correlates with the ability of insulin to induce glucokinase gene expression in cultured hepatocytes from 1-day-old rats, as observed in previous in vivo studies. This suggests that neither a prior exposure to insulin nor a simple aging of the fetal cells in the presence of the hormone in culture are instrumental for the full DNase-I hypersensitivity of the two proximal sites necessary for the neonatal response of the glucokinase gene to insulin. The proximal hypersensitive site-1, which is close to the transcription start site in the liver, does coincide with a sequence (designated IRSL) that is 80% identical to the phosphoenolpyruvate carboxykinase IRS and with a DNase-I footprint that has been identified overlapping this sequence. Nevertheless, functional analysis of this sequence suggested that it is unlikely that the insulin-response sequence like alone is sufficient to mediate the transcriptional effect of insulin on the hepatic glucokinase gene.

Animals↗
PubMed1996

Cyclic AMP and fatty acids increase carnitine palmitoyltransferase I gene transcription in cultured fetal rat hepatocytes.

In the rat, the gene for liver mitochondrial carnitine palmitoyltransferase I (CPT I), though dormant prior to birth, is rapidly activated postnatally. We sought to elucidate which hormonal and/or nutritional factors might be responsible for this induction. In cultured hepatocytes from 20-day-old rat fetus, the concentration of CPT I mRNA, which initially was very low, increased dramatically in a dose-dependent manner after exposure of the cells to dibutyryl cAMP (Bt2cAMP). Similar results were obtained when long-chain fatty acids (LCFA), but not medium-chain fatty acids, were added to the culture medium. The effects of Bt2cAMP and LCFA were antagonized by insulin, also dose dependently. In contrast, CPT II gene expression, which was already high in fetal hepatocytes, was unaffected by any of the above manipulations. Bt2cAMP stimulated CPT I gene expression even when endogenous triacylglycerol breakdown was suppressed by lysosomotropic agents suggesting that the actions of cAMP and LCFA were distinct. Moreover, half-maximal concentrations of Bt2cAMP and linoleate produced an additive effect CPT I mRNA accumulation. While linoleate and Bt2cAMP stimulated CPT I gene transcription by twofold and fourfold, respectively, the fatty acid also increased the half-life of CPT I mRNA (50%). When hepatocytes were cultured in the presence of 2-bromopalmitate, (which is readily converted by cells into its non-metabolizable CoA ester) CPT I mRNA accumulation was higher than that observed with oleate or linoleate. Similarly, the CPT I inhibitor, tetradecylglycidate, which at a concentration of 20 microM did not itself influence the CPT I mRNA level, enhanced the stimulatory effect of linoleate. The implication is that induction of the CPT I message by LCFA does not require mitochondrial metabolism of these substrates; however, formation of their CoA esters is a necessary step. Unlike linoleate, the peroxisome proliferator, clofibrate, increased both CPT I and CPT II mRNA levels and neither effect was offset by insulin. It thus appears that the mechanism of action of LCFA differs from that utilized by clofibrate, which presumably works through the peroxisome proliferator activated receptor. We conclude that the rapid increase in hepatic CPT I mRNA level that accompanies the fetal to neonatal transition in the rat is triggered by the reciprocal change in circulating insulin and LCFA concentrations, coupled with elevation of the liver content of cAMP.

Animals↗
PubMed1996

Regulation of lipogenic enzyme expression by glucose in liver and adipose tissue: a review of the potential cellular and molecular mechanisms.

Regulation of gene expression by nutrients is an important part of the mechanisms allowing mammals to adapt to their nutritional environment. This is especially true for enzymes involved in the storage of energy such as the lipogenic and glycolytic enzymes in the liver and adipose tissue. We review in the present paper the cellular and molecular mechanisms involved in the regulation of glycolytic and lipogenic enzyme gene expression by glucose. In vivo and in vitro experiments have demonstrated that FAS and ACC gene expression is upregulated by glucose in adipose tissue, FAS, ACC and L-PK expression in the liver and ACC and L-PK expression in a pancreatic beta-cell line. This regulation involves the stimulation of their transcription. In order for glucose to act as a gene inducer, it must be metabolized. In adipose tissue, insulin increases indirectly the expression of FAS and ACC by stimulating glucose metabolism through its well-known effect on glucose transport. In the liver, the action of insulin is also indirect by allowing the expression of glucokinase and hence by increasing glucose metabolism. In the liver, fructose has a potentiating effect on the stimulation of gene expression by glucose through its stimulatory effect on glucokinase activity. Several evidences suggest that glucose-6-phosphate is the signal metabolite: (i) the effect of glucose is mimicked by 2-deoxyglucose (a glucose analogue whose metabolism stops after its phosphorylation by hexokinase) in adipose tissue and beta-cell line but not in the liver in which 2-deoxyglucose-6-phosphate does not accumulate, (ii) intracellular glucose-6-phosphate concentration varies in parallel with ACC, FAS and L-PK mRNA concentrations in liver, adipose tissue and beta-cell line, (iii) in vivo, the kinetics of hexose-phosphate fits with the time-related pattern of gene induction. Glucose response elements have been characterized on three genes, L-PK, S14 (a gene which codes for a protein of unknown function but which is directly related to lipogenesis) and FAS. These glucose response elements have all in common the presence of a sequence 5'-CACGTG-3' which binds a transcription factor of the basic domain, helix-loop-helix, leucine zipper family called USF/MLTF, although the organization of the overall glucose response element probably differs from one gene to another. The mechanisms linking glucose-6-phosphate to the glucose responsive transcription complex are presently largely unknown.

Adipose Tissue↗
PubMed1996

[Apheresis tolerance and acceptability in the child weighing 30 kg or less, with the exception of infants].

Over the ten past years, we performed 336 apheresis among 51 children who were 19 months to 15 years old (10 to 30 kg body weight). 3 types of apheresis were carried out. 14 red blood cell exchange, 293 plasma exchanges and 29 peripheral blood stem cell collections (CSP). 5 different types of continuous or discontinuous flow machines have been used. Technical adaptations depending on patient blood volume, hematocrit, type of machine used and apheresis performed permitted us to obtain a very good tolerance and acceptability. According us, the apheresis should be used each time this treatment is needed in the child, because of the very low frequency of side effects.

Adolescent↗
PubMed1996

Effects of growth hormone and IGF-I on glucocorticoid-induced protein catabolism in humans.

The effects of similar increases in total insulin-like growth factor I (IGF-I) plasma concentrations achieved by either recombinant human (rh) growth hormone (GH) or rhIGF-I administration on whole body protein and glucose kinetics were assessed. Twenty-six healthy subjects received methylprednisolone (0.5 mg.kg-1.day-1 orally) during 6 days in combination with either placebo (saline sc), GH (0.3 mg.kg-1.day-1 sc), or IGF-I (80 micrograms.kg-1.day-1 sc) in a double-blind randomized fashion. Glucocorticoid administration resulted in protein catabolism as indicated by an increase in leucine flux and a 62 +/- 13% increase in leucine oxidation ([1-13C]leucine infusion technique); this increase was abolished by GH (-1 +/- 18%) as was statistically insignificant during IGF-I treatment (+53 +/- 25%). GH increased endogenous glucose production by 28 +/- 8%, augmented glucocorticoid-induced insulin resistance of peripheral glucose clearance (euglycemic clamp), and increased circulating lipids. IGF-I administration resulted in both increased endogenous glucose production and increased peripheral glucose clearance such that plasma glucose concentrations remained unchanged by IGF-I. IGF-I lowered circulating GH and insulin and altered IGF binding proteins, which all may have reduced bioactivity of IGF-I. The data demonstrate that, in spite of similar total IGF-I plasma concentrations during treatment, GH and IGF-I exert markedly different effects on whole body leucine, glucose, and lipid metabolism.

Adult↗
PubMed1996

Normalization of insulin secretion by a selective alpha 2-adrenoceptor antagonist restores GLUT-4 glucose transporter expression in adipose tissue of type II diabetic rats.

Noninsulin-dependent mellitus is characterized by the coexistence of defective insulin secretion with insulin resistance in peripheral tissues. Therapeutic objectives are, therefore, to normalize glucose-induced insulin secretion and to restore normal glucose transport into insulin-sensitive tissues. In the present study we evaluate the effects of acute and subchronic administration (2 or 10 days) of the alpha 2-adrenoceptor antagonist SL 84.0418 on glucose tolerance in nondiabetic control rats and type I and type II diabetic rats and the level of the insulin-sensitive glucose transporter GLUT-4, which is exclusively expressed in white and brown adipose tissues, heart, and skeletal muscles. Glucose tolerance and insulin secretion were markedly impaired in type II diabetic rats (neonatal injection of streptozotocin) and were totally corrected by an acute i.p. injection of SL 84.0418. As a consequence of the chronic restoration of insulin secretion, GLUT-4 messenger RNA (mRNA) levels, initially decreased by 67% in white adipose tissue of type II diabetic rats, were normalized by subchronic (10 days), but not acute (2 days), treatment with SL 84.0418. The same results were obtained in brown adipose tissues of type II diabetic rats, whereas no modification of GLUT-4 mRNA levels remained very low in brown adipose tissues of type I diabetic rats (adult injection of streptozotocin) after acute or subchronic administration of SL 84.0418, suggesting that this drug acted by the restoration of insulin secretion. This study reports a decrease in GLUT-4 levels in insulin-sensitive tissues in this model of type II diabetes as well as its regulation after subchronic normalization of insulin secretion. We suggest a direct role for the alpha 2-adrenoceptor antagonist SL 84.0418 in pancreatic beta-cells that allows normalization of glucose tolerance.

Adipose Tissue↗
PubMed1996

Improvement of insulin action in diabetic transgenic mice selectively overexpressing GLUT4 in skeletal muscle.

To investigate the role of glucose transporter expression in whole-body glucose homeostasis, we have created transgenic mice that have a 2.0- to 3.5-fold increase in GLUT4 glucose transporter level in skeletal muscle and heart. This increase is sufficient to significantly improve insulin action and to reduce basal blood glucose levels in transgenic streptozotocin-induced diabetic mice. These results provide the first evidence of a direct causality between skeletal muscle GLUT4 transporter level and overall insulin responsiveness.

Adipose Tissue↗
PubMed1996

Troglitazone inhibits fatty acid oxidation and esterification, and gluconeogenesis in isolated hepatocytes from starved rats.

The effects of troglitazone and pioglitazone on glucose and fatty acid metabolism were studied in hepatocytes isolated from 24-h-starved rats. These thiazolidinediones inhibited long-chain fatty acid (oleate) oxidation and produced a very oxidized mitochondrial redox state. By contrast, thiazolidinediones did not affect the rate of medium-chain fatty acid (octanoate) oxidation or the activity of mitochondrial carnitine palmitoyltransferase (CPT) I. Thiazolidinediones inhibited selectively triglyceride synthesis but not phospholipid synthesis. The combined inhibition of oleate oxidation and esterification by troglitazone was due to a noncompetitive inhibition of mitochondrial and microsomal long-chain acyl-CoA synthetase (ACS) activities. It was suggested that troglitazone must be metabolized into its sulfo-conjugate derivative in liver cells to inhibit mitochondrial and microsomal ACS activities. Thiazolidinediones inhibited glucose production from lactate/pyruvate or from alanine. Analysis of gluconeogenic metabolite concentrations suggested that troglitazone would inhibit gluconeogenesis at the level of pyruvate carboxylase and glyceraldehyde-3-phosphate dehydrogenase reactions. It was concluded that 1) at a similar concentration, troglitazone was more efficient than pioglitazone to inhibit fatty acid metabolism and gluconeogenesis and 2) the inhibition of gluconeogenesis by troglitazone could be the result of the inhibition of long-chain fatty acid oxidation (decrease in acetyl-CoA, NADH-to-NAD+, and ATP-to-ADP ratios).

Animals↗
PubMed1995

Pretranslational regulation of pyruvate dehydrogenase complex subunits in white adipose tissue during the suckling-weaning transition in the rat.

Total pyruvate dehydrogenase complex activity is low in white adipose tissue during the suckling period and increases markedly at weaning on to a high-carbohydrate diet. This is concomitant with an increase in the E1 alpha, E1 beta and E2 subunit protein concentration and their respective mRNAs, suggesting a pretranslational control of this phenomenon. The most marked change is seen for the E1 alpha subunit (17-fold increase in protein concentration). The changes in pyruvate dehydrogenase complex activity and subunit abundance induced by weaning on to a high-carbohydrate diet are precluded if the animals are weaned on to a high-fat diet, suggesting that the nutritional and/or related hormonal changes rather than a developmental stage are responsible for the observed adipose-tissue pyruvate dehydrogenase complex pattern.

Adipose Tissue↗
PubMed1995

Transient increase in obese gene expression after food intake or insulin administration.

Obesity is a disorder of energy balance, indicating a chronic disequilibrium between energy intake and expenditure. Recently, the mouse ob gene, and subsequently its human and rat homologues, have been cloned. The ob gene product, leptin, is expressed exclusively in adipose tissue, and appears to be a signalling factor regulating body-weight homeostasis and energy balance. Because the level of ob gene expression might indicate the size of the adipose depot, we suggest that it is regulated by factors modulating adipose tissue size. Here we show that ob gene exhibits diurnal variation, increasing during the night, after rats start eating. This variation was linked to changes in food intake, as fasting prevented the cyclic variation and decreased ob messenger RNA. Furthermore, refeeding fasted rats restored ob mRNA within 4 hours to levels of fed animals. A single insulin injection in fasted animals increased ob mRNA to levels of fed controls. Experiments to control glucose and insulin independently in animals, and studies in primary adipocytes, showed that insulin regulates ob gene expression directly in rats, regardless of its glucose-lowering effects. Whereas the ob gene product, leptin, has been shown to reduce food intake and increase energy expenditure, our data demonstrate that ob gene expression is increased after food ingestion in rats, perhaps through a direct action of insulin on the adipocyte.

Adipose Tissue↗
PubMed1995

Cloning and characterization of the mouse glucokinase gene locus and identification of distal liver-specific DNase I hypersensitive sites.

We cloned and characterized an 83-kb fragment of mouse genomic DNA containing the entire glucokinase (GK) gene. The 11 exons of the gene span a total distance of 49 kb, with exons 1 beta and 1L being separated by 35 kb. A total of 25,266 bp of DNA sequence information was determined: from approximately -9.2 to approximately +15 kb (24,195 bp), relative to the hepatocyte transcription start site, and from -335 to +736 bp (1071 bp), relative to the transcription start site in beta cells. These sequences revealed that mouse GK is > 94% identical to rat and human GK. Mouse hepatic GK mRNA is regulated by fasting and refeeding, as also occurs in the rat. Alignment of the upstream and downstream promoter regions of the mouse, rat, and human genes revealed several evolutionarily conserved regions that may contribute to transcriptional regulation. However, fusion gene studies in transgenic mice indicate that the conserved regions near the transcription start site in hepatocytes are themselves not sufficient for position-independent expression in liver. Analysis of the chromatin structure of a 48-kb region of the mouse gene using DNase I revealed eight liver-specific hypersensitive sites whose locations ranged from 0.1 to 36 kb upstream of the liver transcription start site. The availability of a single, contiguous DNA fragment containing the entire mouse GK gene should allow further studies of cell-specific expression of GK to be performed.

Animals↗