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Bertrand Perret

Publications and source records attributed to Bertrand Perret.

30 records · Page 2Linked to original sources
PubMed2003

Modulation of phosphoinositide 3-kinase activation by cholesterol level suggests a novel positive role for lipid rafts in lysophosphatidic acid signalling.

Methyl-beta-cyclodextrin (MbetaCD) was used to explore a role for cholesterol-enriched plasma membrane microdomains in coupling lysophosphatidic acid (LPA) stimulation to phosphoinositide 3-kinase (PI3K) activation. Cholesterol depletion strongly inhibited the production of phosphatidylinositol 3,4-bisphosphate and phosphatidylinositol 3,4,5-trisphosphate in Vero cells stimulated with LPA. In agreement, the phosphorylation of Akt/protein kinase B, but not of Erk kinases, was suppressed by MbetaCD. MbetaCD did not interfere with the overall phospholipid metabolism, and its effects were reversed in cholesterol add-back experiments. Finally, PI3K was detected in lipid rafts prepared from control but not MbetaCD-treated cells, suggesting that these microdomains contribute to LPA signalling by compartmentalising component(s) of the PI3K pathway.

Animals↗
PubMed2003

Ectopic beta-chain of ATP synthase is an apolipoprotein A-I receptor in hepatic HDL endocytosis.

The effect of high-density lipoprotein (HDL) in protecting against atherosclerosis is usually attributed to its role in 'reverse cholesterol transport'. In this process, HDL particles mediate the efflux and the transport of cholesterol from peripheral cells to the liver for further metabolism and bile excretion. Thus, cell-surface receptors for HDL on hepatocytes are chief partners in the regulation of cholesterol homeostasis. A high-affinity HDL receptor for apolipoprotein A-I (apoA-I) was previously identified on the surface of hepatocytes. Here we show that this receptor is identical to the beta-chain of ATP synthase, a principal protein complex of the mitochondrial inner membrane. Different experimental approaches confirm this ectopic localization of components of the ATP synthase complex and the presence of ATP hydrolase activity at the hepatocyte cell surface. Receptor stimulation by apoA-I triggers the endocytosis of holo-HDL particles (protein plus lipid) by a mechanism that depends strictly on the generation of ADP. We confirm this effect on endocytosis in perfused rat liver ex vivo by using a specific inhibitor of ATP synthase. Thus, membrane-bound ATP synthase has a previously unsuspected role in modulating the concentrations of extracellular ADP and is regulated by a principal plasma apolipoprotein.

Adenosine Diphosphate↗
PubMed2003

Obesity and alcohol modulate the effect of apolipoprotein E polymorphism on lipids and insulin.

OBJECTIVE: To assess the interaction between apolipoprotein (apo) E polymorphism, alcohol consumption, and BMI on insulin, lipid, and lipoprotein levels in men. RESEARCH METHODS AND PROCEDURES: Cross-sectional study of 266 healthy men without hypolipidemic or antidiabetic drug treatment. BMI, apo E polymorphisms, insulin, and lipid and lipoprotein levels were assessed. Alcohol consumption was assessed by questionnaire. epsilon2/epsilon4 carriers were excluded from the analysis. RESULTS: On bivariate analysis, epsilon2 carriers had lower levels of total and low-density lipoprotein cholesterol and higher levels of apo E and lipoparticle B:E than epsilon3 carriers, the opposite being found for epsilon4 carriers compared with epsilon3 carriers; epsilon4 carriers also had significantly higher insulin levels. On multivariate analysis, significant interactions (p < 0.04) between apo E alleles and increased BMI were found for total and low-density lipoprotein cholesterol and insulin levels, the increase in those parameters with BMI being stronger among epsilon4 carriers than among epsilon3 or epsilon2 carriers. Significant interactions (p < 0.02) between apo E alleles and alcohol consumption were also found for apo B levels, which increased in epsilon2 carriers but remained relatively stable in epsilon3 and tended to decrease in epsilon4 carriers. DISCUSSION: These data suggest that effects of apo E alleles on lipids and insulin levels are partly dependent on environmental variables such as BMI and alcohol intake. These findings highlight the importance of gene x environment interactions on the deleterious effect of obesity on cardiovascular risk factors.

Adult↗
PubMed2002

Phosphoinositide 3-kinase C2alpha is activated upon smooth muscle cell migration and regulated by alpha(v)beta(3) integrin engagement.

The involvement of phosphoinositide 3-kinase C2alpha in vascular smooth muscle cell migration was investigated. Products of phosphoinositide 3-kinase, phosphatidylinositol-3-phosphate, and phosphatidylinositol-3,4-bis-phosphate were increased upon smooth muscle cell migration but their synthesis was affected only partially by phosphoinositide 3-kinase inhibitors, wortmannin and LY-294002. Using specific antibody, we showed that the wortmannin/LY-294002 poorly sensitive phosphoinositide 3-kinase C2alpha is expressed in smooth muscle cells. Measurement of phosphoinositide 3-kinase C2alpha activity in vitro, after immunoprecipitation, clearly demonstrated its activation upon smooth muscle cell migration. Moreover, for the first time, phosphoinositide 3-kinase C2alpha was found to be differentially regulated by alpha(v)beta(3) and alpha(v)beta(5) integrin engagement. Finally, we have identified two new potential phosphoinositide 3-kinase C2alpha-binding proteins, p70 and p110, which both may be tyrosine phosphorylated. Thus, phosphoinositide 3-kinase C2alpha might represent a new regulatory pathway of cell migration downstream of integrin engagement.

Androstadienes↗
PubMed2002

Guinea pig phospholipase B, identification of the catalytic serine and the proregion involved in its processing and enzymatic activity.

Guinea pig phospholipase B (GPPLB) is a glycosylated ectoenzyme of intestinal brush border membrane. It displays a broad substrate specificity and is activated by trypsin cleavage. The primary sequence contains four tandem repeat domains (I to IV) and several serines in lipase consensus sequences. We used site-directed mutagenesis to demonstrate that only the serine 399 present in repeat II is responsible for the various enzymatic activities of GPPLB. Furthermore, we characterized for the first time the retinyl esterase activity of the enzyme. We also constructed and expressed in COS-7 cells, an NH(2)-terminal repeat I deletion mutant which was detected at a very low level by immunoblot. However, confocal microscopy study showed a strong intracellular accumulation with a weak membrane expression of the mutated protein, indicating a role of the NH(2)-terminal repeat I in the processing of GPPLB. Nevertheless, the Western blot-detected protein presented a glycosylation and trypsin sensitivity patterns similar to wild type PLB. The mutant is also fully active without trypsin treatment, in contrast to native enzyme. Thus, we propose a structural model for GPPLB, in which the repeat I constitutes a lid covering the active site and impairing enzymatic activity, its removal by trypsin leading to an active protein.

Animals↗
PubMed2002

Human epidermis is a novel site of phospholipase B expression.

Phospholipase B (PLB) is an enzyme that displays both phospholipase A(2) and lysophospholipase activities. Analysis of human epidermis homogenates indicated the presence of a 97 kDa PLB protein, as well as a phospholipase A(2) activity, both being enriched in the soluble fraction. Immunolabelling and in situ hybridization experiments showed that this enzyme is expressed in the different layers of epidermis with an accumulation at the dermo-epidermis junction. RT-PCR data indicated that PLB is specifically expressed in natural and reconstructed epidermis. By 3'-RACE-PCR and screening of human genome databases, we obtained a 3600 bp cDNA coding for human PLB highly homologous to already described intestinal brush border PLBs. These data led us to conclude that the soluble PLB corresponds to a proteolytic cleavage of the membrane anchored protein. Altogether, our results provide the first characterization of human PLB which should play an important role in epidermal barrier function.

Amino Acid Sequence↗
PubMed2002

Alcohol consumption is associated with enrichment of high-density lipoprotein particles in polyunsaturated lipids and increased cholesterol esterification rate.

BACKGROUND: Alcohol consumption is associated with high levels of high-density lipoproteins (HDLs). Moreover, changes in the fatty acid patterns of red blood cell phospholipids and plasma lipids have been observed in drinkers. The objectives of this study were to characterize the composition of HDL particles with respect to lipid molecular species in regular wine drinkers and to assess the functional properties of those HDLs as regards key steps of reverse cholesterol transport. METHODS: Forty-six subjects were recruited in the frame of a population study performed in Toulouse, southern France, and a nutritional investigation, including daily alcohol consumption, was performed. Subjects were sorted according to their daily alcohol intake (0, < or =35, and >35 g/day), mostly as red wine. The plasma HDL fraction was isolated, and neutral lipid molecular lipids and phospholipid fatty acids were analyzed by gas liquid chromatography. Efflux of cellular cholesterol and rates of cholesterol esterification and cholesteryl ester transfers between lipoproteins were assayed in a cell-plasma incubation system. RESULTS: Wine drinking, at 47 g/day, was associated with an increase in HDL cholesterol and apolipoprotein A-I, but not with triglycerides. Isolated HDL displayed a 27% increase in all cholesteryl ester molecular species. The particles were also enriched in unsaturated phospholipids and, particularly, in those containing arachidonic (+30%) and eicosapentaenoic (+90%) acids. The plasma cholesterol esterification rate, reflecting lecithin cholesterol acyl transferase activity on HDL, was found to be higher (+27%) in drinkers than in nondrinkers, whereas the rate of cellular cholesterol efflux to plasma was identical. CONCLUSIONS: Regular wine consumption is associated with high levels of polyunsaturated lipids in HDL and with increases in the cholesterol esterification rate.

Adult↗
PubMed2002

Coupled assay of sphingomyelin and ceramide molecular species by gas liquid chromatography.

This study reports a single-step analysis of the molecular species of endogenous ceramides and of the ceramide moiety of sphingomyelins in biological samples, using gas liquid chromatography (GLC). Silylated sphingomyelins were quantitatively converted to monosilylated ceramide upon injection into GLC, whereas the free ceramides were di-silylated on the primary and secondary alcohol function, as confirmed by mass spectrometry. The reproducible shift of the retention times between the mono- and di-silylated derivatives enables simultaneous quantification of the variety of sphingomyelin and ceramide molecular species. Overlapping diacylglycerols were first removed by a mild alkaline treatment of the lipid extract. The lowest detection limit (5 pmol) did not allow for identification of free ceramides in human plasma, but 17 molecular species of ceramides derived from sphingomyelins were quantified, from NC16:0 up to NC24:1. By contrast, three major free ceramides (NC16:0, NC24:0, and NC24:1) were quantified in HEPG2 and Chinese hamster ovary (CHO) cells. Upon induction of apoptosis in CHO cells by C6-ceramide, we could follow the disappearance of the C6-ceramide, its partial conversion to C6-sphingomyelin, and the prominent increase of NC16:0 ceramide. Thus, our method represents a unique procedure of simultaneous analysis of sphingomyelin and ceramide molecular species able to monitor the variation of the different pools in biological samples.

Acids↗
PubMed2002

Hepatic lipase: structure/function relationship, synthesis, and regulation.

Hepatic lipase (HL) is a lipolytic enzyme, synthesized by hepatocytes and found localized at the surface of liver sinusoid capillaries. In humans, the enzyme is mostly bound onto heparan-sulfate proteoglycans at the surface of hepatocytes and also of sinusoid endothelial cells. HL shares a number of functional domains with lipoprotein lipase and with other members of the lipase gene family. It is a secreted glycoprotein, and remodelling of the N-linked oligosaccharides appears to be crucial for the secretion process, rather than for the acquisition of the catalytic activity. HL is also present in adrenals and ovaries, where it might promote delivery of lipoprotein cholesterol for steroidogenesis. However, evidence of a local synthesis is still controversial. HL activity is fairly regulated according to the cell cholesterol content and to the hormonal status. Coordinate regulations have been reported for both HL and the scavenger-receptor B-I, suggesting complementary roles in cholesterol metabolism. However, genetic variants largely contribute to HL variability and their possible impact in the development of a dyslipidemic phenotype, or in a context of insulin-resistance, is discussed.

Amino Acid Sequence↗
PubMedDate not supplied

Joint use of clinical parameters, biological markers and CAGE questionnaire for the identification of heavy drinkers in a large population-based sample.

AIMS: Alcohol consumption in France is one of the highest in the world. Factors associated with excessive alcohol drinking are numerous. However, taken separately, none of the existing clinical or biological markers of excessive alcohol intake enables an adequate identification of heavy drinkers. The aim of this cross-sectional survey was to identify socio-demographic, clinical and biological factors associated with excessive alcohol drinking, to develop a model and to assess its reliability, thus enabling the detection of heavy drinkers. METHODS: Subjects were 1619 men and 1559 women, aged 35-64 years, living in three French areas (Lille, Strasbourg and Toulouse) and randomly selected from polling lists. Socio-demographic status, lifestyle, reported alcohol intake and answers to the CAGE questionnaire (alcohol dependence) were obtained by questionnaire. A blood sample was taken for quantification of biological parameters. Men who drank 60 g of ethanol a day (g/day) or above and women who drank 30 g/day or above were classified as heavy drinkers. The reference class (RC) gathered non-drinkers and moderate drinkers together. The sample was divided into two sub-samples: the first was used to estimate the parameters of a logistic regression model (heavy drinkers vs others), and the second to assess the accuracy of this model for the identification of heavy drinkers, using receiver operating characteristic (ROC) curves. A specific analysis was performed for each gender. RESULTS: Fourteen per cent of men and 40.8% of women were non-drinkers. Nine per cent of women and 14.4% of men were heavy drinkers. Wine was the most consumed alcoholic beverage. In the univariate analyses, differences were observed between the two groups of alcohol consumers for most of the socio-demographic, clinical and biological variables considered. In the multivariate analyses, low educational level, smoking, apoprotein B, high density lipoprotein cholesterol, mean corpuscular volume (MCV), gamma-glutamyl-transferase (GGT) and the CAGE score for men, and living area, age, MCV, GGT and the CAGE score for women remained independently and significantly associated with heavy drinking. In the validation sub-sample, these models combining different types of markers enabled a good discrimination between heavy drinkers and the RC, with an area under the ROC curve of 82% for men and of 79% for women. CONCLUSIONS: In this study, socio-demographic, clinical and biological factors and the CAGE score were independently related to excessive alcohol drinking and their joint utilization in a screening model enabled a good recognition of heavy drinkers.

Adult↗