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S Pierre

Publications and source records attributed to S Pierre.

At least 19 recordsLinked to original sources
PubMed2001

Protein associated with Myc (PAM) is a potent inhibitor of adenylyl cyclases.

Using the yeast two-hybrid assay and the second of the two large cytosolic domains of type V adenylyl cyclase (ACV) as bait, we identified a small region (amino acids 1028-1231) in the protein associated with Myc (PAM) as an interaction site for ACV. This small region of PAM as well as purified full-length PAM inhibited the activity of ACV. Additionally, full-length PAM was a very potent inhibitor of ACI and AC activities in S49 cyc(-) cells and HeLa cells with IC(50) values in the pm and low nm range. Moreover, the regulator of chromatin condensation 1-like domain of PAM (amino acids 446-1062) was sufficient and as potent as full-length PAM at inhibiting the activity of ACV. Interestingly, full-length PAM did not inhibit ACII activity that was stimulated by either forskolin of Galpha(s). When endogenous levels of PAM in HeLa cells were decreased using antisense oligodeoxynucleotides, the basal cAMP content was elevated, and the dose-response curve for vasoactive intestinal peptide-elicited cAMP accumulation in HeLa cells was shifted to the left. Therefore, we conclude that PAM is a very potent, novel inhibitor of specific isoforms of AC. Furthermore, the regulator of chromatin condensation 1-like domain of PAM is sufficient to exert the effects of the full-length protein on AC and decreases in endogenous PAM levels in HeLa cells can modulate both basal and agonist stimulated cAMP accumulation.

Adaptor Proteins, Signal Transducing↗
PubMed2001

Molecular biological approaches to unravel adenylyl cyclase signaling and function.

Signal transduction through the cell membrane requires the participation of one or more plasma membrane proteins. For many transmembrane signaling events adenylyl cyclases (ACs) are the final effector enzymes which integrate and interpret divergent signals from different pathways. The enzymatic activity of adenylyl cyclases is stimulated or inhibited in response to the activation of a large number of receptors in virtually all cells of the human body. To date, ten different mammalian isoforms of adenylyl cyclase (AC) have been cloned and characterized. Each isoform has its own distinct tissue distribution and regulatory properties, providing possibilities for different cells to respond diversely to similar stimuli. The product of the enzymatic reaction catalyzed by ACs, cyclic AMP (cAMP) has been shown to play a crucial role for a variety of fundamental physiological cell functions ranging from cell growth and differentiation, to transcriptional regulation and apoptosis. In the past, investigations into the regulatory mechanisms of ACs were limited by difficulties associated with their purification and the availability of the proteins in any significant amount. Moreover, nearly every cell expresses several AC isoforms. Therefore, it was difficult to perform biochemical characterization of the different AC isoforms and nearly impossible to assess the physiological roles of the individual isoforms in intact cells, tissue or organisms. Recently, however, different molecular biological approaches have permitted several breakthroughs in the study of ACs. Recombinant technologies have allowed biochemical analysis of adenylyl cyclases in-vitro and the development of transgenic animals as well as knock-out mice have yielded new insights in the physiological role of some AC isoforms. In this review, we will focus mainly on the most novel approaches and concepts, which have delineated the mechanisms regulating AC and unravelled novel functions for this enzyme.

Adenylyl Cyclases↗
PubMed2001

High-altitude pulmonary edema is initially caused by an increase in capillary pressure.

BACKGROUND: High-altitude pulmonary edema (HAPE) is characterized by severe pulmonary hypertension and bronchoalveolar lavage fluid changes indicative of inflammation. It is not known, however, whether the primary event is an increase in pressure or an increase in permeability of the pulmonary capillaries. METHODS AND RESULTS: We studied pulmonary hemodynamics, including capillary pressure determined by the occlusion method, and capillary permeability evaluated by the pulmonary transvascular escape of 67Ga-labeled transferrin, in 16 subjects with a previous HAPE and in 14 control subjects, first at low altitude (490 m) and then within the first 48 hours of ascent to a high-altitude laboratory (4559 m). The HAPE-susceptible subjects, compared with the control subjects, had an enhanced pulmonary vasoreactivity to inspiratory hypoxia at low altitude and higher mean pulmonary artery pressures (37 +/- 2 versus 26 +/- 1 mmHg, P<0.001) and pulmonary capillary pressures (19 +/- 1 versus 13 +/- 1 mmHg, P < 0.001) at high altitude. Nine of the susceptible subjects developed HAPE. All of them had a pulmonary capillary pressure >19 mm Hg (range 20 to 26 mmHg), whereas all 7 susceptible subjects without HAPE had a pulmonary capillary pressure < 19 mm Hg (range 14 to 18 mm Hg). The pulmonary transcapillary escape of radiolabeled transferrin increased slightly from low to high altitude in the HAPE-susceptible subjects but remained within the limits of normal and did not differ significantly from the control subjects. CONCLUSIONS: HAPE is initially caused by an increase in pulmonary capillary pressure.

Adult↗
PubMed2001

The C terminus of sprouty is important for modulation of cellular migration and proliferation.

The Drosophila Sprouty (SPRY) protein has been shown to inhibit the actions of epidermal growth factor and fibroblast growth factor. However, the role of mammalian SPRY proteins has not been clearly elucidated. We postulated that human Sprouty2 (hSPRY2) is an inhibitor of cellular migration and proliferation. Indeed, using stably transfected HeLa cells, which expressed hemagglutinin (HA)-tagged hSPRY2 or hSPRY2 tagged at the C terminus with red fluorescent protein, we demonstrated that hSPRY2 inhibits the migration of cells in response to serum, epidermal growth factor, fibroblast growth factor, and platelet-derived growth factor. Additionally, hSPRY2 also inhibited the growth of HeLa cells in response to serum. Previously, two C-terminal domains on hSPRY2, which are necessary for its colocalization with microtubules (residues 123-177) or translocation to membrane ruffles (residues 178-194), have been identified (Lim, J., Wong, E. S., Ong, S. H., Yusoff, P., Low, B. C., and Guy, G. R. (2000) J. Biol. Chem. 275, 32837-32845). Therefore, using TAT-tagged hSPRY2 and its mutants, we determined the role of these two C-terminal domains in the inhibition of cell migration and proliferation. Our data show that the deletion of either of these two regions in hSPRY2 abrogates its ability to modulate cell migration in response to different growth factors and proliferation in response to serum. Therefore, we conclude that hSPRY2 inhibits the actions of a number of growth factors, and its C terminus, which is homologous among various SPRY isoforms, is important in mediating its biological activity.

Amino Acid Sequence↗
PubMed2001

Inhibition of hypoxic pulmonary vasoconstriction by carbon monoxide in dogs.

OBJECTIVE: We tested the hypothesis that carbon monoxide might participate in the modulation of hypoxic pulmonary vasoconstriction (HPV) by prostacyclin (PGI2) and nitric oxide. DESIGN: Prospective, interventional study. SETTING: University laboratory. SUBJECTS: Nineteen intact anesthetized mongrel dogs. INTERVENTIONS: Right heart catheterization for the measurements of mean pulmonary artery pressure (Ppa), left atrial pressure estimated from occluded Ppa (Ppao), pulmonary capillary pressure (Pcp) calculated from the Ppa decay curve after balloon occlusion, and cardiac output (Q); inferior vena cava balloon for the control of Q by manipulation of venous return; ventilation in hyperoxia (fraction of inspired O2, 0.4) or in hypoxia (Fio2, 0.1); inhibition of cyclooxygenase by indomethacin (Indo); inhibition of nitric oxide synthase by NG-nitro-l-arginine (L-NA); inhibition of heme oxygenase by mesoporphyrin IX (SnMP); inhalation of nitric oxide (20 ppm); and inhalation of carbon monoxide (100 ppm). MEASUREMENTS AND MAIN RESULTS: The first seven dogs were weak responders to hypoxia as assessed by a hypoxia-induced increase in the gradient between Ppa and Ppao, measured at one level of Q kept constant, by an average of only 2 mm Hg (p = NS). This HPV was markedly increased by the combined administration of Indo and L-NA. A further enhancement of HPV was observed after the addition of SnMP, leading to severe pulmonary hypertension with an average increase in Ppa to 39 mm Hg. Inhaled nitric oxide inhibited HPV only after the combined administration of Indo, L-NA, and SnMP. Inhaled carbon monoxide had no effect. The next 12 dogs were stronger responders to hypoxia, as assessed by a hypoxia-induced increase in the gradient between Ppa and Ppao, measured at several levels of Q, by an average of 3 mm Hg (p <.05). This HPV was of the same magnitude after administration of placebo (n = 6) or SnMP (n = 6). Addition of Indo enhanced HPV to the same extent in the placebo and in the SnMP groups. Addition of L-NA induced a further enhancement of HPV, which was, however, greater in the SnMP group. There was a slight increase in the capillary-venous segment relative to the arterial segment in hypoxic conditions, but the partitioning of pulmonary vascular resistance was otherwise unaffected by nitric oxide, carbon monoxide, or PGI2. CONCLUSIONS: Endogenous carbon monoxide modulates canine HPV only in the absence of nitric oxide. The vasodilation mediated by nitric oxide, PGI2, or carbon monoxide is essentially distributed between proximal and distal sites proportionally to the degree of constriction produced during hypoxia.

Administration, Inhalation↗
PubMed2001

RT-PCR detection of the Na,K-ATPase beta3-isoform in human heart.

The Na,K-ATPase is a heterodimer composed of an alpha-catalytic and a beta-glycoprotein subunit. At present, three different alpha-polypeptides (alpha1, alpha2, alpha3) and two distinct beta-isoforms (beta1 and beta2) have been detected in human heart. The aim of the present study was to determine whether or not the beta3-isoform of the Na,K-ATPase can be detected in human heart. Using the highly sensitive method of RT-PCR, we here show that human heart expresses the beta3-isoform of the Na,K-ATPase. Given the differences in pharmacological properties of the nine different Na,K-ATPase isoenzymes (containing all combinations of the subunit isoforms), the study of beta3-isoform regulation in human heart may be of interest in understanding the altered response of human myocardium to digitalis therapy during heart failure.

Animals↗
PubMed2001

RT-PCR detection of Na,K-ATPase subunit isoforms in human umbilical vein endothelial cells (HUVEC): evidence for the presence of alpha1 and beta3.

The endothelial Na,K-ATPase is an active component in maintaining a variety of normal vascular functions. The enzyme is characterized by a complex molecular heterogeneity that results from differential expression and association of multiple isoforms of both its alpha- and beta-subunits. The aim of the present study was to determine which isoforms of the Na,K-ATPase are expressed in human endothelial cells. HUVEC (human umbilical vein endothelial cells) were used as a model of well known human endothelial cells. The high sensitive method RT-PCR was used with primers specific for the various isoforms of the alpha- and beta-subunits of the Na,K-ATPase. The results show that HUVEC express alpha1-, but not alpha2-, alpha3- or alpha4-isoforms of the catalytic subunit and that beta3- but not beta2- or beta1-isoforms is present in these cells. These findings are in contradiction with our previous detection of Na,K-ATPase isoforms in HUVEC using antibodies (14). Such results raise the technical problem of the specificity of the available antibodies directed against the different isoforms as well as the question of the physiological relevance of the diversity of the Na,K-ATPase isoforms.

Endothelium, Vascular↗
PubMed2000

Effect of streptozotocin-induced diabetes on rat liver Na+/K+-ATPase.

Na+/K+-ATPase during diabetes may be regulated by synthesis of its alpha and beta subunits and by changes in membrane fluidity and lipid composition. As these mechanisms were unknown in liver, we studied in rats the effect of streptozotocin-induced diabetes on liver Na+/K+-ATPase. We then evaluated whether fish oil treatment prevented the diabetes-induced changes. Diabetes mellitus induced an increased Na+/K+-ATPase activity and an enhanced expression of the beta1 subunit; there was no change in the amount of the alpha1 and beta3 isoenzymes. Biphasic ouabain inhibition curves were obtained for diabetic groups indicating the presence of low and high affinity sites. No alpha2 and alpha3 isoenzymes could be detected. Diabetes mellitus led to a decrease in membrane fluidity and a change in membrane lipid composition. The diabetes-induced changes are not prevented by fish oil treatment. The results suggest that the increase of Na+/K+-ATPase activity can be associated with the enhanced expression of the beta1 subunit in the diabetic state, but cannot be attributed to changes in membrane fluidity as typically this enzyme will increase in response to an enhancement of membrane fluidity. The presence of a high-affinity site for ouabain (IC50 = 10-7 M) could be explained by the presence of (alphabeta)2 diprotomeric structure of Na+/K+-ATPase or an as yet unknown alpha subunit isoform that may exist in diabetes mellitus. These stimulations might be related, in part, to the modification of fatty acid content during diabetes.

Animals↗
PubMed2000

Specific up-regulation of mitochondrial F0F1-ATPase activity after short episodes of atrial fibrillation in sheep.

INTRODUCTION: Ventricular fibrillation induced by either digitalis intoxication or electrical stimulation is reported to alter myocardial energy by impairing the sarcolemmal Na,K-ATPase or the receptor for digitalis and the mitochondrial ATPase synthase or F0F1-ATPase. However, little is known about these membrane functions during atrial fibrillation (AF). METHODS AND RESULTS: We analyzed the effects of electrically induced AF on biochemical activities of atrial F0F1-ATPase and Na,K-ATPase in sheep. A group of six sheep was subjected to direct short electrical stimulation of the right atrium to induce AF. Sham-operated sheep served as a control group. Microsomal and mitochondrial membranes of atrial muscle were isolated and tested for enzymatic activity. All paced sheep developed multiple episodes of sustained AF, with a mean total duration of 110 minutes over a 2-hour period. Data showed that short-term pacing-induced AF significantly activated membrane F0F1-ATPase activity (P < 0.05) without changes in cytochrome-c oxidase activity, Na,K-ATPase activity, ouabain sensitivity, and alpha1-subunit expression. CONCLUSION: Specific activation of F0F1-ATPase activity is an early molecular consequence of sustained AF in sheep.

Animals↗
PubMed2000

Inhibition of Na,K-ATPase by external electrical cardioversion in a sheep model of atrial fibrillation.

INTRODUCTION: Electrical external cardioversion commonly used to treat atrial fibrillation (AF) is associated with myocardial membrane damage and disturbances in ionic homeostasis (hemodynamically unstable). The present study was designed to investigate whether alterations in ionic homeostasis observed were due in part to changes in the myocardial activity of Na,K-ATPase. METHODS AND RESULTS: AF was induced by pacing in ten anesthetized sheep divided into two groups. Group I (n = 4) received a single external countershock of 360 J after three episodes of AF lasting 10 minutes. Group II (n = 6) served as controls. Activity, responsiveness to ouabain, and membrane expression of catalytic alpha and beta subunits of Na,K-ATPase in sarcolemmal myocardial membrane fractions were investigated. Membrane fluidity and fatty acid composition, and plasma levels of atrial natriuretic factor (ANF) also were measured. One shock after episodes of AF significantly decreased ventricular Na,K-ATPase activity up to 50% (P < 0.001) without modification of atrial activity at the membrane level. Sites with low affinity to ouabain showed a fivefold lower affinity for ouabain in the cardioversion group than in the control group (IC50 = 7.9 micromol/L vs 40 micromol/L ouabain, P < 0.05). Plasma levels of ANF were significantly increased in the cardioversion group compared with the control group. These changes were independent of membrane modulation in terms of expression of Na,K-ATPase, membrane fluidity, and fatty acid composition. CONCLUSION: This study suggests that left ventricular perturbation of ionic homeostasis subsequent to transthoracic cardioversion could result from inactivation of Na,K-ATPase activity.

Animals↗
PubMed2000

Echocardiographic and invasive measurements of pulmonary artery pressure correlate closely at high altitude.

Exaggerated hypoxia-induced pulmonary hypertension is a hallmark of high-altitude pulmonary edema (HAPE) and plays a major role in its pathogenesis. Many studies of HAPE have estimated systolic pulmonary arterial pressure (SPAP) with Doppler echocardiography. Whereas at low altitude, Doppler echocardiographic estimation of SPAP correlates closely with its invasive measurement, no such evidence exists for estimations obtained at high altitude, where alterations of blood viscosity may invalidate the simplified Bernoulli equation. We measured SPAP by Doppler echocardiography and invasively in 14 mountaineers prone to HAPE and in 14 mountaineers resistant to this condition at 4,559 m. Mountaineers prone to HAPE had more pronounced pulmonary hypertension (57 +/- 12 and 58 +/- 10 mmHg for noninvasive and invasive determination, respectively; means +/- SD) than subjects resistant to HAPE (37 +/- 8 and 37 +/- 6 mmHg, respectively), and the values measured in the two groups as a whole covered a wide range of pulmonary arterial pressures (30-83 mmHg). Spearman test showed a highly significant correlation (r = 0.89, P < 0.0001) between estimated and invasively measured SPAP values. The mean difference between invasively measured and Doppler-estimated SPAP was 0.5 +/- 8 mmHg. At high altitude, estimation of SPAP by Doppler echocardiography is an accurate and reproducible method that correlates closely with its invasive measurement.

Adult↗
PubMed1999

Ginkgo biloba extract (EGb 761) protects Na,K-ATPase activity during cerebral ischemia in mice.

Neuroprotective drugs such as Ginkgo biloba extract (EGb 761) could prevent the ischemia-induced impairment of the Na,K-ATPase activity. In this study, Na,K-ATPase activity and expression, contents in fatty acids and malondialdehyde, an index of lipoperoxidation, were compared in the ipsilateral (ischemic) and the contralateral (unlesioned) cortices after 1 h of unilateral focal cortices cerebral ischemia in the mouse. EGb 761 (110 mg/kg) was administered daily to half of the animals for 10 days before ischemia. Ischemia significantly reduced Na,K-ATPase activity by about 40% and increased malondialdehyde content; EGb 761 pretreatment abolished these effects. The free radical scavenger properties of EGb 761 are a potential mechanism by which Na,K-ATPase injury and lipoperoxidation are prevented.

Animals↗
PubMed1999

Localization of Na,K-ATPase alpha/beta isoforms in rat sciatic nerves: effect of diabetes and fish oil treatment.

The localization of the Na,K-ATPase isoenzymes in sciatic nerve remains controversial, as well as diabetes-induced changes in Na,K-ATPase isoforms. Some of these changes could be prevented by fish oil therapy. The aim of this study was to determine by confocal microscopy the distribution of Na,K-ATPase isoforms (alpha1, alpha2, alpha3, beta1, and beta2) in the sciatic nerve, the changes induced by diabetes, and the preventive effect of fish oil in diabetic neuropathy. This study was performed in three groups of rats. In the first two groups, diabetes was induced by streptozotocin and rats were supplemented daily with fish oil or olive oil at a dosage of 0.5 g/kg of body weight. The third one was a control group that was supplemented with olive oil. Five antibodies against specific epitopes of Na,K-ATPase isoenzymes were applied to stained dissociated nerve fibers with fluorescent secondary antibodies. The five isoenzymes were documented in nonspecific regions, Schwann cells (myelin), and the node of Ranvier. The localization of the alpha1, alpha2, and beta1 isoenzymes was not affected by diabetes. In contrast, diabetes induced a decrease of the alpha2 subunit (p < 0.05) and an up-regulation of the beta2 subunit (p < 0.05). These modifications were noted in both regions for alpha2 and were localized at the myelin domain only for the beta2. Fish oil supplementation prevented the diabetes-induced changes in the alpha2 subunit with an additional up-regulation. The beta2 subunit was not modified. A phenotypic change similar to nerve injury was induced by diabetes. Fish oil supplementation partially prevented some of these changes.

Animals↗
PubMed1999

A quantitative immunocytochemical study of Na+,K+-ATPase in rat hepatocytes after STZ-induced diabetes and dietary fish oil supplementation.

Because diabetes causes alterations in hepatic membrane fatty acid content, these changes may affect the Na+,K+-ATPase. In this study we documented the effects of streptozotocin (STZ)-induced diabetes on hepatic Na+,K+-ATPase catalytic alpha1-subunit and evaluated whether these changes could be normalized by fish oil supplementation. Two groups of diabetic rats received fish oil or olive oil supplementation. Both groups had a respective control group. We studied the localization of catalytic alpha1-subunit on bile canalicular and basolateral membranes using immunocytochemical methods and confocal laser scanning microscopy, and the Na+, K+-ATPase activity, membrane fluidity, and fatty acid composition on isolated hepatic membranes. A decrease in the alpha1-subunit was observed with diabetes in the bile canalicular membranes, without changes in basolateral membranes. This decrease was partially prevented by dietary fish oil. Diabetes induces significant changes as documented by enzymatic Na+,K+-ATPase activity, membrane fluidity, and fatty acid content, whereas little change in these parameters was observed after a fish oil diet. In conclusion, STZ-induced diabetes appears to modify bile canalicular membrane integrity and dietary fish oil partly prevents the diabetes-induced alterations.

Animals↗
PubMed1999

Respectability.

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Career Mobility↗
PubMed1996

A MGF/STAT5 binding site is necessary in the distal enhancer for high prolactin induction of transfected rabbit alpha s1-casein-CAT gene transcription.

The rabbit alphas1-casein gene contains a distal prolactin-dependent enhancer 3442-3285 bp 5' to the site of initiation of transcription. We have reported previously that four DNA/protein-binding sites (F1-F4) are located within this distal enhancer. We now show that one of this binding site (the F4 site) binds in vitro a MGF/STAT5-like factor. The functional importance of the F4 site was estimated by cotransfection of CHO cells with a chimeric gene containing or not the F4 sequence linked to the (-391/+1774)CAT gene and a plasmid encoding the rabbit mammary prolactin receptor. The F4 site is necessary for maximal response, of the enhancer to prolactin. However, this site has to be associated to the Fl-F3 fragment. It can be replaced by a genuine MGF/STAT5-binding site. A mutational analysis indicates that F4 and F1 sites are simultaneously involved to confer a high prolactin sensitivity.

Animals↗
PubMed1996

Effects of carbon dioxide embolism with nitrous oxide in the inspired gas in piglets.

We have compared cardiorespiratory variables in anaesthetized piglets whose lungs were ventilated with oxygen in nitrous oxide (N2O group) or nitrogen (N group) after right ventricular carbon dioxide boluses (0.5 or 1 ml kg-1; n = 12) or slow graded injections (n = 6). Boluses affected all variables studied significantly (P < 0.05) except mean systolic arterial pressure. Significant changes in PE'CO2 (P = 0.012) and PaO2 (P = 0.048) values were observed in the N2O group. Changes in PaCO2 were related to volumes of injected carbon dioxide (P = 0.044). Boluses of 1.0 ml kg-1 induced severe circulatory collapse in two piglets in the N2O group. Slow embolization altered respiratory variables significantly (P < 0.001)). PaO2 decreased significantly in the N2O group (P < 0.0001). Mean pulmonary arterial pressure increased significantly over time (P = 0.001) and lasted longer in the N2O group (P < 0.05). Volumes and time required to induce a 50% increase in mean pulmonary arterial pressure differed significantly between groups (P < 0.05). We conclude that nitrous oxide worsened the effects of rapid and slow carbon dioxide emboli on cardiopulmonary variables. Rapid carbon dioxide embolism altered respiratory and haemodynamic variables, while slow carbon dioxide embolism changed only respiratory variables.

Anesthesia, General↗