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G Thibault

Publications and source records attributed to G Thibault.

At least 73 records · Page 4Linked to original sources
PubMed1995

Basal and angiotensin II-induced cytosolic free calcium in adult rat cardiomyocytes and fibroblasts after volume overload.

This study investigates basal and angiotensin II (Ang II)-induced [Ca2+]i concentrations in cells from hearts of rats that have undergone cardiac hypertrophy due to volume overload. [Ca2+]i measurements assessed by digital imaging using fura 2 methodology were performed on isolated ventricular cardiomyocytes and fibroblasts from adult rat hearts with a 4-week aortocaval shunt. Long-term aortocaval shunt induced a significant increase in atrial (72%) and ventricular (41%) weights and a large elevation in plasma atrial natriuretic peptide-(1-98) concentration (160%). For adult cardiomyocytes [Ca2+]i measurements are reported as diastolic (average of the lowest points) and systolic intracellular Ca2+ values (average of the maximum points corresponding to the diastolic points) over a 30-second time interval. Basal diastolic [Ca2+]i (99 +/- 4.1 nmol/L for experimental cells versus 90 +/- 4.8 for control cells) was not altered, whereas basal systolic [Ca2+]i was significantly greater in ventricular cardiomyocytes from overload hearts (155 +/- 2.3 versus 129 +/- 4.4 nmol/L for control cells, P < .05). Ang II increased intracellular Ca2+ spike frequency in a concentration-dependent manner in cardiomyocytes from control and overload myocardium. Basal and Ang II-induced intracellular Ca2+ spike frequencies were not modified in cardiomyocytes from hypertrophied hearts. Basal [Ca2+]i in ventricular fibroblasts from overload myocardium was significantly increased (128 +/- 5.1 nmol/L for fibroblasts from hypertrophied hearts versus 104 +/- 3.5 for control cells, P < .05). Ang II-induced [Ca2+]i was lower in fibroblasts from overload myocardium (P < .05).(ABSTRACT TRUNCATED AT 250 WORDS)

Analysis of Variance↗
PubMed1995

Localization of atrial natriuretic factor receptors in the mesenteric arterial bed. Comparison with angiotensin II and endothelin receptors.

Although receptors for atrial natriuretic factor (ANF) and angiotensin II (Ang II) have been reported in rat mesenteric arteries, both peptides induce weak biological responses. Endothelin-1 (ET-1) evokes a potent vasoconstriction in the mesenteric artery. To identify the tissue localization of ANF, Ang II, and ET-1 receptors, radioligand binding experiments with 125I-ANF, 125I-[Sar1,Ile8]Ang II, and 125I-ET-1 were performed in defatted mesenteric arteries and in the surrounding adipose tissue. 125I-ANF binding assays in adipose tissue showed a single class of high-affinity binding sites (Bmax, 420 +/- 16 fmol/mg protein; Kd, 343 +/- 16 pmol/L). In vascular membranes, most 125I-ANF binding was nonspecific. The majority of receptors present in adipose tissue recognized ANF, C-type natriuretic peptide (CNP), and des-[Gln18,Ser19,Gly20,Leu21,Gly22]ANF-(4- 23) (C-ANF) with close affinities, with C-ANF competing for > 98% of the binding sites. In adipocytes, ANF and CNP stimulated cGMP generation. cGMP production by mesenteric arteries was stimulated by sodium nitroprusside but not by ANF or CNP. Autoradiographic localization of 125I-ANF and 125I-ET-1 showed that in the case of ANF, most specific binding occurred in adipocytes, whereas for ET-1, specific binding was present in both adipose tissue and mesenteric arteries. Cross-linking of 125I-ANF followed by SDS-PAGE revealed two receptor species of 130 and 70 kD in adipose membranes and none in vascular tissue. Both were completely displaced by ANF, CNP, and C-ANF. 125I-[Sar1,Ile8]Ang II binding assays in adipose tissue exhibited a single class of binding sites (Bmax, 211 +/- 4 fmol/mg protein; Kd, 520 +/- 10 pmol/L.(ABSTRACT TRUNCATED AT 250 WORDS)

Adipose Tissue↗
PubMed1995

Tissue factor activity of syncytiotrophoblast plasma membranes and tumoral trophoblast cells in culture.

During pregnancy, important modifications of hemostasis occur resulting in mothers in hypercoagulability and the role of placental cells such as trophoblast cells has been hypothesized. In this study, we first showed that syncytiotrophoblast plasma membranes, isolated from normal human placenta, expressed a strong tissue factor (TF) activity. We then studied TF activity of two continuous trophoblast cell lines (JEG-3 and BeWo) in comparison to human umbilical vein endothelial cells (HUVEC) and transformed human endothelial cells (ECV-304). TF assays were performed on intact detached confluent cells. Unstimulated JEG-3 and BeWo cells exhibited a very high TF activity which slightly increased after 2 to 4 h TNF-alpha stimulation. In contrast, HUVEC and ECV-304 had a lower basal TF activity which was mainly inducible by TNF-alpha, with a maximum effect after 4 to 6 h stimulation. For both cell types, TF activity was decreased to basal value after 16-hour TNF-alpha stimulation. These results support that trophoblast cells are able to express TF but the involvement of this property in the hemostatic physiological changes observed during pregnancy, remains to be demonstrated.

Cell Line, Transformed↗
PubMed1995

Modulation of Ca2+ transients in neonatal and adult rat cardiomyocytes by endothelin-1.

Endothelin-1 (ET-1) may exert inotropic and chronotropic effects in cardiac muscle by modulating intracellular Ca2+. This study assesses effects of ET-1 on intracellular free Ca2+ concentration ([Ca2+]i) in neonatal and adult rat cardiomyocytes. [Ca2+]i was measured by Fura 2 methodology. ET receptor subtypes were determined by binding studies. ET-1 increased neonatal and adult [Ca2+]i and spike frequency in a dose-dependent manner. It decreased [Ca2+]i amplitude in neonatal cardiomyocytes but had no effect on [Ca2+]i amplitude in adult cells. The ET(A) receptor antagonist BQ-123 reduced ET-1-induced responses and the ETB receptor agonist IRL-1620 increased [Ca2+]i. ET-1 binding studies demonstrated 85% displacement by BQ-123 and 15% by the ETB receptor agonist sarafotoxin S6c, suggesting a predominance of ET(A) receptors. These data demonstrate that in neonatal and adult cardiomyocytes ET-1 has stimulatory effects on [Ca2+]i which are mediated predominantly via ET(A) receptors. Therefore, ET-1 may influence cardiac development and function.

Age Factors↗
PubMed1995

Reduced affinity of iodinated forms of Tyr0 C-type natriuretic peptide for rat natriuretic peptide receptor B.

Tyr(O)CNP is an analogue of C-type natriuretic peptide (CNP) with a tyrosine residue added to the NH2 terminus to allow its iodination. In the present study, the suitability of iodinated Tyr(O)CNP as a ligand was tested, and its potency was compared with that of other natural rat natriuretic peptides or structural analogues by radioligand binding experiments. Binding studies were performed on membranes of COS-1 cells transfected with expression plasmids for either rat natriuretic peptide receptor (NPR)-A, rat NPR-B, or bovine NPR-C. 125I-ANP(99-126) was used as a ligand to assess the binding characteristics of NPR-A and -C, and 125I-Tyr(O)CNP was used to study NPR-B. Binding associated to membranes of nontransfected COS cells was always < 3% of the total binding observed in membranes from cells transfected with receptor expression plasmids. Receptor densities in transfected cells ranged from 500 to 2500 fmol/mg of protein. High performance liquid chromatography and ionspray mass spectrometry analyses revealed that the reagents used in the course of iodination (lactoperoxidase, chloramine T, or N-chloromorpholine altered the structure of Tyr(O)CNP, most likely by changing the thiol of the Met17 residue into a sulfoxide. To further evaluate the usefulness of forms of iodinated Tyr(O)CNP on the cGMP responses in cells transfected with NPR-B. In conclusion, the suitability iodinated forms of Tyr(O)CNP as radioligands, we performed iodination of the peptide with cold iodine (Na-127I-). After purification by high performance liquid chromatography, three different modified peptides (i.e. Tyr(O)Met(O)17CNP, 127I-Tyr(O)Met(O)17CNP, and 127I2-Tyr(O)Met(O)17CNP) were recovered, and they were compared with CNP-22, Tyr(O)CNP, ANP(99-126), BNP-32, and des[Gin18, Ser19, Gly20, Leu21, Gly22]ANP(4-23) NH2 (c-ANP) for their ability to bind to transfected receptors. The binding affinity of Tyr(O)CNP for NPR-A and -B receptors is similar to that of CNP. However, oxidation of the Met17 residue into methionine sulfoxide reduces the affinity of the compound for NPR-B by > 10-fold, whereas the addition of one or two iodines did not further reduce its affinity. Similar results were obtained on evaluation of the ability of the oxidized form of monoiodinated Tyr(O)CNP on the cGMP responses in cells transfected with NPR-B. In conclusion, the suitability of iodinated forms of Tyr(O)CNP as radioligands for binding studies on rat NPR-B is not optimal, and the results of studies using such compounds for the detection, identification, and quantification of this receptor should be interpreted with caution.

Animals↗
PubMed1994

Adhesive properties of choriocarcinoma cells toward lymphocytes activated or not by interleukin-2.

Choriocarcinoma cells (CC) in vitro are resistant to NK lysis but sensitive to lysis by blood or decidual effectors activated by interleukin-2 (IL-2). Because lytic activity requires a step of adhesion, the adhesive properties of the choriocarcinoma cells BeWo, JEG-3, and JAR were examined functionally toward peripheral blood lymphocytes. The adhesion of lymphocytes to choriocarcinoma cells was very low and did not increase after stimulating lymphocytes with IL-2. As demonstrated by cytofluorimetry analysis, choriocarcinoma cells and cytotrophoblast cells prepared from term placenta expressed intercellular adhesion molecule-1 (ICAM-1), whereas only CC expressed CD56. Tumor necrosis factor-alpha or interferon-gamma increased the expression of ICAM-1 on choriocarcinoma cells without modifying the adhesion of lymphocytes to choriocarcinoma cells. These results suggest that resistance of choriocarcinoma cells to lysis by cytotoxic effectors could partially be attributed to the low level of lymphocyte adhesion to these cells.

Cell Adhesion↗
PubMed1994

Effect of intravenous atrial natriuretic peptide on gas exchange in humans.

The aim of this work was to establish whether a physiological increase in atrial natriuretic peptide (ANP) plasma levels affects pulmonary gas exchange in humans. Ten volunteers received an infusion of either ANP (4 pmol.kg-1.min-1) or physiological saline, for 60 min. Baseline measures of the alveolar-arterial PO2 difference and of the physiological dead space were within normal limits and remained stable during and after the infusion of ANP or saline, although plasma ANP and cGMP rose significantly (p < 0.01) (mean +/- SEM: ANP: 13.4 +/- 3.9 to 56.0 +/- 10.4 pmol/l; cyclic GMP: 3.8 +/- 0.3 to 17.0 +/- 3.8 nmol/l). We conclude that a physiological increase in plasma ANP does not affect pulmonary gas exchange significantly in humans.

Adult↗
PubMed1994

Identification of a calcium dependent atrial secretory binding protein as calmodulin.

OBJECTIVE: The role of calcium in the regulation of atrial natriuretic factor (ANF) secretion has focused attention on calcium binding proteins as regulatory components of the secretory machinery for ANF. Two members of the annexin family of calcium binding proteins have emerged as candidates for this role. The aim of this paper is to report the isolation, purification, and identification of another calcium dependent atrial secretory granule binding protein. METHODS: A 20 kDa calcium binding protein was isolated from rat cardiac tissue by affinity chromatography and purified by ion exchange high pressure liquid chromatography followed by reverse phase high pressure liquid chromatography. Identification of the protein was based on its electrophoretic mobility, chromatographic properties, amino acid composition, and ability to stimulate the activity of calmodulin dependent phosphodiesterase. RESULTS: Analysis of this protein and its interaction with atrial secretory granules revealed that it was identical to calmodulin and that the calcium dependent interaction with atrial secretory granules takes place at physiological calcium concentrations. CONCLUSIONS: The study identifies calmodulin as a calcium dependent atrial secretory binding protein. The interaction of calmodulin with atrial secretory granules at physiological calcium concentrations suggest a possible role for calmodulin in the regulation of ANF release from the secretory granules.

Animals↗
PubMed1994

Calcium is involved in both positive and negative modulation of the secretory system for ANP.

The calcium dependence of the atrial natriuretic peptide (ANP) secretory system is controversial. Some studies clearly support a stimulatory role, whereas others favor an inhibitory role for calcium in this endocrine system. We demonstrate that calcium is involved in both a positive modulatory role and a negative modulatory role, thereby providing some explanation for the seemingly irreconcilable findings previously published. The negative modulatory role performed by calcium is evident during basal secretion, whereas the positive modulatory role is especially evident in the sustained phase of the secretory response to stimulation. Furthermore, we demonstrate the calcium dependence of processing of the prohormone to the mature circulating form in a cell culture system. This supports the concept that processing is a function of the atrial myocyte rather than of the mesenchymal cells of the atrium. We have demonstrated previously that calcium is important for packaging of the prohormone into secretory granules. Together these findings support a multifaceted role for calcium in the regulation of the secretory apparatus for ANP.

Animals↗
PubMed1994

Cardiac and plasma atrial natriuretic peptide after 9-day hindlimb suspension in rats.

To determine atrial natriuretic peptide (ANP) adaptation to simulated weightlessness, immunoreactive plasma (ir-NH2- and ir-COOH-terminals) and atrial (ir-COOH-terminal) ANP levels, atrial mRNA expression, immunoreactive cardiocyte ANP levels (ir-NH2- and ir-COOH-terminals), and ultrastructural observations of granules in atrial cardiocytes were assessed in male Wistar rats after a 9-day hindlimb suspension. Plasma ir-NH2- and ir-COOH-terminal ANP concentrations decreased by 17 (P < 0.05) and 37% (P < 0.05), respectively, in suspended rats. A concomitant ir-COOH-terminal ANP content reduction was also observed in left (31%; P < 0.01) and right atria (25%; P < 0.05). Atrial ANP mRNA expression was severely depleted in the right atrium and less so in the left atrium after 9 days of hindlimb suspension. Immunocytochemistry observations demonstrated lowered NH2- and COOH-terminal ANP immunoreactivities in left and right atria from suspended rats. A reduced number of storage granules (dense granules) in both atria was also noted on ultrastructural analysis. It was concluded that ANP biosynthesis, storage, and release were decreased after a 9-day hindlimb suspension.

Animals↗
PubMed1994

Endothelin-stimulated secretion of natriuretic peptides by rat atrial myocytes is mediated by endothelin A receptors.

Endothelin (ET), a potent vasoconstrictor peptide, is known to enhance the secretion of atrial natriuretic factor (ANF) by the heart. In the present study, we investigated the potency of ET isopeptides to stimulate ANF and brain natriuretic peptide (BNP) secretion in primary cultures of neonatal atrial myocytes, and we characterized the receptor mediating these effects. All ET isopeptides caused a twofold increase of ANF and BNP secretion with the following order of potency: ET-1 approximately ET-2 > sarafotoxin 6b >> ET-3. Secretion of the natriuretic peptides was blocked by BQ-123, an ETA-receptor antagonist, but was not affected by either IRL-1620 or [Ala1,3,11,15]ET-1, two ETB-receptor agonists. ET receptors were localized by autoradiography on the surface of atrial myocytes, indicating that contaminating cells were not responsible for 125I-ET-1 binding. Competition binding analyses were then used to assess the ET-receptor subtype on atrial myocyte membrane preparations. A high-affinity (100 pmol/L) binding site with high density (approximately 1500 fmol/mg) was found to preferentially bind the ET isopeptides in the following order: ET-1 > or = ET-2 > or = sarafotoxin 6b > ET-3. Binding was totally displaced by BQ-123 but not by IRL-1620. The ET binding site therefore had the characteristics of an ETA-like receptor. Analysis by cross-linking and sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that it possessed a molecular mass of approximately 50 kD. Northern blot analysis of both ETA- and ETB-receptor mRNAs allowed only the detection of the former, indicating that the ETB receptor may be expressed in very small amounts. These results demonstrate that ANF and BNP secretion by atrial myocytes is enhanced by ET via binding to an ETA-like receptor.

Animals↗
PubMed1993

IL-4, but not tumor necrosis factor-alpha, increases endothelial cell adhesiveness for lymphocytes by activating a cAMP-dependent pathway.

IL-4 and TNF-alpha increase endothelial cell adhesiveness for PBL by promoting the expression of adhesion molecules. We investigated the intracellular cAMP involvement in the increased endothelial cell adhesivity induced by IL-4 or TNF-alpha. We showed that both IL-4 and TNF-alpha increased intracellular cAMP in endothelial cells (EC). Furthermore, dibutyryl-cAMP and forskolin (which increased intracellular cAMP) increased basic EC adhesivity for PBL. The co-stimulation of EC with cAMP elevating agents and TNF-alpha, but not IL-4, resulted in an additive increase in EC adhesiveness. 2',5' dideoxyadenosine, an inhibitor of adenylate cyclase, decreased PBL adhesion to IL-4- but not TNF-alpha-treated EC. Similarly, HA1004, a protein kinase A inhibitor, totally reversed the IL-4 but not TNF-alpha effect on EC adhesiveness, whereas H7, a protein kinase C inhibitor, did not antagonise cytokine-enhanced EC adhesivity. These results indicate that IL-4, but not TNF-alpha, uses a cAMP-dependent pathway to increase PBL adhesion. Furthermore, we showed that cAMP elevation in EC did not induce vascular cell adhesion molecule 1, the only identified adhesion molecule induced by IL-4, indicating that a rise in cAMP in EC promotes an as yet unidentified adhesion pathway. Our results show that IL-4 increases EC adhesiveness for PBL through activation of protein kinase A by promoting an unidentified adhesion pathway.

Cell Adhesion↗
PubMed1993

Renal effects of a nonhypotensive i.v. dose of felodipine.

To evaluate the natriuretic effect of a nonhypotensive dose of felodipine, 11 healthy volunteers (age: 21-28 years) on a high-sodium diet received the drug or its vehicle in a double-blind, randomized, crossover study. Administered intravenously at a dose level of 7.5 micrograms/min for 30 minutes followed by 5 micrograms/min for 120 minutes, felodipine increased natriuresis (546 +/- 69 vs. 454 +/- 39 mumol/min, P < 0.001) and diuresis (8.9 +/- 0.6 vs. 7.5 +/- 0.5 mL/min), compared to its vehicle. Renal plasma flow tended to be augmented, but there was a significant reduction of renal vascular resistance (0.085 +/- 0.004 vs. 0.101 +/- 0.012 mm Hg/mL/min, P < 0.03). The glomerular filtration rate was slightly decreased and proximal sodium reabsorption was diminished with no measurable effect on distal function. Felodipine stimulated plasma renin activity, but produced no changes in plasma atrial natriuretic factor, cGMP, aldosterone, and atrial vasopressin levels. In conclusion, felodipine induced natriuresis and diuresis while reducing proximal tubular sodium reabsorption.

Adult↗
PubMed1993

Identification and immunolocalisation of annexins V and VI, the major cardiac annexins, in rat heart.

OBJECTIVE: The heart contains proteins of the annexin family, a unique group of calcium binding proteins. This study was aimed at identifying the major cardiac annexins and determining their distribution in the rat heart. METHODS: Annexins were isolated by affinity chromatography and purified by ion exchange high pressure liquid chromatography. Identification of isolated proteins by immunoblotting was confirmed by partial amino acid sequence determination. Antisera raised against the isolated proteins were used for immunohistochemistry by the avidin-biotin-peroxidase technique. RESULTS: Two annexins were isolated and purified. Amino acid sequencing confirmed their identities as annexin V and VI. Immunohistochemistry showed that both annexins were present in cardiac myocytes and non-myocytes, but a distinct pattern of distribution was seen for each annexin. Annexin V immunoreactivity was enhanced in the atria compared with the ventricles, whereas annexin VI was more uniformly distributed. In individual cardiac myocytes annexin V was distributed throughout the cell by contrast with annexin VI, which localised to the sarcolemma. Intercalated discs displayed immunoreactivity for both annexins, most prominently for annexin VI. The most striking immunoreactivity for annexin V occurred in vascular endothelial cells, both in the microcirculation and in the major coronary vessels. Immunoreactivity for annexin VI in vascular structures was localised to the nuclei of endothelial and smooth muscle cells. CONCLUSIONS: Annexins V and VI are the major cardiac annexins. The localisation of these annexins to different components of cardiac myocytes will serve to direct the search for their functions in the heart. The striking immunoreactivity for annexins, particularly annexin V, in the coronary vessels indicates that the functions of cardiac annexins include a role in the coronary circulatory system.

Amino Acid Sequence↗
PubMed1993

Characterization of receptors for the atrial natriuretic factor in rat renal microvessels.

OBJECTIVE: To characterize [125I]-atrial natriuretic factor [ANF-(99-126)] binding sites in the renal preglomerular microvasculature of Sprague-Dawley rats. METHODS: Renal preglomerular microvessels were isolated by infusion of a magnetized iron oxide solution into the renal arteries and detachment from non-vascular tissue by a magnetic field. In order to characterize [125I]-ANF-(99-126) binding sites, saturation and competitive binding experiments were performed. To evaluate the proportions of ANF receptor subtypes (ANF-R1, ANF-R2), competition curves were charted in the presence of 10(-6) mol/l C-ANF-(4-23), a specific ligand of ANF-R2 (ANP-C). RESULTS: [125I]-ANF binding to vascular membranes was saturable and of high affinity. Equilibrium saturation binding curves suggested the presence of one group of high-affinity receptors [equilibrium dissociation constant (Kd) 22 +/- 6 pmol/l; binding capacity (Bmax) 118 +/- 6 fmol/mg protein]. In competitive inhibition experiments, no significant differences were found in binding capacity between experiments performed either in the presence or in the absence of an excess (1 mumol/l) of C-ANF (94 +/- 27 versus 151 +/- 35 fmol/mg protein, respectively), suggesting that most receptors in the renal vasculature are of the subtype ANF-R1. Incubation of renal microvessels with ANF-(99-126) stimulated cyclic GMP production in a dose-related manner. In parallel studies, the proportion of ANF-R1 (ANP-A, -B) and ANF-R2 (ANP-C) receptors in glomeruli, calculated from competitive inhibition experiments, was 86 +/- 2 and 14 +/- 2%, respectively (P < 0.005). CONCLUSIONS: These results indicate that rat renal preglomerular microvessels contain a high proportion of guanylate cyclase-coupled ANF-R1 (ANP-A, -B) and a low density of ANF-R2 (ANP-C) receptors. This difference in the proportion of ANF receptor subtypes, compared to that reported in glomeruli and other vascular beds, may have physiological significance.

Animals↗