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D Escande

Publications and source records attributed to D Escande.

At least 37 records · Page 2Linked to original sources
PubMed2000

Patch-clamp study of liver nuclear ionic channels reconstituted into giant proteoliposomes.

Nuclear ionic channels (NICs) represent ubiquitous structures of living cells, although little is known about their functional properties and encoding genes. To characterize NICs, liver nuclear membrane vesicles were reconstituted into either planar lipid bilayers or proteoliposomes. Reconstitution of nuclear envelope (NE) vesicles into planar lipid bilayer proceeded with low efficiency. NE vesicle reconstitution into proteoliposomes led to NIC observations by the patch-clamp technique. Large conductance, voltage-gated, K(+)-permeant and Cl(-)-permeant NICs were characterized. An 80-105-pS K(+)-permeant NIC with conducting sub-state was also recorded. Our data establish that NICs can be characterized upon reconstitution into giant proteoliposomes and retain biophysical properties consistent with those described for native NICs.

Animals↗
PubMed2000

Steady-state versus non-steady-state QT-RR relationships in 24-hour Holter recordings.

The aim of the present study was to investigate the QT-RR interval relationship in ambulatory ECG recordings with special emphasis on the physiological circumstances under which the QT-RR intervals follow a linear relation. Continuous ECG recordings make it possible to automatically measure QT duration in individual subjects under various physiological circumstances. However, identification of QT prolongation in Holter recordings is hampered by the rate dependence of QT duration. Comparison of QT duration and QT interval rate dependence between different individuals implies that the nature of the QT-RR relationship is defined in ambulatory ECG. Holter recordings were performed in healthy volunteers at baseline and after administration of dofetilide, a Class III antiarrhythmic drug. After dofetilide, beat-to-beat automated QT measurements on Holter tapes were compared with manually measured QT intervals on standard ECGs matched by time. The QT-RR relationship was analyzed at baseline in individual and group data during three different periods: 24-hour, daytime, and nighttime. Data were collected under steady-state or non-steady-state conditions of cycle length and fitted with various correction formulae. Our study demonstrated an excellent agreement between manually and automated measurements. The classic Bazett correction formula did not fit the QT-RR data points in individual or group data. When heart beats were selected for a steady rhythm during the preceding minute, QT-RR intervals fit a linear relationship during the day and night periods, but not during the 24-hour period in both individual and group data. In contrast, in the absence of beat selection, data fit a more complex curvilinear relationship irrespective of the period. Our study provides the basis for comparison of QT interval durations and QT-RR relationships between individuals and between groups of subjects.

Adolescent↗
PubMed2000

Differential expression of KvLQT1 isoforms across the human ventricular wall.

Long Q-T mutant (KvLQT1) K(+) channels associate with their regulatory subunit IsK to produce the slow component of the delayed rectifier potassium (I(Ks)) cardiac current. The amplitude of KvLQT1 current depends on the expression of a KvLQT1 splice variant (isoform 2) that exerts strong dominant negative effects on the full-length KvLQT1 protein (isoform 1). We used RNase protection assays to determine the relative expression of KvLQT1 isoforms 1 and 2 and IsK mRNAs in human ventricular layers. Overall expression of KvLQT1 and IsK genes was similar in the three layers. However, there was a significant difference in the ratio between KvLQT1 isoforms 1 and 2. Isoform 2 represented 25.2 +/- 2.3%, 31.7 +/- 1.2%, and 24.9 +/- 1.7% of total KvLQT1 expression in left ventricular endocardial, midmyocardial, and epicardial tissues, respectively. Similar data were obtained from right ventricular samples. COS-7 cells were intranuclearly injected with KvLQT1 isoforms 1 or 2 plus IsK cDNAs, using two different isoform 2-to-isoform 1 ratios. Cells injected with an isoform 2-to-isoform 1 ratio mimicking that in the midmyocardium showed a K(+) current with approximately 75% reduced amplitude compared with those injected with a ratio mimicking that in the epicardium. Our results suggest that differential expression of KvLQT1 isoform 2 in endocardial, midmyocardial, and epicardial tissues is responsible for differential I(Ks) amplitude and contributes to the regional action potential heterogeneity observed across the ventricular wall.

Action Potentials↗
PubMed1999

beta3-adrenoceptor control the cystic fibrosis transmembrane conductance regulator through a cAMP/protein kinase A-independent pathway.

In human cardiac myocytes, we have previously identified a functional beta3-adrenoceptor in which stimulation reduces action potential duration. Surprisingly, in cardiac biopsies obtained from cystic fibrosis patients, beta3-adrenoceptor agonists produced no effects on action potential duration. This result suggests the involvement of cystic fibrosis transmembrane conductance regulator (CFTR) chloride current in the electrophysiological effects of beta3-adrenoceptor stimulation in non-cystic fibrosis tissues. We therefore investigated the control of CFTR activity by human beta3-adrenoceptors in a recombinant system: A549 human cells were intranuclearly injected with plasmids encoding CFTR and beta3-adrenoceptors. CFTR activity was functionally assayed using the 6-methoxy-N-(3-sulfopropyl)quinolinium fluorescent probe and the patch-clamp technique. Injection of CFTR-cDNA alone led to the expression of a functional CFTR protein activated by cAMP or cGMP. Co-expression of CFTR (but not of mutated DeltaF508-CFTR) with high levels of beta3-adrenoceptor produced an increased halide permeability under base-line conditions that was not further sensitive to cAMP or beta3-adrenoceptor stimulation. Patch-clamp experiments confirmed that CFTR channels were permanently activated in cells co-expressing CFTR and a high level of beta3-adrenoceptor. Permanent CFTR activation was not associated with elevated intracellular cAMP or cGMP levels. When the expression level of beta3-adrenoceptor was lowered, CFTR was not activated under base-line conditions but became sensitive to beta3-adrenoceptor stimulation (isoproterenol plus nadolol, SR 58611, or CGP 12177). This later effect was not prevented by protein kinase A inhibitors. Our results provide molecular evidence that CFTR but not mutated DeltaF508-CFTR is regulated by beta3-adrenoceptors expression through a protein kinase A-independent pathway.

Animals↗
PubMed1999

In memoriam

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Journal Article↗
PubMed1999

Mutations in a dominant-negative isoform correlate with phenotype in inherited cardiac arrhythmias.

The long QT syndrome is characterized by prolonged cardiac repolarization and a high risk of sudden death. Mutations in the KCNQ1 gene, which encodes the cardiac KvLQT1 potassium ion (K+) channel, cause both the autosomal dominant Romano-Ward (RW) syndrome and the recessive Jervell and Lange-Nielsen (JLN) syndrome. JLN presents with cardiac arrhythmias and congenital deafness, and heterozygous carriers of JLN mutations exhibit a very mild cardiac phenotype. Despite the phenotypic differences between heterozygotes with RW and those with JLN mutations, both classes of variant protein fail to produce K+ currents in cultured cells. We have shown that an N-terminus-truncated KvLQT1 isoform endogenously expressed in the human heart exerts strong dominant-negative effects on the full-length KvLQT1 protein. Because RW and JLN mutations concern both truncated and full-length KvLQT1 isoforms, we investigated whether RW or JLN mutations would have different impacts on the dominant-negative properties of the truncated KvLQT1 splice variant. In a mammalian expression system, we found that JLN, but not RW, mutations suppress the dominant-negative effects of the truncated KvLQT1. Thus, in JLN heterozygous carriers, the full-length KvLQT1 protein encoded by the unaffected allele should not be subject to the negative influence of the mutated truncated isoform, leaving some cardiac K+ current available for repolarization. This is the first report of a genetic disease in which the impact of a mutation on a dominant-negative isoform correlates with the phenotype.

Adult↗
PubMed1999

Respiratory epithelial ion transport in patients with disseminated bronchiectasis.

The nosological limits between disseminated bronchiectasis and cystic fibrosis (CF) remain unclear. In patients with isolated congenital bilateral absence of the vas deferens, a forme fruste of the CF disease, a normal baseline nasal transepithelial potential difference (PD) but an impaired response to pharmacological interventions have been reported. The purpose of the present study was to explore ion transport in respiratory epithelium from patients with disseminated bronchiectasis. The PD under both baseline and pharmacological interventions was investigated in 13 healthy subjects, six patients with genetically proven CF and 15 patients with disseminated bronchiectasis as confirmed by computed tomography scan. Baseline PD was similar in the control and bronchiectasis groups but, as expected, was significantly more negative in the CF group. Patients with bronchiectasis responded to pharmacological tests (sequential perfusion with amiloride, chloride-free solution, isoprenaline and uridine triphosphate (UTP) similarly to healthy subjects. In contrast, CF patients exhibited an increased response to amiloride and an impaired response to chloride-free solution and isoprenaline. The data show that patients with disseminated bronchiectasis exhibit normal electrophysiological properties in their nasal epithelium. Nasal transepithelial potential difference including pharmacological tests may appear a valuable diagnostic procedure for cystic fibrosis with disseminated bronchiectasis.

Adolescent↗
PubMed1999

Interspecies differences in the cardiac negative inotropic effects of beta(3)-adrenoceptor agonists.

The aim of the present study was to compare the effects of three preferential (BRL 37344, SR 58611, CL 316 243) and a partial (CGP 12177) beta-adrenoceptor (beta(3)-AR) agonists on the contractility of ventricular strips sampled from various mammalian species including humans. In the human heart, all beta(3)-AR agonists tested decreased contractility by 40 to 60% below control with an order of potency: BRL 37344 > CL 316 243 = SR 58611 >> CGP 12177. In the dog, the negative inotropic effects produced by beta(3)-AR stimulation were less pronounced than in humans, approximately 30% below control. The order of potency of beta(3)-AR agonists was CGP 12177 > BRL 37344 = SR 58611 >> CL 316 243; i.e., very different from that observed in humans. In rat, only BRL 37344 was efficient to decrease contractility. In guinea pig, only CL 316 243 significantly reduced peak tension. In both species, the reduction in peak tension did not exceed 20 to 30%. Finally, in the ferret, none of the agonists tested induced a negative inotropic effect. In dog, the negative inotropic effects of CGP 12177 were not modified by nadolol, but were abolished by bupranolol, a beta(1-3)-AR. beta(3)-AR transcripts were detected in the dog but not in the rat ventricle by using a reverse transcription-polymerase chain reaction assay. We conclude that cardiac negative inotropic effects related to beta(3)-AR agonist stimulation vary markedly depending on the species. A comparable interspecies variation previously has been reported concerning the lipolytic effects of beta(3)-AR agonist stimulation. Our study demonstrates that the pharmacological profile of a beta(3)-AR agonist on the human myocardium cannot be extrapolated from usual animal models.

Adrenergic beta-Agonists↗
PubMed1998

Adult KCNE1-knockout mice exhibit a mild cardiac cellular phenotype.

The KCNE1 gene encodes a channel regulator IsK which in association with the KvLQT1 K+ channel protein determines the slow component of the cardiac delayed rectifier current. We have investigated the cellular electrophysiological characteristics of adult KCNE1-knockout mouse hearts by means of the standard microelectrode technique. Action potential parameters from the ventricular endocardium of KCNE1 -/- mice were indistinguishable from those of KCNE1 +/+ animals. In particular, KCNE1 -/- hearts did not exhibit prolonged repolarization. E-4031, a specific blocker of erg K+ channels consistently prolonged repolarization in KCNE1 +/+ but not in KCNE1 -/- hearts. By contrast, the chromanol compound 293B, a specific blocker of KvLQT1 K+ channel produced comparable effects on repolarization in KCNE1 -/- and KCNE1 +/+ mice. We conclude that invalidation of the mouse KCNE1 gene by homologous recombination leads to a mild cardiac phenotype at the cellular level.

Action Potentials↗
PubMed1998

Polyethylenimine but not cationic lipids promotes transgene delivery to the nucleus in mammalian cells.

The beta-galactosidase reporter gene, either free or complexed with various cationic vectors, was microinjected into mammalian cells. Cationic lipids but not polyethylenimine or polylysine prevent transgene expression when complexes are injected in the nucleus. Polyethylenimine and to a lesser extent polylysine, but not cationic lipids, enhance transgene expression when complexes are injected into the cytoplasm. This latter effect was independent of the polymer vector/cDNA ionic charge ratio, suggesting that nucleic acid compaction rather than surface charge was critical for efficient nuclear trafficking. Cell division was not required for nuclear entry. Finally, comparative transfection and microinjection experiments with various cell lines confirm that barriers to gene transfer vary with cell type. We conclude that polymers but not cationic lipids promote gene delivery from the cytoplasm to the nucleus and that transgene expression in the nucleus is prevented by complexation with cationic lipids but not with cationic polymers.

Animals↗
PubMed1998

A dominant negative isoform of the long QT syndrome 1 gene product.

Mutations in the KvLQT1 gene are the cause of the long QT syndrome 1. KvLQT1 gene product is associated with the regulator protein IsK to produce a component of the delayed rectifier K+ current in cardiac myocytes. We identified an N-terminal truncated isoform of the KvLQT1 gene product, referred to as isoform 2. In RNase protection assays, isoform 2 represented 28.1 +/- 0.6% of the total KvLQT1 expression in the human adult ventricle. COS-7 cells injected intranuclearly with KvLQT1 isoform 1 cDNA exhibited a fast-activating K+ current, whereas those injected with a KvLQT1 isoform 1 plus IsK cDNA showed a slow-activating K+ current. Cells injected with KvLQT1 isoform 2 plasmid showed no detectable K+ current. Those injected with a 1/1 isoform 2/isoform 1 ratio showed no detectable K+ current. Those injected with 1/5 and 2/5 ratios showed a K+ current with markedly reduced amplitude. Coexpression of the IsK regulator consistently reduced the dominant negative effects of isoform 2. Our results indicate that KvLQT1 isoform 2 exerts a pronounced negative dominance on isoform 1 channels and that the cardiac KvLQT1 K+ channel complex is composed of at least three different proteins as follows: isoform 1, isoform 2, and IsK.

Animals↗
PubMed1998

Hyperexpression of recombinant CFTR in heterologous cells alters its physiological properties.

We investigated whether high levels of expression of the cystic fibrosis transmembrane conductance regulator (CFTR) would alter the functional properties of newly synthesized recombinant proteins. COS-7, CFPAC-1, and A549 cells were intranuclearly injected with a Simian virus 40-driven pECE-CFTR plasmid and assayed for halide permeability using the 6-methoxy-N-(3-sulfopropyl)quinolinium fluorescent probe. With increasing numbers of microinjected pECE-CFTR copies, the baseline permeability to halide dose dependently increased, and the response to adenosine 3',5'-cyclic monophosphate (cAMP) stimulation decreased. In cells hyperexpressing CFTR, the high level of halide permeability was reduced when a cell metabolism poisoning cocktail was applied to decrease intracellular ATP and, inversely, was increased by orthovanadate. In CFPAC-1 cells investigated with the patch-clamp technique, CFTR hyperexpression led to a time-independent nonrectifying chloride current that was not sensitive to cAMP stimulation. CFPAC-1 cells hyperexpressing CFTR exhibited no outward rectifying chloride current nor inward rectifying potassium current either spontaneously or under cAMP stimulation. We conclude that hyperexpression of recombinant CFTR proteins modifies their properties inasmuch as 1) CFTR channels are permanently activated and not susceptible to cAMP regulation and 2) they lose their capacity to regulate heterologous ionic channels.

Animals↗
PubMed1997

Vasomotor dysfunction early after exposure of normal rabbit arteries to an adenoviral vector.

We aimed to investigate whether infection of normal rabbit arteries with a recombinant adenovirus vector would result per se in alterations in contractile and endothelial functions. In one group of rabbits, right or left femoral and ear artery segments were injected in vivo with a replication-deficient adenoviral vector expressing a beta-galactosidase (beta-Gal) reporter gene (4 x 10(10) pfu/ml) to demonstrate efficient gene transfer. Contralateral arteries were injected with the same concentration of a recombinant adenoviral vector carrying no transgene (Ad.MLPnull). In another group of animals, Ad.MLPnull was injected into the lumen of femoral and ear artery segments. The contralateral arteries were used as controls with the injection of vehicle alone. Histochemical assessment of gene transfer using beta-Gal activity (group 1) or in vitro contractility and endothelial function (group 2) was performed 3 days after adenoviral infection. Gene transfer was efficient and reproducible in the endothelium and was associated with the presence of inflammatory cells in the media. In Ad.MLPnull-injected arteries, in vitro contractile response of femoral artery rings to either KCl 60 mM or phenylephrine (10 microM) was reduced to 10.5 +/- 2.3% (n = 14; p < 0.001) and 8.8 +/- 2.0% (n = 7; p < 0.001) of the control values, respectively. Furthermore, in arteries injected with Ad.MLPnull, the endothelium-dependent relaxation produced by acetylcholine (10 microM) was virtually abolished. Similarly, the relaxant effects of the alpha 2-adrenoreceptor agonist UK14304 (1 microM) or the Ca2+ ionophore A23187 (1 microM) were also abolished. By contrast, sodium nitroprusside (10 microM) was still able to relax adenovirus-infected arteries. We conclude that infection with a recombinant adenoviral vector can induce early severe vasomotor alterations in both contractile function and endothelium-mediated relaxation of normal rabbit arteries.

Acetylcholine↗
PubMed1997

[Human life and energy production. Prospects opened up by controlled thermonuclear fusion].

The massive and presently increasing energy production is going to confront mankind with a very important problem in the forthcoming decades, in particular due to the vanishing of resources and to the greenhouse effect. The share of fossil fuels in the energy production will have to decrease, and other energy sources will be needed. Among them controlled thermonuclear fusion has may assets due to its non-radioactive fuel with plentiful supply, its non radioactive and non polluting ashes, its safety, its weak environmental impact, and its irrelevance to nuclear proliferation in a normal setting. During the last three decades, physicists have made a series of steps toward the peaceful use of the dominant source of energy in the Universe. They have learned how to confine by magnetic fields plasmas at temperatures of 200 millions degrees centigrade, and they have developed several specific technologies. This way, they produced 11 million watts of nuclear power by fusing two isotopes of hydrogen. These investigations are conducted in a responsible spirit, that of ecoproduction, where possible negative consequences are anticipated, are made as low as reasonably achievable, and their management is studied. Yet several fundamental issues still have to be solved before on economically efficient industrial thermonuclear power plant be operated. A huge international collaboration involving Japan, the USA, the Russian Federation, and the European Union joined with Switzerland and Canada, is presently designing the first experimental thermonuclear reactor, the International Thermonuclear Experimental Reactor (ITER). It would cost 9 billion dollars, a cost similar to other large scientific projects. This is an important step toward an electricity producing thermonuclear reactor that would be both safe and respectful of human health and of environment.

Conservation of Energy Resources↗