Biomedical subjects

C Breton

Publications and source records attributed to C Breton.

At least 37 records · Page 2Linked to original sources

PubMed2000

Estradiol-stimulated nitric oxide release in human granulocytes is dependent on intracellular calcium transients: evidence of a cell surface estrogen receptor.

We tested the hypothesis that estrogen acutely stimulates constitutive nitric oxide synthase activity in human granulocytes by acting on a cell surface estrogen receptor (ER). The release of nitric oxide was measured in real time with an amperometric probe. Exposure of granulocytes to 17beta-estradiol stimulated NO release within seconds in a concentration-dependent manner. The NO release was also stimulated by 17beta-estradiol conjugated to bovine serum albumin (E(2)-BSA), which suggests mediation by a cell surface receptor. Tamoxifen, an ER inhibitor, antagonized the action of both 17beta-estradiol and E(2)-BSA, whereas ICI 182,780, an inhibitor of the nuclear ER, had no effect. Using dual emission microfluorometry in a calcium-free medium, the 17beta-estradiol-stimulated release of NO from granulocytes was shown to be dependent on intracellular calcium ([Ca(2+)]i) transients in a tamoxifen-sensitive process. Exposure to BAPTA-AM (1,2bis-(-aminophenoxy)ethans-N,N,N', N'-tetraacetic acid tetra(acetoxyymethyl) ester), a [Ca(2+)]i chelator, reduced [Ca(2+)]i in response to E(2)-BSA, and depleting [Ca(2+)]i stores abolished the effect of 17beta-estradiol on NO release. Confocal photomicrographs using E(2)-BSA-FITC (fluorescein isothiocyanate) revealed cell membrane reactivity. Estrogen-stimulated NO release had an immunosuppressive effect, and it initiated granulocyte rounding and loss of adherence in a tamoxifen-sensitive manner. Finally, using reverse transcriptase-polymerase chain reaction, human neutrophil granulocytes expressed ERalpha but not ERbeta, suggesting that ERalpha may be the membrane receptor for 17beta-estradiol. The study demonstrated that a physiological dose of estrogen down-regulates granulocyte activity by acutely stimulating NO release via the activation of a cell surface ER which is coupled to increases in [Ca(2+)]i. (Blood. 2000;95:3951-3958)

Animals↗

PubMed2000

Cell-surface estrogen receptors mediate calcium-dependent nitric oxide release in human endothelia.

BACKGROUND: Although estrogen replacement therapy has been associated with reduction of cardiovascular events in postmenopausal women, the mechanism for this benefit remains unclear. Because nitric oxide (NO) is considered an important endothelium-derived relaxing factor and may function to protect blood vessels against atherosclerotic development, we investigated the acute effects of physiological levels of estrogen on NO release from human internal thoracic artery endothelia and human arterial endothelia in culture. METHODS AND RESULTS: We tested the hypothesis that estrogen acutely stimulates constitutive NO synthase activity in human endothelial cells by acting on a cell-surface receptor. NO release was measured in real time with an amperometric probe. 17beta-Estradiol exposure to internal thoracic artery endothelia and human arterial endothelia in culture stimulated NO release within seconds in a concentration-dependent manner. 17beta-Estradiol conjugated to bovine serum albumin also stimulated NO release, suggesting action through a cell-surface receptor. Tamoxifen, an estrogen receptor inhibitor, antagonized this action. We further showed with the use of dual emission microfluorometry that 17beta-estradiol-stimulated release of endothelial NO was dependent on the initial stimulation of intracellular calcium transients. CONCLUSIONS: Physiological doses of estrogen immediately stimulate NO release from human endothelial cells through activation of a cell-surface estrogen receptor that is coupled to increases in intracellular calcium.

Adult↗

PubMed2000

[Laparoscopic excision of a duodenal neuroendocrine tumor].

A 75-year-old woman with melena was found to have a carcinoid tumor in the posterior wall of the duodenal bulb. The biology was normal. The tumor measured 10 mm in size, and endoscopic ultrasonography showed only submucosal involvement. There was no liver metastasis and no regional lymph nodes. Tumoral resection was performed laparoscopically with success. Postoperative course was uneventful. Laparoscopic resection could be an appropriate minimally invasive treatment for selected small size duodenal tumors.

Aged↗

PubMed2000

Comparative biology of the endocannabinoid system possible role in the immune response.

In this review we discuss data showing that the endogenous cannabinoid system, represented by cannabinoid receptors, endogenous cannabinoid receptor ligands and enzymes for the biosynthesis and degradation of these ligands, is conserved throughout evolution from coelenterates to man. This signaling system has been suggested to play several roles in animals, including the regulation of cell development and growth, nervous functions, reproduction and feeding behavior. In this article, however, we shall describe with more detail the possible function of the endogenous cannabinoid system in the modulation of immune response in organisms from the lower to the higher levels of animal evolution.

Animals↗

PubMed2000

Structure and expression of the variant melanin-concentrating hormone genes: only PMCHL1 is transcribed in the developing human brain and encodes a putative protein.

PMCHL1 and PMCHL2 are two copies of the so-called variant melanin-concentrating hormone (MCH) gene that are located, respectively, on human chromosome 5p14 and 5q13 and that emerged recently during primate evolution. They correspond to a 5'-end truncated version of the MCH gene mapped on chromosome 12q23 and encoding a neuropeptide precursor. The gene organization and regulation of the expression of the variant MCH genes in the human brain are the central issues we investigated. First, the structure and fine chromosomal mapping of the 5p and 5q variant MCH genes were established. These revealed several point mutations and length variations of one CA/TA repeat which allow discrimination between each copy. Using a combination of RACE-PCR, RT-PCR, and sequencing analysis, we provided strong evidence for the expression of the PMCHL1 gene but not the PMCHL2 gene in the human fetal, newborn, and adult brains. Sense, potentially coding, RNAs, as well as noncoding antisense RNAs, were identified and displayed a region-specific expression in the human brain. Strikingly, sense unspliced RNAs of the PMCHL1 gene carried a novel open reading frame and may produce an NLS-containing protein of 8 kDa named VMCH-p8. These transcripts were translated in vitro and in transfected COS cells. Therefore, the PMCHL1 gene provides a unique example of the generation of a gene in the Hominoidae lineage which is specifically transcribed in the developing human brain and has the capacity to be translated into a putative novel protein.

Amino Acid Sequence↗

PubMed2000

Thermobacillus xylanilyticus gen. nov., sp. nov., a new aerobic thermophilic xylan-degrading bacterium isolated from farm soil.

An aerobic, thermophilic, xylanolytic, spore-forming bacterium, XETP (T = type strain; P = patent strain), has been isolated from farm soil situated underneath a manure heap in northern France. Strain XETP, which stained negative in the Gram test, occurs as short rods which sometimes form chains. Its spores are ellipsoidal, central to subterminal and occur in swollen sporangia. It grows at temperatures up to 63 degrees C and in the pH range 6.5-8.5. When grown on glucose in optimal conditions, its doubling time was found to be 33 min. CO2 was observed to have a growth-stimulating effect at the start of the culture. In addition to glucose, the isolate utilizes xylose, arabinose, mannose, cellobiose, galactose, maltose, sucrose, xylan and starch. Growth is inhibited by 5% NaCl. The G+C content of strain XETP is 57.5 mol%. The 16S rDNA sequence analysis indicated that strain XETP falls into the radiation of the Bacillus-Lactobacillus-Streptococcus subdivision of the Gram-positive phylum. Its three closest phylogenetic relatives are 'Bacillus viscosus', Paenibacillus curdlanolyticus and Bacillus popilliae with identity values of 91.15, 90.94 and 90.92%, respectively. The major cellular fatty acids are 14-methyl pentadecanoic acid (16:0 iso), hexadecanoic acid (16:0) and 14-methyl hexadecanoic acid (17:0 anteiso). On the basis of 16S rRNA sequence and chemotaxonomic characteristics, the isolate is different enough for it to be considered as a member of a new genus. It is therefore proposed that this isolate represents a new genus and species: Thermobacillus xylanilyticus. Strain XETP, the type strain of Thermobacillus xylanilyticus, has been deposited in the Collection Nationale de Cultures Microbiennes (CNCM I-1017) as a patent strain.

Aerobiosis↗

PubMed1999

Estradiol coupling to human monocyte nitric oxide release is dependent on intracellular calcium transients: evidence for an estrogen surface receptor.

We tested the hypothesis that estrogen acutely stimulates constitutive NO synthase (cNOS) activity in human peripheral monocytes by acting on an estrogen surface receptor. NO release was measured in real time with an amperometric probe. 17beta-estradiol exposure to monocytes stimulated NO release within seconds in a concentration-dependent manner, whereas 17alpha-estradiol had no effect. 17beta-estradiol conjugated to BSA (E2-BSA) also stimulated NO release, suggesting mediation by a membrane surface receptor. Tamoxifen, an estrogen receptor inhibitor, antagonized the action of both 17beta-estradiol and E2-BSA, whereas ICI 182,780, a selective inhibitor of the nuclear estrogen receptor, had no effect. We further showed, using a dual emission microfluorometry in a calcium-free medium, that the 17beta-estradiol-stimulated release of monocyte NO was dependent on the initial stimulation of intracellular calcium transients in a tamoxifen-sensitive process. Leeching out the intracellular calcium stores abolished the effect of 17beta-estradiol on NO release. RT-PCR analysis of RNA obtained from the cells revealed a strong estrogen receptor-alpha amplification signal and a weak beta signal. Taken together, a physiological dose of estrogen acutely stimulates NO release from human monocytes via the activation of an estrogen surface receptor that is coupled to increases in intracellular calcium.

Calcium↗

PubMed1999

A single domain thermophilic xylanase can bind insoluble xylan: evidence for surface aromatic clusters.

A clone expressing xylanase activity in Escherichia coli has been selected from a genomic plasmid library of the thermophilic Bacillus strain D3. Subcloning from the 9-kb insert located the xylanase activity to a 2.7-kb HindII/BamHI fragment. The DNA sequence of this clone revealed an ORF of 367 codons encoding a single domain type-F or family 10 enzyme, which was designated as XynA. Purification of the enzyme following over-expression in E. coli produced an enzyme of 42 kDa with a temperature optimum of 75 degrees C which can efficiently bind and hydrolyse insoluble xylan. The pH optimum of the enzyme is 6.5, but it is active over a broad pH range. A homology model of the xylanase has been constructed which reveals a series of surface aromatic residues which form hydrophobic clusters. This unusual structural feature is strikingly similar to the situation observed in the structure determined for the type-G xylanase from the Bacillus D3 strain and may constitute a common evolutionary mechanism imposed on different structural frameworks by which these xylanases may bind potential substrates and exhibit thermostability.

Amino Acid Sequence↗

PubMed1999

Docking of linear peptide antagonists into the human V(1a) vasopressin receptor. Identification of binding domains by photoaffinity labeling.

A novel photoactivatable linear peptide antagonist selective for the V(1a) vasopressin receptor, [(125)I][Lys(3N(3) Phpa)(8)]HO-LVA, was synthesized, characterized, and used to photolabel the human receptor expressed in Chinese hamster ovary cells. Two specific glycosylated protein species at 85-90 and 46 kDa were covalently labeled, a result identical to that obtained with a previous photosensitive ligand, [(125)I]3N(3)Phpa-LVA (Phalipou, S., Cotte, N. , Carnazzi, E., Seyer, R., Mahe, E., Jard, S., Barberis, C., and Mouillac, B. (1997) J. Biol. Chem. 272, 26536-26544). To identify contact sites between the new photoreactive analogue and the V(1a) receptor, the labeled receptors were digested with Lys-C or Asp-N endoproteinases and chemically cleaved with CNBr. Fragmentation with CNBr, Lyc-C, and Asp-N used alone or in combination, led to the identification of a restricted receptor region spanning the first extracellular loop. The results established that sequence Asp(112)-Pro(120) could be considered as the smallest covalently labeled fragment with [(125)I][Lys(3N(3)Phpa)(8)]HO-LVA. Based on the present experimental result and on previous photoaffinity labeling data obtained with [(125)I]3N(3)Phpa-LVA (covalent attachment to transmembrane domain VII), three-dimensional models of the antagonist-bound receptors were constructed and then verified by site-directed mutagenesis studies. Strikingly, these two linear peptide antagonists, when bound to the V(1a) receptor, could adopt a pseudocyclic conformation similar to that of the cyclic agonists. Despite divergent functional properties, these peptide antagonists could interact with a transmembrane-binding site significantly overlapping that of the natural hormone vasopressin.

Amino Acid Sequence↗

PubMed1999

Up-regulation of vasopressin V(1a) receptor mRNA in rat facial motoneurons following axotomy.

We have reported previously that axotomy induced a marked increase of vasopressin receptor binding in the adult rat facial nucleus, suggesting an increased number of vasopressin receptors. These receptors were pharmacologically undistinguishable from peripheral V(1a) vasopressin receptors. In the present study, we show, using in situ hybridization and reverse transcriptase-polymerase chain reaction (RT-PCR), that axotomy regulates the expression of the vasopressin V(1a) receptor mRNA in the facial nucleus. Results were obtained from adult male rats killed 1 week following crush of the right facial nerve. In situ hybridization was performed with a (35)S-labelled riboprobe. A specific hybridization signal was detected in both left and right facial nuclei, with a significantly higher intensity in the nucleus ipsilateral to the lesion. V(1a) receptor transcripts were found associated with large facial motoneuronal cell bodies, not with other cells present in the nucleus, i.e., glial or epithelial cells. RT-PCR analysis of unlesioned facial tissue revealed the presence of mRNAs encoding vasopressin V(1a), vasopressin V(1b) and oxytocin receptors, whereas only the V(1a) receptor mRNA was found to be increased following axotomy in the lesioned facial tissue. These data suggest that the axotomy-induced expression of vasopressin receptors in the rat facial nucleus is due, at least to a large extent, to an increase of the V(1a) vasopressin receptor mRNA in facial motoneurons.

Animals↗

PubMed1999

[Results of laparoscopic hernioplasty. A study of 401 cases in 318 patients].

STUDY AIM: The aim of this retrospective study was to report a series of laparoscopic hernioplasty performed in two surgical centers, and to evaluate the results with a mean follow-up of 31 months. PATIENTS AND METHODS: From January 1992 to January 1997, 318 patients with 401 inguinal hernias were operated on through laparoscopy by six senior surgeons and six junior surgeons. There were 302 men and 16 women (mean age: 53 years). The operation was performed through an extra-peritoneal approach (TEP) in 298 hernias, a trans-abdomino-preperitoneal approach (TAPP) in 62 hernias, and an intra-abdominal approach (IPOM) in 41 hernias. RESULTS: Conversion into open surgery was necessary in 7% of the patients. There was no postoperative death. The postoperative morbidity rate was 10%. The average hospital stay was three days. With a 1 to 5 year follow-up, 4% of the 94% of the patients who answered the questionnaire showed a recurrence (3% in the extra-peritoneal group; 4% in the trans abdomino-preperitoneal group; 10% in the intra-abdominal group). CONCLUSION: Laparoscopic hernioplasty seems as efficient as traditional hernoplasty with the advantages of mini-invasive surgery. The extra-peritoneal approach was preferred and performed in most cases of this series. The intra-peritoneal approach was abandoned.

Abdomen↗

PubMed1999

[Laparoscopic splenorraphy using a resorbable prosthesis in splenic injuries. Apropos of 5 cases].

STUDY AIM: The aim of this retrospective study is to report five cases of laparoscopic splenorraphy with an absorbable perisplenic mesh for splenic injury. PATIENTS AND METHOD: From January 1996 to February 1998, three men and two women (mean age: 52 years) were included in this study. The splenic lesions were due to either a fall (n = 3), a traffic accident (n = 1), or pleural paracenthesis in a patient with mediastinitis after valvular replacement. Splenic injury was recognized by ultrasonography. The patients were operated as either emergency cases (n = 2), or within 24 hours (n = 3). The procedure included evacuation of the hemoperitineum, total liberation of the spleen, and splenic hemostasis with a perisplenic mesh which was used in open surgery. The mesh placed behind the spleen, covering its superior and inferior poles, was unrolled forwards and burses progressively tightened. RESULTS: There was no conversion, no mortality, no morbidity. In the four injured patients, the mean duration of surgery was 120 minutes (70-180), without any blood transfusion, and the patients were discharged on d4 or 5. The fifth patient, after valvular replacement, was operated on with anticoagulation. The mean duration of surgery was 270 minutes. Four blood units were necessary. He was discharged at d26. CONCLUSION: This technique combines the advantages of the perisplenic mesh which is efficient and safe, with the advantages of laparoscopic surgery which simplifies the postoperative course. It can only be used in case of isolated splenic injury in patients with stable hemodynamic condition.

Absorbable Implants↗

PubMed1999

How can laparoscopic management assist radiation treatment in cervix carcinoma?

PURPOSE: To determine the role of laparoscopic lymphadenectomy (pelvis +/- para-aortic nodes) and laparoscopic hysterectomy in cervical cancer compared to 'classic radical surgery' in patients undergoing surgery in comparison with modern imaging in patients treated with radiotherapy alone. MATERIALS AND METHODS: The limitations of modern imaging are presented as well as how complication rates can be increased when classic laparotomy is followed by radiation therapy. Laparoscopic procedures are described with particular emphasis on how to provide information on lymph node metastases with the risk of overlooking microscopic involvement. A number of clinical experiences are cited to illustrate this problem and show how treatment approaches can be adapted. RESULTS: The role of laparoscopy is evaluated according to different clinical situations and treatment protocols emphasizing the possibilities offered by this method to the radiotherapist. CONCLUSION: When developing laparoscopic techniques for the management of cervical carcinoma, caution must be exercised to ensure that these techniques are not detrimental to the prognosis.

Female↗

PubMed1999

Structure/function studies of glycosyltransferases.

Glycosyltransferases are the enzymes that synthesize oligosaccharides, polysaccharides and glycoconjugates. The analysis of the wealth of sequences that are now available in databases allowed the determination of conserved peptide motifs for each class of enzyme. Recent experimental data demonstrated their importance in donor and acceptor substrate binding and in catalysis. Fold-recognition studies provided the first models of the catalytic domains of some of these enzymes, while the first successes in glycosyltransferase crystallography are opening new routes in structural glycobiology.

Amino Acid Sequence↗

PubMed1999

Divergent evolution of fucosyltransferase genes from vertebrates, invertebrates, and bacteria.

On the basis of function and sequence similarities, the vertebrate fucosyltransferases can be classified into three groups: alpha-2-, alpha-3-, and alpha-6-fucosyltransferases. Thirty new putative fucosyltransferase genes from invertebrates and bacteria and six conserved peptide motifs have been identified in DNA and protein databanks. Two of these motifs are specific of alpha-3-fucosyltransferases, one is specific of alpha-2-fucosyltransferases, another is specific of alpha-6-fucosyltransferases, and two are shared by both alpha-2- and alpha-6-fucosyltranserases. Based on these data, literature data, and the phylogenetic analysis of the conserved peptide motifs, a model for the evolution offucosyltransferase genes by successive duplications, followed by divergent evolution is proposed, with either two different ancestors, one for the alpha-2/6-fucosyltransferases and one for the alpha-3-fucosyltransferases or a single common ancestor for the two families. The expected properties of such an hypothetical ancestor suggest that the plant or insect alpha-3-fucosyltransferases using chitobiose as acceptor might be the present forms of this ancestor, since fucosyltransferases using chitobiose as acceptor are expected to be of earlier appearance in evolution than enzymes using N -acetyllactosamine. However, an example of convergent evolution of fucosyltransferase genes is suggested for the appearance of the Leaepitopes found in plants and primates.

Amino Acid Sequence↗

PubMed1999

Fold recognition study of alpha3-galactosyltransferase and molecular modeling of the nucleotide sugar-binding domain.

The structure and fold of the enzyme responsible for the biosynthesis of the xenotransplantation antigen, namely pig alpha3 galactosyltransferase, has been studied by means of computational methods. Secondary structure predictions indicated that alpha3-galactosyltransferase and related protein family members, including blood group A and B transferases and Forssman synthase, are likely to consist of alternating alpha-helices and beta-strands. Fold recognition studies predicted that alpha3-galactosyltransferase shares the same fold as the T4 phage DNA-modifying enzyme beta-glucosyltransferase. This latter enzyme displays a strong structural resemblance with the core of glycogen phosphorylase b. By using the three-dimensional structure of beta-glucosyltransferase and of several glycogen phosphorylases, the nucleotide binding domain of pig alpha3-galactosyltransferase was built by knowledge-based methods. Both the UDP-galactose ligand and a divalent cation were included in the model during the refinement procedure. The final three-dimensional model is in agreement with our present knowledge of the biochemistry and mechanism of alpha3-galactosyltransferases.

Amino Acid Sequence↗

PubMed1999

Identification of a plasmid-borne locus in Rhizobium etli KIM5s involved in lipopolysaccharide O-chain biosynthesis and nodulation of Phaseolus vulgaris.

Screening of derivatives of Rhizobium etli KIM5s randomly mutagenized with mTn5SSgusA30 resulted in the identification of strain KIM-G1. Its rough colony appearance, flocculation in liquid culture, and Ndv(-) Fix(-) phenotype were indicative of a lipopolysaccharide (LPS) defect. Electrophoretic analysis of cell-associated polysaccharides showed that KIM-G1 produces only rough LPS. Composition analysis of purified LPS oligosaccharides from KIM-G1 indicated that it produces an intact LPS core trisaccharide (alpha-D-GalA-1-->4[alpha-D-GalA-1-->5]-Kdo) and tetrasaccharide (alpha-D-Gal-1-->6[alpha-D-GalA-1-->4]-alpha-D-Man-1-->5Kdo), strongly suggesting that the transposon insertion disrupted a locus involved in O-antigen biosynthesis. Five monosaccharides (Glc, Man, GalA, 3-O-Me-6-deoxytalose, and Kdo) were identified as the components of the repeating O unit of the smooth parent strain, KIM5s. Strain KIM-G1 was complemented with a 7.2-kb DNA fragment from KIM5s that, when provided in trans on a broad-host-range vector, restored the smooth LPS and the full capacity of nodulation and fixation on its host Phaseolus vulgaris. The mTn5 insertion in KIM-G1 was located at the N terminus of a putative alpha-glycosyltransferase, which most likely had a polar effect on a putative beta-glycosyltransferase located downstream. A third open reading frame with strong homology to sugar epimerases and dehydratases was located upstream of the insertion site. The two glycosyltransferases are strain specific, as suggested by Southern hybridization analysis, and are involved in the synthesis of the variable portion of the LPS, i.e., the O antigen. This newly identified LPS locus was mapped to a 680-kb plasmid and is linked to the lpsbeta2 gene recently reported for R. etli CFN42.

Base Sequence↗

PubMed1999

Vasopressin receptors in human adrenal medulla and pheochromocytoma.

The nature of vasopressin (VP) receptors present in normal and tumoral human adrenal was investigated using various experimental approaches. Specific VP-binding sites were detected by autoradiography using [3H]arginine VP as a radioligand in adrenal cortex and medulla. The V1a receptor subtype was expressed in the two parts of the gland, as shown by pharmacological studies and RT-PCR experiments. By contrast, the V1b receptor subtype was only expressed in medullary chromaffin cells. This was confirmed by the characterization of V1b transcripts detected in adrenal medulla tissues. In pheochromocytoma, we also detected functional V1b receptors. These receptors triggered intracellular calcium mobilization from intracellular pools and were involved in catecholamine secretion. Binding experiments performed on pheochromocytoma plasma membrane preparations also revealed V1a vasopressin-binding sites, whose roles and cellular localization have not yet been determined. RT-PCR experiments confirmed these data; 100% and 80% of the five tumors tested exhibited V1a and V1b transcripts, respectively. Perifusion experiments also demonstrated that some pheochromocytomas may secrete large amounts of VP. Our findings imply that VP locally secreted by human adrenal medulla may regulate adrenal function by acting on V1a or V1b receptors. More interestingly, we demonstrate that one pheochromocytoma oversecretes VP. In this particular case, this may contribute to the increase in blood pressure observed.

Adrenal Gland Neoplasms↗