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B Fournier

Publications and source records attributed to B Fournier.

At least 19 recordsLinked to original sources
PubMed1999

Strength and regulation of the different promoters for chromosomal beta-lactamases of Klebsiella oxytoca.

The two groups of chromosomal beta-lactamases from Klebsiella oxytoca (OXY-1 and OXY-2) can be overproduced 73- to 223-fold, due to point mutations in the consensus sequences of their promoters. The different versions of promoters from blaOXY-1 and blaOXY-2 were cloned upstream of the chloramphenicol acetyltransferase (CAT) gene of pKK232-8, and their relative strengths were determined in Escherichia coli and in K. oxytoca. The three different mutations in the OXY beta-lactamase promoters resulted in a 4- to 31-fold increase in CAT activity compared to that of the wild-type promoter. The G-->T transversion in the first base of the -10 consensus sequence caused a greater increase in the promoter strength of the wild-type promoter than the two other principal mutations (a G-to-A transition of the fifth base of the -10 consensus sequence and a T-to-A transversion of the fourth base of the -35 sequence). The strength of the promoter carrying a double mutation (transition in the Pribnow box and the transversion in the -35 hexamer) was increased 15- to 61-fold in comparison to that of the wild-type promoter. A change from 17 to 16 bp between the -35 and -10 consensus sequences resulted in a ninefold decrease of the promoter strength. The expression of the blaOXY promoter in E. coli differs from that in K. oxytoca, particularly for promoters carrying strong mutations. Furthermore, the blaOXY promoter appears not to be controlled by DNA supercoiling or an upstream curved DNA, but it is dependent on the gene copy number.

Base Sequence↗
PubMed1998

DNA complexes obtained with the integron integrase IntI1 at the attI1 site.

Integrons are genetic elements that are able to capture genes by a site-specific recombination mechanism. Integrons contain a gene coding for a lambda-like integrase that carries out site-specific recombination by interacting with two different target sites; the attI site and the palindromic sequence attC (59 base element). Cassette integrations usually involve the attI site, while cassette excisions use attC . Therefore, the integrase should bind both sites to cleave DNA and perform site-specific recombination reactions. We have used purified maltose-binding protein fused with the integrase (MBP-IntI1) and native IntI1 protein and gel retardation assays with fragments containing the complete and partial attI1 site to show formation of four complexes in this region. Chemical modification of specific nucleotides within the attI1 site was used to investigate their interference with binding of the integrase protein. We attribute IntI1 specific binding to four regions in the attI1 site and a GTTA consensus sequence is found in three of the four regions. Interference by modified guanine and thymine residues in the DNA major groove and adenine residues in the minor groove were observed, indicating that the integrase interacts with both sides of the helix. Binding of IntI1 to attC is also discussed.

ATP-Binding Cassette Transporters↗
PubMed1998

[Long-term follow-up of 9 islets of Langerhans autografts after resection of the pancreas].

INTRODUCTION: Preservation of physiological endocrine pancreatic function represents a major problem in pancreatectomised patients. In 40 to 100%, pancreatic resection can result in diabetes, which is often difficult to manage. Islet autotransplantation has been proposed to prevent this severe metabolic consequence. PATIENTS AND METHODS: Between March 1992 and January 1997 we performed 9 human islet autotransplantations in Geneva. The patients comprised 6 males and 3 females aged 40 to 81 years (median: 51 years). The pancreatectomy was total in 3 cases, subtotal (95%) in 2 cases and partial (40-80%) in the others. Indications for resection were alcohol-induced chronic pancreatitis (6 cases) and focal benign pancreatic pathologies (3 cases). After collagenase digestion, unpurified islets were injected intraportally and embolised into the liver. Patient metabolic status was regularly tested by 24-hour serum glucose profile, measurement of glycosylated haemoglobin, oral and intravenous glucose tolerance tests and glucagon stimulation test. RESULTS: Immediately after autotransplantation, one patient had persistent insulin-dependent diabetes. Among the 8 patients who were insulin-independent soon after the graft, 4 presented a progressive deterioration of endocrine pancreatic function and required insulin therapy 5, 8, 24 and 36 months after the graft. Currently, 4 patients are insulin-independent: three of them have normal glucose tolerance tests (24, and 48 months after the graft) and the last presented with glucose intolerance 22 months after the graft but still does not require exogenous insulin. CONCLUSION: Islet autotransplantation can be considered a useful therapeutic option serving to prevent or delay the occurrence of surgically-induced diabetes. In our opinion, islet autotransplantation should be offered to any non diabetic patient needing to undergo major pancreatic resection.

Adult↗
PubMed1998

[Isolation of the pig islets of Langerhans: evaluation of in vitro and in vivo function].

Pig islets are considered the best alternative to human islets in the treatment of insulin-dependent diabetes. Pigs could represent a potential islet donor for xenotransplantation in humans because of the close similarity between human and porcine insulin and the theoretically unlimited availability of porcine pancreas. From November 1991 to January 1997 we performed 221 pig islet isolations from 3 pig sources: group 1: minipigs (age 9-18 months) and white pigs (3-8 months), group 2: large white pigs (5-8 months), group 3: large white pigs (12-24 months). Islets were isolated according to a semi-automated method using enzymatic digestion and purification through discontinuous Euro-Ficoll gradients. The pancreases were surgically removed in our laboratory for group 1, while pancreases from groups 2 and 3 were removed at the slaughterhouse with an average warm ischemia time of 15 minutes. In vitro islet function was assessed by static incubations and perifusions, and in vivo islet function by transplantation under the kidney capsule of nude diabetic mice. The results were as follows: [table: see text] Insulin secretion increased twofold after in vitro glucose stimulation. We obtained restoration of euglycemia in diabetic mice which survived > 3 months after the graft and returned to diabetes after nephrectomy. This study shows that our isolated pig islets are viable and functional in vitro and in vivo after transplantation.

Animals↗
PubMed1998

Prolactin stimulation of phosphoinositide metabolism in CHO cells stably expressing the PRL receptor.

PRL receptor (PRL-R) activation by PRL triggers a cascade of intracellular events including homodimerization of the receptor, activation of cytoplasmic receptor-associated tyrosine kinase and tyrosine-phosphorylation of various signal transducers. In CHO cells, transfected with the long form of PRL-R, an increase in [Ca2+]i was observed following PRL stimulation whereas Ca2+ is generally coupled with the phosphoinositide metabolism. In this study, we investigated phosphoinositide involvement in the PRL transduction pathway. We report that PRL induces rapid increases in two novel inositol phospholipids, almost certainly PtdIns(4,5)P2 and PtdIns(3,4,5)P3. Pre-traitment of CHO cells with wortmanin, a specific PtdIns3-kinase inhibitor, considerably reduces the PRL-induced increase in PtdInd(3,4,5)P3, thus suggesting an involvement of this enzyme in the cascade of activation of cytoplasmic kinase proteins. A pathway beginning with the activation of PtdIns3-kinase, phosphorylation of PtdIns(4,5)P2 and rapid synthesis of PtdIns(3,4,5)P3 is proposed. PtIns(3,4,5)P3 may acts as a lipid second messenger, directly or indirectly responsible for some of the multiple cell changes attributed to PRL.

Androstadienes↗
PubMed1998

Pharmacological studies of proctolin receptors on foregut and hindgut of Blaberus craniifer.

Proctolin (Arg-Tyr-Leu-Pro-Thr) and proctolin analogs modified at position 1, 2, or 5 caused dose dependent contractions of Blaberus fore- and hindgut. The varying contractile effects between both tissues revealed the possible presence of receptor subtypes as identified by [GABA1]-proctolin. A single population of binding sites (Kd approximately 100 nM) was deduced from Scatchard analysis. In addition, nanomolar concentrations of proctolin induced a dose-dependent hydrolysis of phosphoinositides (PIns) augmented by GTPgammaS (1 microM) on foregut membranes but no accumulation of cAMP. Proctolin induced contractions are likely mediated via a phospholipase C linked to a heptahelical receptor bound to heterotrimeric G-proteins.

Adenylyl Cyclases↗
PubMed1998

Isolation, sequence determination, physical and physiological characterization of the neuroparsins and ovary maturing parsins of Schistocerca gregaria.

Neurosecretory products immunologically related to either neuroparsin (NP) or ovary maturing parsin (OMP) of Locusta migratoria (Lom) were purified from the nervous corpora cardiaca of Schistocerca gregaria (Scg). The determination of both their molecular masses by mass spectrometry and their sequences by automated Edman degradation established that they are members of the NP and OMP families respectively. NP molecules of Schistocerca (Scg NPs) consisted of two major forms having about the same molecular masses as NPA and NPB of Locusta and 88% primary structure similarity. They had also the same antidiuretic activity. OMP molecules of Schistocerca (Scg OMPs) were composed in young adults of four isoforms: two long isoforms corresponding to Lom OMP, and differing by a tripeptide insertion (Pro-Ala-Ala) at position 21 and two short isoforms deprived of the 13-residue N-terminal peptide of Lom OMP and differing by the same tripeptide insertion. The PAA isoforms were observed in low amounts as compared to the other isoforms. In mature adults, only the two short isoforms were present. The complete sequence of PAA Scg OMP presents a large degree of sequence homology with Lom OMP (83%). The mixed Scg OMPs had the same biological effects as Lom OMPs. They induced precocious occurrence of both ecdysteroids and vitellogenin in the haemolymph and stimulated oöcyte growth.

Aging↗
PubMed1998

Mutations in topoisomerase IV and DNA gyrase of Staphylococcus aureus: novel pleiotropic effects on quinolone and coumarin activity.

Previous studies have shown that topoisomerase IV and DNA gyrase interact with quinolones and coumarins in different ways. The MICs of coumarins (novobiocin and coumermycin) for MT5, a Staphylococcus aureus nov mutant, are higher than those for wild-type strains. Sequencing the gyrB gene encoding one subunit of the DNA gyrase revealed the presence of a double mutation likely to be responsible for this resistance: at codon 102 (Ile to Ser) and at codon 144 (Arg to Ile). For single-step flqA mutant MT5224c9, previously selected on ciprofloxacin, the fluoroquinolone MIC was higher and the coumarin MIC was lower than those for its parent, MT5. Sequencing the grlB and grlA genes of topoisomerase IV of MT5224c9 showed a single Asn-470-to-Asp mutation in GrlB. Genetic outcrosses by transformation with chromosomal DNA and introduction of plasmids carrying either the wild-type or the mutated grlB gene indicated that this mutation causes both increased MICs of fluoroquinolones and decreased MICs of coumarins and that the mutant grlB allele is codominant for both phenotypes with multicopy alleles. Integration of these plasmids into the chromosome confirmed the codominance of fluoroquinolone resistance, but grlB+ appeared dominant over grlB (Asp-470) for coumarin resistance. Finally, the gyrA (Leu-84) mutation previously described as silent for fluoroquinolone resistance increased the MIC of nalidixic acid, a nonfluorinated quinolone. Combining the grlA (Phe-80) and grlB (Asp-470) mutations with this gyrA mutation also had differing effects. The findings indicate that alterations in topoisomerases may have pleiotropic effects on different classes of inhibitors as well as on inhibitors within the same class. A full understanding of drug action and resistance at the molecular level must take into account both inhibitor structure-activity relationships and the effects of different classes of topoisomerase mutants.

4-Quinolones↗
PubMed1998

Effects of mutations in GrlA of topoisomerase IV from Staphylococcus aureus on quinolone and coumarin activity.

The grlA genes of Staphylococcus aureus ISP794 (wild type), MT5224c4 (grlA [Phe-80]), MT5224c2 (grlA [Pro-116]), and MT111 (grlA [Glu-116]) were cloned in pSK950, a shuttle vector, and introduced into S. aureus strains derived from strain RN4220. The mutations at position 116 of GrlA (Ala-->Pro or Glu) caused an increase in the level of fluoroquinolone resistance and a decrease in the level of coumarin susceptibility, whereas the mutation at position 80 (Ser-->Phe) caused only an increase in the level of fluoroquinolone resistance. In multicopy alleles, both types of mutations were codominant for fluoroquinolone resistance, and mutations at position 116 were also codominant for coumarin resistance.

Alleles↗
PubMed1998

Human islet allotransplantation: world experience and current status.

Currently type-I diabetes mellitus is treated with exogenous insulin administration, but traditional insulin therapy does not prevent long-term systemic complications and therefore alternatives should be sought. Presently the only option is to substitute the insulin-producing beta cells in order to obtain a more physiological way to deliver insulin. Simultaneous kidney and pancreas transplantation is an established procedure. Pancreas transplantation, however, is a major surgical procedure with a high rate of complications. Transplantation of the isolated insulin-secreting islets of Langerhans is an alternative approach, which is easier and safer than whole organ transplantation. Since 1990, clinical trials of islet transplantation have begun in a few specialized centers worldwide and the International Islet Registry counting 305 human islet allografts at the end of 1995. Insulin independence at 1 year was achieved in 8% of the patients, but 20% of cases showed a graft function with a normal basal C peptide and improved glycemic regulation. We review here the different aspects of human islets of Langerhans allotransplatation, namely historical aspects, indications, isolation and purification procedures and the results obtained.

Cryopreservation↗
PubMed1998

Biological effects of eleven combined oral contraceptives on serum triglycerides, gamma-glutamyltransferase, alkaline phosphatase, bilirubin and other biochemical variables.

The objectives of this paper are to update and quantify the biological effects of the most commonly used oral contraceptives (OC) on 15 biochemical tests currently determined in clinical laboratories and to compare these effects between the different types of OC. The sample population was constituted by 1604 women using combined OCs and the control group comprised 3466 women in the same age range not taking medication. Women taking OC were divided into 11 groups according to the estrogen/progestogen combination. The effects of OCs were studied after adjustment for age, weight, height, body mass index and alcohol and tobacco consumption. The changes observed with the new progestogens were less important than in the past. In comparison with the controls, the mean serum triglyceride concentration was significantly increased by +8.5% to +36.0% (p<0.05 to p<0.001) in each group while those of total cholesterol and gamma-glutamyltransferase were increased only in 3 and 4 estrogen/progestogen combinations respectively. Conversely, the mean concentrations of alkaline phosphatase, total bilirubin, phosphate and albumin were significantly decreased. Using a discriminant analysis, three main groups according to the type of progestogen were defined: cyproterone acetate, DL-norgestrel and levonorgestrel, and all other progestogens. The changes in serum triglyceride concentration induced by OC intake must be considered by the clinician and are useful for taking a clinical and risk decision in an individual woman.

Adolescent↗
PubMed1998

Prolactin induces an inward current through voltage-independent Ca2+ channels in Chinese hamster ovary cells stably expressing prolactin receptor.

There is still only limited understanding of the early steps of prolactin (PRL) signal transduction in target cells. Recent studies have identified some of the essential first steps: these include the rapid association of the PRL receptor with JAK tyrosine kinases and tyrosine phosphorylation of a number of proteins, including members of the signal transducer and activator of transcription (Stats) family. On the other hand, binding of PRL to its receptor is rapidly followed by calcium influx. However, PRL-induced ionic events and the related ionic channels involved have not been clearly established. This work was undertaken to characterise the channels responsible for calcium influx and to obtain an insight into their activation processes. Using the patch-clamp technique in the cell-attached configuration, single Ca2+ channel currents were recorded following PRL application (10 nM) in Chinese hamster ovary (CHO) cells stably expressing PRL receptor (CHO-E32). Statistical analysis showed that the recorded currents were voltage-independent, with a slope conductance of 16 pS. Although these channels were present in excised patches, the fact that PRL was unable to activate them suggested that a soluble cytoplasmic component may be required. Application of the purified inositol phosphate, Ins(1,3,4,5)P4 (2 microM), to the inside of the excised patch membrane activated the voltage-independent 16 pS Ca2+ channel. The open probability (Popen) was enhanced. The inositol phosphates Ins(1,2,3,4,5)P5 and Ins(1,4,5)P3 did not affect channel activity while InsP6 (20 microM) had some effect, although less marked than that of Ins(1,3,4,5)P4. Using the anion-exchange HPLC technique, we then studied the effects of PRL (10 nM) on the turnover of inositol phosphates (InsPs) in CHO-E32. Our studies showed that PRL induces rapid increases in the production of Ins(1,3,4,5)P4 (207% at 30 s), InsP5 (171% at 30 s), and InsP6 (241% at 30 s). Conversely, Ins(1,4,5)P3 showed a transient decrease at 5 s, accompanied by a concomitant increase in Ins(1,3,4,5)P4, suggesting that the former could be transiently phosphorylated to produce the latter. Comparison of the production kinetics of Ins(1,4,5)P3, Ins(1,3,4,5)P4, InsP5, and InsP6 indicated the possibility of additional metabolic routes which have yet to be determined. This study suggests that PRL promotes Ca2+ entry through voltage-independent Ca2+ channels that may be activated by Ins(1,3,4,5)P4 and InsP6.

Animals↗
PubMed1998

[Perceived health and pregnancy: study of several behavioral and environmental determinants among a population of pregnant women].

The aim of this study, based on data of the Centre for Preventive Medicine, is to compare the perceptions women have of their health within a population of pregnant women that was divided into two categories: one group of women who feel they are in a poor health, the others feel they are in good health. The study results seem to differentiate between both the psychological and physical profiles of the two groups of pregnant women. The women who are not followed by a doctor don't seem to have a more negative perception of their health status. It seems, in addition, that the negative perception of health status has not only psychological roots, but also behavioural and environmental ones.

Adolescent↗
PubMed1997

The human peroxisomal multifunctional protein involved in bile acid synthesis: activity measurement, deficiency in Zellweger syndrome and chromosome mapping.

The dehydrogenation of 24R,25R-varanoyl-CoA, the physiological intermediate formed during the peroxisomal breakdown of the bile acid intermediate trihydroxycoprostanic acid, was studied in human liver. The reaction appeared to be catalyzed by two different enzymes. A first one, present in the cytosol, did not discriminate between the four possible varanoyl-CoA isomers and did not require the CoA moiety. The second enzymic activity was associated with peroxisomes and acted only on the 24R,25R-isomer, in which the 24-hydroxy group possesses the D-configuration. The D-specific dehydrogenase is part of a 79 kDa protein which represents the human counterpart of a recently discovered second multifunctional protein in rat liver peroxisomes, named multifunctional protein 2 (MFP-2). Human MFP-2, like its rat counterpart, is also responsible for the formation (by hydratation) of 24R,25R-varanoyl-CoA. A deficiency of MFP-2 in Zellweger liver could be demonstrated immunologically by using antibodies against the rat enzyme and enzymically -- after removal of the cytosol -- by using 24R,25R-varanoyl-CoA. The gene coding for MFP-2 was mapped to chromosome 5q2.3.

3-Hydroxyacyl CoA Dehydrogenases↗
PubMed1997

Expression of neuroparsin cDNA in insect cells using baculovirus vectors.

The cDNA encoding neuroparsin A, a polytropic neurohormone of the locust, Locusta migratoria, was inserted into the genome of Autographa californica nuclear polyhedrosis virus such that transcription was under control of the p10 promoter. A polypeptide having the same charge and the same apparent molecular weight as the authentic neuroparsin A and that was reactive against neuroparsin immune serum was produced in recombinant virus-infected lepidopteran cell lines but not in control virus-infected cells. The baculovirus-expressed polypeptide was purified by two steps of liquid chromatography (anion exchange and reversed phase) which were previously used to purify the natural neuroparsin. The purified baculovirus-expressed polypeptide enhanced fluid reabsorption of everted rectum preparations, as did the natural neuroparsin. Thus, this gene expression system produced a polypeptide identical to authentic neuroparsin.

Animals↗
PubMed1997

Assessment of the relationship between broadband ultrasound attenuation and bone mineral density at the calcaneus using BUA imaging and DXA.

The purposes of this study was to determine the relationship between broadband ultrasound attenuation (BUA) and bone mineral density (BMD) measured at different regions of the calcaneus with identical site-matched regions of interest (ROIs). Dual-energy X-ray absorptiometry (DXA) measurements of the calcaneus and BUA imaging were performed in 30 women (15 premenopausal and 15 postmenopausal). Four square ROIs were located in the great tuberosity and one square ROI in the foramen calcaneus. A ROI adapted to the shape and size of the whole calcaneus was also considered. All ROIs were analyzed three times with both techniques to minimize intra-observer variability. The correlation coefficient between attenuation and frequency was used as an index of BUA measurement error. Before accepting a measurement of BUA in inhomogeneous material, it could be useful to map the spatial variations of the measurement error. In all ROIs we found the BUA and BMD were strongly related (r = 0.78-0.91, p < 0.001). The correlation between BUA and BMD was slightly higher in the inferior part of the posterior tuberosity than in the superior part and in the foramen calcaneus. The very high correlation between attenuation and frequency found in all ROIs (r = 0.99) suggests that measurement errors of propagation were probably not significant. Ultrasound imaging yields the opportunity for studying the spatial acoustic properties in the calcaneus and their relation to bone mass or structural parameters provided by independent imaging techniques. BUA measured with current transmission techniques reflects mainly bone mass, and microarchitecture to a smaller extent.

Absorptiometry, Photon↗
PubMed1997

Quantitative ultrasound imaging at the calcaneus using an automatic region of interest.

A new approach to measuring bone properties at the calcaneus using ultrasound parametric imaging has recently emerged. However, an additional source of observer-related error is the substantial regional variations in the pattern of ultrasound parameters. The contribution of intra-observer and inter-observer variability to the coefficient of variation can be eliminated using an algorithm which selects the region of interest (ROI) completely automatically. The objective of the present study was the clinical assessment of an automatic ROI for both broadband ultrasonic attenuation (BUA) and speed of sound (SOS) measurement using ultrasound parametric imaging. The automatic ROI was defined as the circular region of lowest attenuation in the posterior tuberosity of the calcaneus. We have tested this algorithm using clinical images of the calcaneus from 265 women. Mean coefficients of variation were 1.6% (95% confidence interval 1.4%-1.9%) and 0.26% (95% confidence interval 0.23%-0.32%) for BUA and SOS respectively (standardized CV was 2.1% for BUA and 2.6% for SOS). Z-scores in an osteoporotic group were -0.61 and -0.52 for BUA and SOS respectively. In healthy women, the age-related decline was -0.50 dB/ MHz per year (0.7%/year) for BUA and -1.2 m/s per year (0.08%/year) for SOS. In the subgroup of healthy postmenopausal women, using stepwise multiple regression, we found that BUA was predicted best by years since menopause (YSM) and weight, with overall model r2 = 0.28; SOS was predicted best by YSM only (r2 = 0.21). Neither the range of biological variation of ultrasound parameters nor the clinical value were affected by the choice of the region of lowest attenuation for measurement. The automatic procedure was totally independent of operator interaction, therefore excluding loss of precision due to intra- or inter-observer variability. The results showed the high precision and robustness of the procedure. These factors make this approach viable for routine clinical use.

Aged↗