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Biomedical subjects

B Dupuy

Publications and source records attributed to B Dupuy.

At least 19 recordsLinked to original sources
PubMed1999

Relationships between striatal dopamine denervation and frontal executive tests in Parkinson's disease.

Indirect evidence from human and monkey investigations supports the idea that impaired frontal tasks in Parkinson's disease (PD) may result from striato-frontal disruption caused by dopamine (DA) denervation of the caudate nucleus. To directly investigate this hypothesis, we used PET with 11C-S-Nomifensine (11C-S-NMF), a sensitive marker of striatal DA denervation, in 10 non-demented PD patients in whom two frontal executive tests, the object alternation (OA) and the conditional associative learning (CAL) tasks, thought to reflect mainly set-shifting/inhibition and planning, respectively, were given. In addition, the central executive function of verbal working memory was assessed with the Brown Peterson paradigm (BPP). We found a highly significant correlation between right caudate 11C-S-NMF specific binding and OA performance, less significant and reverse-direction correlations between CAL performance and putamen 11C-S-NMF binding, and no significant correlation with BPP performance. Thus, caudate DA denervation may subtend poor set-shifting/inhibition process in PD. Our results also point to distinct and complex relationships between striatal DA and specific frontal tasks.

Adult↗
PubMed1998

Report of the European DNA Profiling Group (EDNAP)--an investigation of the hypervariable STR loci ACTBP2, APOAI1 and D11S554 and the compound loci D12S391 and D1S1656.

This paper describes the results of three collaborative exercises which continues the EDNAP theme to explore whether uniformity of DNA profiling results could be achieved between European laboratories using STRs. In an earlier exercise, complex hypervariable AAAG-repeat STR loci were investigated, but reproducibility was found to be poor because of the variation of techniques used by participating laboratories. In the exercise reported here, an internal allelic ladder composed of ACTBP2 and D11S554 fragments was distributed. This ladder was used to size ACTBP2 analysed by a "singleplex" PCR amplification and D11S554 combined with APOAI1 in a separate "duplex" reaction. Laboratories were asked to test 7 blood stains, one of which was a known control, and to report the results to the co-ordinating laboratory. The exercise demonstrated that ACTBP2 showed good reproducibility between laboratories, whereas further testing would be needed to validate APOAI1 and D11S554 for interlaboratory comparisons. In separate exercises, the simple loci D12S391 and D1S1656 were tested; both of these showed excellent reproducibility between laboratories.

Alleles↗
PubMed1998

Reproducibility of mtDNA analysis between laboratories: a report of the European DNA Profiling Group (EDNAP).

The aim of this collaborative exercise was to determine whether uniformity of mtDNA sequencing results could be achieved among different EDNAP laboratories. Laboratories were asked to sequence mtDNAHV1 region (16024-16365) from three bloodstains, proceeding in accordance with the protocol and strategies currently used in each individual laboratory. Cycle sequencing was used by 11 laboratories and solid phase single stranded sequencing was used by one laboratory. Different PCR strategies and PCR conditions were used by the different laboratories. Three laboratories used semi-nested PCR, two nested PCR, three direct amplification of HV1 and four amplification of overlapping fragments covering the HV1 region. Despite the diversity of methodologies used, all the laboratories reported the same results. The successful result of this exercise shows that PCR based mtDNA typing by automated sequencing is a valid, robust and reliable means of forensic identification despite the different strategies and methodologies used by the different laboratories.

DNA Fingerprinting↗
PubMed1998

Quantification of the inflammatory response in exudates to three polymers implanted in vivo.

Flow cytometry was used to quantify an inflammatory reaction in vivo as a new approach to evaluating the biocompatibility of biomaterials. The exudate formed inside cylindrical tubes composed of polyvinyl chloride (PVC), silicone elastomer (SIL), or polyurethane (PU) implanted subcutaneously in the dorsal region of rats was collected over a 3-week period. The volume, number of cells, and concentration of fibrinogen were determined in the exudate for the three biomaterials. The exudate was analyzed using a flow cytometry technique after labeling of the leukocytes with a monoclonal anti-CD45 antibody. Fibrinogen rose progressively over the 3-week period for the three polymers. After the different leukocyte lines were identified in rat blood samples, their determination in the exudate revealed differences among the three biomaterials. At day 2, PVC induced a predominantly neutrophilic inflammatory reaction whereas PU and SIL gave a mixture of monocytes and neutrophils. At day 9, the aspect of the cytograms was different, but the identification of the subpopulations was still possible. At day 23, the number of cell events became too low to distinguish the subpopulations. An even more detailed approach might be possible using specific labeling for each leukocyte line to establish a comparison among the three biomaterials. Flow cytometry associated with histomorphometric assessment might provide a precise quantitative in vivo test for determining the biocompatibility of materials.

Animals↗
PubMed1998

Evaluation of proliferation and protein expression of human bone marrow cells cultured on coral crystallized in the aragonite of calcite form.

The two crystalline forms of CaCO3, aragonite (from natural coral) and calcite (from natural limestone), have been used with success as bone graft substitutes. However, natural coral transformed into calcite by heating has never been tested. The objective of this work was to study the proliferation and alkaline phosphatase, osteonectin, and osteocalcin expression of human bone marrow cells cultured on CaCO3 crystallized both in the aragonite form (natural coral) and in the calcite form (natural coral modified by heating). The methods used to characterize calcite obtained from the coral were volumic porosimetry, scanning electron microscopy (SEM) and X-ray diffraction. Cell colonization of the material was assessed by SEM performed on days 1, 7, 20, and 30 and [3H]thymidine incorporation was performed on days 3, 7, 12, 18, 25, and 32. Phenotypic expression was assessed by using in situ cytochemistry (alkaline phosphatase), immunocytochemistry (osteonectin and osteocalcin), and hybridization (osteocalcin, beta-actin, and alkaline phosphatase mRNA). Results showed the transformation of aragonite into calcite after heating, the conservation of macroporosity, and a modification of the surface. Calcite appeared to have a smoother and more uniform surface than aragonite crystals. As for [3H]thymidine there was an increase incorporation from days 3 to 18, a stabilization from days 18 to 25, and a decrease from days 25 to 32. After 20 days of culture, immunological studies using monoclonal antibodies to osteocalcin, osteonectin, cytochemical analysis of alkaline phosphatase activity, and in situ hybridization using osteocalcin, beta-actin, and alkaline phosphatase cDNA indicated that the cells had not lost their osteoblastic phenotype. These experiments demonstrate that coral crystallized in the aragonite or calcite form present a similar degree of specific cytocompatibility.

Animals↗
PubMed1998

Cellular biocompatibility and resistance to compression of macroporous beta-tricalcium phosphate ceramics.

The main problem for macroporous structures used as bone substitutes is their lower resistances when compared to that of cancellous bone. The present investigation aimed to improve the strength of ceramics with 65% porosities based on beta-TCP. The initial mixtures were rendered plastic by addition of non-ionic carbohydrate binders. Macropores were created using substances which were eliminated by heat. Mechanical tests indicated that the resistance of the ceramics depended more on the quantity than the nature of the binders. Porosity measurements were done with a mercury porosimeter, and cellular biocompatibility was evaluated by performing cellular attachment tests and observing the proliferation of differentiated cells.

Biocompatible Materials↗
PubMed1998

Regulated transcription of Clostridium difficile toxin genes.

The Clostridium difficile toxA and toxB genes, encoding cytotoxic and enterotoxic proteins responsible for antibiotic-associated colitis and pseudomembranous colitis, were shown to be transcribed both from gene-specific promoters and from promoters of upstream genes. However, the gene-specific transcripts represented the majority of tox gene mRNAs. The 5' ends of these mRNAs were shown to correspond to DNA sequences that had promoter activity when fused to the Escherichia coli beta-glucuronidase (gusA) gene and introduced into C. perfringens. The appearance of tox mRNA in C. difficile was repressed during exponential growth phase but increased substantially as cells entered stationary phase. When glucose or other rapidly metabolizable sugars were present in the medium, the stationary phase-associated induction was inhibited, indicating that the toxin genes are subject to a form of catabolite repression. This glucose effect was general to many toxinogenic strains having varying levels of toxin production.

Bacterial Toxins↗
PubMed1998

"In vitro" dissolution of coral in peritoneal or fibroblast cell cultures.

Previous studies have shown that in vivo coral resorption involves a biphasic process: First, the edges of the coral block become powdery, then extracellular fluid and phagocytosis contribute to the dissolution of the crystals. The authors examined some types of cells that could be involved in phagocytosis, particularly the ability of both dermal fibroblasts and mouse-resident peritoneal cells to phagocytose and dissolve coral powder "in vitro". Radioactive coral was incubated for 24, 48, or 72 hrs with cells in the presence or absence of cytochalasin B (a phagocytic inhibitor) or chloroquine (a lysosomotropic agent). Furthermore, to specify the role of crystal cell contacts in the solubilization process, they incubated radioactive coral in conditioned media (obtained from two-day human fibroblastic or macrophagic cell culture in the presence or absence of non-radioactive coral) or at a distance from the cells using culture inserts. Measurements of the radioactivity in the different supernatants were performed. Transmission electron microscopy was carried out on the cells cultivated in the presence or absence of radioactive coral. The data suggest that both fibroblasts and macrophages dissolve the coral, and that the intracellular degradation in phagolysosomes is one of the mechanisms explaining coral powder dissolution.

Animals↗
PubMed1997

Report of the European DNA profiling group (EDNAP): an investigation of the complex STR loci D21S11 and HUMFIBRA (FGA).

This paper describes a collaborative exercise which was intended to demonstrate whether uniformity of DNA profiling results could be achieved between European laboratories using two complex short tandem repeat (STR) loci. The loci D21S11 and HUMFIBRA (FGA) were chosen because they are commonly used by different European laboratories. D21S11 has approximately 14 common alleles (f > 0.001), whereas HUMFIBRA has 19 common alleles. Laboratories were asked to test seven blood stains, one of which was a known control, and to report the results to the coordinating laboratory. The exercise demonstrated that complex STRs were amenable to standardisation.

Alleles↗
PubMed1997

Clostridium perfringens urease genes are plasmid borne.

Although many bacteria are ureolytic, and in some cases urease acts as a virulence factor, the urease phenotype has not been analyzed in the anaerobic pathogen Clostridium perfringens. In this study, approximately 2% of C. perfringens strains, representing the principal biotypes, were found to harbor the urease structural genes, ureABC, and these were localized on large plasmids that often encode, in addition, the lethal epsilon or iota toxins or the enterotoxin. This represents the first report of a plasmid-encoded urease in a gram-positive bacterium. The C. perfringens enzyme was highly similar to the ureases of other bacteria and cross-reacted with antibodies raised against the urease purified from Helicobacter pylori. Urease production was inhibited by urea and induced under growth conditions where the availability of nitrogen sources was limiting. To date, this form of regulation has been observed only for chromosomal ureABC genes.

Amino Acid Sequence↗
PubMed1996

Genome mapping of Clostridium perfringens strains with I-CeuI shows many virulence genes to be plasmid-borne.

The intron-encoded endonuclease I-CeuI from Chlamydomonas eugametos was shown to cleave the circular chromosomes of all Clostridium perfringens strains examined at single sites in the rRNA operons, thereby generating ten fragments suitable for the rapid mapping of virulence genes by pulsed-field gel electrophoresis (PFGE). This method easily distinguishes between plasmid and chromosomal localisations, as I-CeuI only cuts chromosomal DNA. Using this approach, the genes for three of the four typing toxins, beta, epsilon, and tau, in addition to the enterotoxin and lambda-toxin genes, were shown to be plasmid-borne. In a minority of strains, associated with food poisoning, where the enterotoxin toxin gene was located on the chromosome, genes for two of the minor toxins, theta and mu, were missing.

Bacterial Toxins↗
PubMed1996

A two-step embedding process for better preservation of soft tissue surrounding coral implants.

Infiltration of paraffin or embedding polymers proceeds more quickly in soft than in mineralized tissue specimens (bone or biomaterial). The proposed method takes advantage of this difference to protect soft tissue from the action of decalcifying agents. After embedding a bone-soft-tissue sample in a hydrophobic polymer, it is cleared of the resin on one of its sides to permit access of the decalcifying solution to the mineral component. A second infiltration with another polymerizable solution that cures in the empty pores created by the dissolution makes it possible to obtain a homogeneous hard block, make thin slices, and perform enzyme histochemistry and immunostaining on well-preserved soft tissue with most of its antigenic and enzymatic properties intact.

Animals↗
PubMed1996

Influence of the structure of three corals on their resorption kinetics.

The aim of this study was to investigate the influence of the structure of corals on their resorption kinetics after implantation in subcutaneous areas. Three types of coral (Porites astreoides, Montastrea annularis and Dichocoenia stokesi) identical in composition but different in structure were implanted for periods of 1 and 2 months in subcutaneous sites in OF1 mice. The resorption of the implants was studied by means of qualitative (histology, scanning electron microscopy, fluorochrome labelling method) and quantitative approaches (gravimetric method). The results of the qualitative study revealed a process of irregular deterioration of the coral, linked to the detachment of crystals at the surface of the implant. The results of the quantitative study showed that the speed of resorption increases with the implantation time and the open porosity of the coral. These reactions are explained by the increase of the surface exchange area in contact with factors responsible for resorption: biological medium and cells. When considering the choice of coral as a bone substitute, these factors must be taken into account to allow the in situ maintenance of the implant over a sufficiently long period of time according to the clinical situation.

Animals↗
PubMed1995

The resorption of bone-implanted corals varies with porosity but also with the host reaction.

Three different exoskeletons of coral species Porites asteroides (P), Montastrea annularis (M), and Dichocoenia stokesi (D) were implanted for 2-20 weeks in rabbits. At 2, 4, 8, or 20 weeks, the exoskeletons presented variations in their resorptions depending on the species. To understand the variations in the decreasing speed of the implants despite their similar chemical composition, a study of the surface and architecture of the coral was carried out using scanning electronic microscopy, porosity was evaluated, and growth and differentiation of osteogenic cells cultured in vitro were observed for more than 1 month. At the cellular level, the surface of the implants was identical. Three-dimensional structures of the implants were variable, but the porosity values [P = 42.7%, M = 40.7%, and D = 17.4%] could not completely account for the differences in the resorbing process of the species. Standard histologic studies performed at 2, 4, 8, and 20 weeks after implantation produced the same pattern with P or M, showing aspects of rapid resorption; however, with D there were images resembling those of a foreign-body reaction. It seems that when resorption is not quick enough, a foreign body reaction develops which further slows down the process. This work focuses on the importance of porosity when using coral as bone substitute.

Animals↗
PubMed1995

Cholate-induced disruption of calcitonin-loaded liposomes: formation of trypsin-resistant lipid-calcitonin-cholate complexes.

PURPOSE: The work was performed to obtain a better understanding why the oral administration of calcitonin (CT)-loaded liposomes to rats results in a hypocalcemia, while liposomes are normally disrupted in the gastro-intestinal tract and cannot protect the hormone from enzymatic digestion. METHODS: In vitro comparisons between the stability of calcein and CT-loaded liposomes in the presence of cholate solutions led to an interpretation of the results observed. By means of gel filtration, turbidimetry, and fluorescence measurements, the interactions between CT and lipids were studied after sonicated liposomes had been broken down by cholate. RESULTS: Experiments showed that CT in the external medium of a liposome suspension had no effect on the vesicles. Gel filtration of cholate-treated liposomes loaded with calcein and CT resulted in a total separation of calcein from the lipid fraction for detergent concentrations higher than 4 mM. However, 50% of the CT was reencapsulated even when the cholate-to-phospholipid molar ratio was increased up to 100. Incubation of cholate-solubilized liposomes with 1% trypsin resulted in a partial CT-breakdown. CONCLUSIONS: These results strongly suggest that during membrane solubilization by cholate, lipid-CT complexes are formed which retain most of the CT initially embedded in the liposomal membrane, and which offer some protection to CT under the action of trypsin. The existence of these complexes could be one of the reasons for the reported hypocalcemia in rats after oral administration of CT-loaded liposomes.

Animals↗
PubMed1995

Rapid expansion of the physical and genetic map of the chromosome of Clostridium perfringens CPN50.

The physical map of the 3.6-megabase chromosome of Clostridium perfringens CPN50 was extended by positioning sites for the endonucleases SfiI and I-CeuI, and in parallel, the gene map was expanded by using a genome scanning strategy. This involved the cloning and sequencing of random chromosomal fragments, identification of the functions of the putative genes by database searches, and then hybridization analysis. The current gene map comprises almost 100 markers, many of which encode housekeeping functions while others are involved in sporulation or pathogenesis. Strikingly, most of the virulence genes were found to be confined to a 1,200-kb segment of the chromosome near oriC, while the pleiotropic regulatory locus, virRS, was situated toward the putative replication terminus. A comparison of the gene maps of three endospore-forming bacilli, C. perfringens, Clostridium beijerinckii, and Bacillus subtilis, revealed a similar order and distribution of key sporulation and heat shock genes which might reflect an ancient evolutionary relationship.

Chromosome Mapping↗
PubMed1994

Calcitonin-loaded liposomes: stability under acidic conditions and bile salts-induced disruption resulting in calcitonin-phospholipid complex formation.

Calcitonin-loading in liposomes composed of phosphatidylcholine, cholesterol and stearylamine or dipalmitoyl phosphatidylglycerol was studied at low pH values and in the presence of bile salts to check whether liposomal entrapment could be a possible means of protecting the peptide against the aggressive conditions present in the gastrointestinal tract. The association of calcitonin with the lipidic vesicles was monitored using radioactive labelling of the peptide and gel-filtration separation of the free and liposome-associated fractions. The results show that for all phospholipid compositions tested, loading was preserved in light acidic or basic buffers, and that only a slight disruption was observed at pH 2.5. Cholate caused a significant but only partial release of calcitonin even when the cholate-to-phospholipid ratio was increased. To understand the mode of calcitonin entrapment in the vesicles, the release of liposome-entrapped calcein was monitored concomitantly and taken as a stability criterion. Liposome integrity appears to be resistant at low pHs but to be totally destroyed by 4 mM cholate in a manner quasi-independent of the phospholipid concentration. These results strongly suggest that bile salts induce a disruption of the liposomes which results in the formation of new lipidic structures involving calcitonin and probably cholate.

Bile Acids and Salts↗