Étudier les effets de l'hydroxyde de calcium et de la dermaseptine-1 sur Enterococcus faecalis et les cellules pulpaires

Les médications intra canalaires, comme l'hydroxyde de calcium, sont utilisées en endodontie pour désinfecter le système radiculaire. Mais, les traitements endodontiques peuvent échouer en raison d'une infection persistante causée principalement par la subsistance de certaines bactéries dans le canal dentaire. Pour contrôler la croissance bactérienne, l'usage de peptides antimicrobiens comme la dermaseptine semble être une alternative prometteuse. Les objectifs de ce projet de recherche étaient de comparer les propriétés antibactériennes de la dermaseptine S1 (DS1) et de l'Ultracal XS (hydroxyde de calcium) sur la croissance d'Enterococcus faecalis et leur compatibilité respective sur les cellules souches de la pulpe dentaire. Méthode : E. faecalis est cultivé en présence ou non d'Ultracal XS (Ultradent), d'hydroxyde de calcium, de sulfate de baryum, de propylène glycol ou de DS1 (10, 50 and 100 µm/mL). La croissance bactérienne est évaluée pendant 24 h par spectrophotométrie. Les effets des différents composés sur les cellules souches pulpaires furent évalués par microscope optique à 24 et 48 h et par test de MTT à 48 h. Les données furent collectées de 3 expériences et soumises à une analyse statistique ANOVA à une issue et au test de Tukey. Résultats : L'Ultracal XS diminue légèrement la croissance bactérienne, alors que l'hydroxyde de calcium pur, le sulfate de baryum et le propylène glycol n'ont pas eu d'effet sur la bactérie. La DS1, même à de faibles concentrations, a significativement réduit la croissance bactérienne. L'effet inhibiteur a été présent pendant 24 heures, avec des concentrations de DS1 de 50 et 100 µm/mL. De plus, l'Ultracal XS et ses constituants ont montré un effet toxique important sur l'adhésion et la croissance des cellules souches pulpaires, alors que la DS1 montre seulement un effet toxique modéré à des concentrations de 100 µm/mL. Conclusion : L'Ultracal XS diminue modérément la croissance d' E. faecalis alors que la DS1 la réduit de façon importante. Les effets toxiques au niveau des cellules souches pulpaires sont modérés pour la DS1 en comparaison avec l'Ultracal XS. Cette étude a démontré qu'il pourrait être intéressant d'utiliser les dermaseptines en médication intracanalaire en endodontie.Intracanal medication is used in endodontics to disinfect the root canal system, but sometimes endodontic treatments fail due to the persistence of infection. Antimicrobial peptides such as dermaseptins might be a good alternative to calcium hydroxide for intracanal medication. Endodontic treatments can fail due to the persistence of infection. Antimicrobial peptides might be a good alternative to calcium hydroxide for intracanal medication. This study aims to compare the antimicrobial properties of dermaseptin S1 (DS1) and Ultracal XS against E. faecalis growth and their compatibility with dental pulp stem cells (DPSC). Methods: E. faecalis cells were cultured in the presence or not of Ultracal XS (Ultradent), calcium hydroxide, barium sulfate, propylene glycol, or DS1 (10, 50, and 100 µm/mL). The bacterial growth was assessed over 24 h by spectrophotometry. The effect of the different chemicals on the shape and growth of DPSCs were evaluated using an optical microscope at 24 and 48 h and an MTT assay after 48 h. Data were collected from 3 experiments and subjected to statistical analysis using one-way ANOVA and Tukey's test. Results: Ultracal XS, slightly decreased the growth of bacteria, while calcium hydroxide, barium sulfate, and propylene glycol do not affect the growth of E. faecalis. Interestingly, DS1 significantly reduced the growth of E. faecalis. The inhibitory effect was present up to 24 h culture, with 50 and 100 ug/ml of DS1. Ultracal XS and its constituents have a high toxic effect on DPSC adhesion and growth, while DS1 showed moderate adverse effects only at high concentrations (100 µg/ml). Conclusions: Ultracal XS moderately, while DS1 highly decreased the growth of E. faecalis. The toxic effect of DS1 on DPSCs was moderate compared to Ultracal XS. This study demonstrated the potential use of dermaseptin as an intracanal medication

Expression in E. coli and characterization of the catalytic domain of Botrytis cinerea chitin synthase

<p>Abstract</p> <p>Background</p> <p>Chitin synthase 3a (CHS3a) from <it>Botrytis cinerea </it>(Bc) catalyses the multiple transfer of <it>N</it>-acetylglucosamine (GlcNAc) residues to the growing chitin chain. Chitin, a β-1,4 linked GlcNAc homopolymer, is an essential cell wall component of filamentous fungi. Chitin synthase, processive membranous protein, has been recognized as a promising target for new antifungicides. Enzymatic characterizations of chitin synthases have been limited, mainly because purity and amounts of integral enzyme obtained after purification procedures have not been sufficient.</p> <p>Findings</p> <p>We undertook the preparation of two BcCHS3a fragment proteins, containing only the central domain and devoid of the N-terminal and transmembrane C-terminal regions. The central domain of CHS3a, named SGC (Spsa GntI Core), is conserved in all UDP-glycosyltransferases and it is believed to contain the active site of the enzyme. CHS3a-SGC protein was totally expressed as inclusion bodies in <it>Escherichia coli</it>. We performed recombinant CHS3a-SGC purification in denaturing conditions, followed by a refolding step. Although circular dichroism spectra clearly exhibited secondary structures of renatured CHS3a-SGC, no chitin synthase activity was detected. Nevertheless CHS3a-SGC proteins show specific binding for the substrate UDP-GlcNAc with a dissociation constant similar to the Michaelis constant and a major contribution of the uracil moiety for recognition was confirmed.</p> <p>Conclusions</p> <p>Milligram-scale quantities of CHS3a-SGC protein with native-like properties such as specific substrate UDP-GlcNAc binding could be easily obtained. These results are encouraging for subsequent heterologous expression of full-length CHS3a.</p

Saccharomyces cerevisiae chitin biosynthesis activation by N-acetylchitooses depends on size and structure of chito-oligosaccharides

<p>Abstract</p> <p>Background</p> <p>To explore chitin synthesis initiation, the effect of addition of exogenous oligosaccharides on <it>in vitro </it>chitin synthesis was studied. Oligosaccharides of various natures and lengths were added to a chitin synthase assay performed on a <it>Saccharomyces cerevisiae </it>membrane fraction.</p> <p>Findings</p> <p><it>N</it>-acetylchito-tetra, -penta and -octaoses resulted in 11 to 25% [¹⁴C]-GlcNAc incorporation into [¹⁴C]-chitin, corresponding to an increase in the initial velocity. The activation appeared specific to <it>N</it>-acetylchitooses as it was not observed with oligosaccharides in other series, such as beta-(1,4), beta-(1,3) or alpha-(1,6) glucooligosaccharides.</p> <p>Conclusions</p> <p>The effect induced by the <it>N</it>-acetylchitooses was a saturable phenomenon and did not interfere with free GlcNAc and trypsin which are two known activators of yeast chitin synthase activity <it>in vitro</it>. The magnitude of the activation was dependent on both oligosaccharide concentration and oligosaccharide size.</p

Les beta-cetoamides: synthese et reactivite. Application a la synthese de squelettes alcaloidiques

SIGLEAvailable from INIST (FR), Document Supply Service, under shelf-number : TD 80443 / INIST-CNRS - Institut de l'Information Scientifique et TechniqueFRFranc

Oxidation of Δ4- and Δ5-Steroids with Hydrogen Peroxide Catalyzed by Porphyrin Complexes of MnIIIand FeIII

In this paper we describe a new environmentally friendly method to promote the stereoselective epoxidation of Delta4- and Delta5-steroids. Metalloporphyrins efficiently catalyze the epoxidation reactions of 17beta-acetoxy-4-androstene (1), 4-cholestene (2) and 3beta-acetoxy-5-cholestene (3) in the presence of H2O2 as oxygen donor. Modeling the molecular structure of the porphyrin as well as the central metal allows the control of the preferential formation of alpha- or beta-epoxides. Porphyrins with bulky, electron-withdrawing groups in the ortho positions of the meso phenyls and with MnIII as the central metal ion, such as [Mn(TDCPP)Cl], gave preferentially the beta-epoxide of Delta4- and Delta5-steroids. [Fe(TPFPP)Cl] catalyzes preferentially the alpha-epoxidation of Delta4-steroids and also increases the stereoselectivity for the alpha-epoxide in Delta5-steroids, similar to the results obtained with m-CPBA (m-chloroperbenzoic acid) as oxidant. The substrate structure strongly influences the chemoselectivity of the reactions. The X-ray structures of two main products were determined, and two-dimensional NMR techniques allowed the full assignment of 1H and 13C NMR resonances as well as the stereochemistry of these products. A mechanistic proposal involving oxo species for the beta-approach and peroxy species for the alpha-approach is proposed. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004