Peroxidases regulation of Candida albicans oral biofilms
Among the prosthetic and implant biomaterials in the oral cavity currently exciting tremendous interest, titanium and resin are the main components of implants and dentures respectively. From their introduction in the oral cavity, a highly complex heterogeneous biofilm coats these biomaterials. The...
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| Format: | Doctoral Thesis |
| Language: | en |
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Universite Libre de Bruxelles
2012
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| Online Access: | https://dipot.ulb.ac.be/dspace/bitstream/2013/209728/1/23f4aa44-4a94-42cb-b0e7-0a1bf6990a6e.txt https://dipot.ulb.ac.be/dspace/bitstream/2013/209728/4/586e24b9-5ece-484b-a9d7-f2a34b2479d7.txt http://hdl.handle.net/2013/ULB-DIPOT:oai:dipot.ulb.ac.be:2013/209728 |
| Summary: | Among the prosthetic and implant biomaterials in the oral cavity currently exciting tremendous interest, titanium and resin are the main components of implants and dentures respectively. From their introduction in the oral cavity, a highly complex heterogeneous biofilm coats these biomaterials. The present thesis analyzes the complex relationships that are formed between one single microorganism (Candida albicans), one defence system of the oral cavity (oral peroxidases) and the two aforementioned biomaterials (titanium and resin). These biomaterials are indeed in contact with peroxidases: myeloperoxidase from neutrophils and sialoperoxidase from salivary secretions. Oral peroxidases belong to the salivary non immune innate defence mechanisms that control the oral microbial flora. In the presence of hydrogen peroxide (H2O2), they catalyze in vivo the oxidation of thiocyanate (SCN-) into hypothiocyanite (OSCN-), in vitro the oxidation of iodide (I-) into hypoiodite (OI-). In the salivary compartment and in oral biofilms, H2O2 is mainly formed by bacteria. In our investigations, H2O2 was produced by an enzyme sequence glucose (G) / glucose-oxidase (GOD). The OSCN- and OI- are antibacterial, antiviral and antifungal oxidants. Few studies have considered their action on Candida biofilms.<p>Candida albicans is a commensal yeast of the oral cavity which can turn parasitic when the host immune defences are weakened. This fungus forms biofilms on biomaterials within the mouth, especially on dentures, the decontamination of these prostheses is therefore essential to avoid the risk of candidosis. (Candida et prothèses dentaires. Ahariz M. Loeb I. Courtois Ph. Rev. Stomatol. Chir. Maxillofac. 111: 74-78, 2010).<p>In vitro, our investigations aimed to analyze the relationships between peroxidase systems and Candida. The effect of peroxidase systems (G / GOD / KI or KSCN / peroxidase) on Candida suspensions, on biofilms already formed or in formation was evaluated with various inputs of hydrogen peroxide and was studied by incorporation of enzymatic sequences in the culture media used for Candida biofilms formation. The susceptibility of Candida albicans ATCC 10231 to OSCN- versus to OI-, produced by lactoperoxidase (LPO), was studied in three different experimental models: - in a liquid culture medium – on a solid medium (with agarose gel), - in a biofilm model developed in the context of this work. The latter consisted of titanium powder suspended in Sabouraud broth contaminated with Candida albicans. The growth of Candida in the supernatant (planktonic phase) was evaluated by turbidimetry and the biomass of yeasts adherent to biomaterials (attached phase) by the tetrazolium salt MTT method. Enzymatic studies have allowed the optimization of the concentrations and activities of peroxidase systems components and the illustration of the competition between thiocyanate and iodide for lactoperoxidase. Peroxidase systems G/GOD/I-/LPO and G/GOD/SCN-/LPO prevented or limited the growth of Candida in the planktonic and attached phases on titanium powder for at least 21 days. At a dose of GOD (0.2 U / ml), the system G/GOD/I-/LPO has limited the development of planktonic and attached phases for 4 days while the system G/ GOD/SCN-/LPO has shown an inhibitory effect only in the first 2 days of incubation. (Candida albicans susceptibility to lactoperoxidase-generated hypoiodite. Ahariz M. Courtois Ph. Clinical, cosmetic and investigational dentistry; 2: 69-78, 2010).<p>In other experiments, peroxidase was adsorbed on titanium sheets in order to modify their surface and give them the property of inhibiting biofilm formation of Candida after addition of the enzyme substrates. Enzymatic studies and X-ray photoelectron spectroscopy (XPS) showed the adsorption of lactoperoxidase to titanium. In vivo, peroxidases are adsorbed on titanium healing abutments. (Adsorption of peroxidase on titanium surfaces: A pilot study. Ahariz M. Mouhyi J. Louette P. Van Reck J. Malevez C. Courtois P. J. Biomed. Mater. Res. 52: 567-571, 2000).<p>The development of the Candida biofilm was followed on titanium (powder or sheets) as well as on resin. Planktonic and attached phases have been monitored for 21 days. The presence of an exopolysaccharide matrix secreted by the yeasts has been observed with light microscopy and confirmed with fluorescence using the calcofluor method. In a series of experiments on titanium and resin sheets, an attached phase was demonstrated by the same techniques. The efficiency of the peroxidase system using iodide as a substrate was demonstrated when the enzyme was in solution and when it was preadsorbed on titanium (Candida albicans biofilm on titanium: effect of peroxidase precoating. Ahariz M. Courtois Ph. Medical Devices: Evidence and Research; 3: 33-40, 2010).<p>The incorporation in oral gels of other molecules present in exocrine secretions is a research direction that was also discussed: the present studies have demonstrated the in vitro benefits of peroxidase systems (with thiocyanate, chloride or especially iodide as substrates) acting in synergy with colostrum, lactoferrin and lysozyme. But the formulation of specialities that contain these natural antimicrobials is difficult to transpose in vivo as the complexity of the oral environment is very large (Denture contamination by yeasts in the elderly. Vanden Abbeele A. de Meel H. Ahariz M. Perraudin J.-P. Beyer I. Courtois P. Gerodontology; 25: 222-228, 2008).<p>Investigations pursued in vivo in 155 patients allowed the determination of the wild strains of Candida sp present on the fitting surface of the removable dental appliance and on the corresponding palatal mucosa. The link between the presence of yeasts and a reduced salivary flow was confirmed. These wild strains were directly grown and identified on Petri dishes (ChromAgar™ medium) from the macroscopic morphology of colonies and from additional tests (germination test in human serum, formation of chlamydoconidies on RAT medium, API™ galleries identification system ). For a period of two weeks, 14 patients accepted the daily application of a gel on the fitting surface of their denture. It was a double-blind comparison of an active gel containing the thiocyanate - lysozyme - lactoferrin - colostrum complete system with a control gel inactivated by heating. Data analysis showed a reduction in the number of colonies on the palatal mucosa by Candida sp, but not on the denture itself. By contrast, decontamination ex vivo of dentures by immersion in a bath (at room temperature or 37° C) containing either G/GOD producing H2O2 or the complete peroxidase system G/GOD/KI/L producing OI- demonstrated the efficiency of hypoiodite.<p><p><p>Parmi les biomatériaux prothétiques et implantaires qui connaissent actuellement un essor considérable dans la sphère orale, le titane est le composant principal des implants et la résine celui des prothèses dentaires. Dès leur introduction dans la cavité orale, un biofilm hétérogène très complexe les recouvre. Cette thèse analyse la complexité des relations qui se nouent entre un seul micro-organisme (Candida albicans), un système de défense de la cavité orale (les peroxydases orales) et les 2 biomatériaux précités (titane, résine). Ces biomatériaux sont en effet, dans le milieu oral, au contact de peroxydases: la myéloperoxydase des neutrophiles et la sialoperoxydase des sécrétions salivaires. Les peroxydases orales appartiennent aux mécanismes salivaires de défense innée non immunitaires qui contrôlent la flore microbienne orale. En présence de peroxyde d’hydrogène (H2O2), elles catalysent in vivo l’oxydation du thiocyanate (SCN-) en hypothiocyanite (OSCN-) et in vitro l’oxydation d’iodure (I-) en hypoiodite (OI-). Dans le compartiment salivaire et dans les biofilms oraux, l’H2O2 provient essentiellement des bactéries. Dans nos investigations, l’H2O2 était produit par une séquence enzymatique glucose (G) / glucose-oxydase (GOD). L’OSCN- et l’OI- sont des oxydants antibactériens, antiviraux et antifongiques. Peu d’études envisagent leur action sur les biofilms à Candida.<p>\ === Doctorat en Sciences dentaires === info:eu-repo/semantics/nonPublished |
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