| Popis: |
A literature review of dental plaque biofilms formation, progression and detachment mechanisms is presented in this thesis. Various strategies that have been employed to reduce or eliminate dental biofilms are discussed. The focus of the thesis was on the mechanical properties and disruption of dental biofilms, especially from hard-to-access areas of the oral cavity, such as the interproximal (IP) sites between the teeth. Various methods to measure mechanical properties of dental biofilms were investigated, and physical and chemical strategies to disrupt these biofilms were employed. Streptococcus mutans, the bacterium responsible for initiation of dental plaque biofilms, was used in our studies. A uniaxial compressive test was utilized to characterize the mechanical behaviour of biofilms, while manipulating the chemical microenvironment. Initially, the mechanical properties of a dextran gel were characterized. The gel was used as an artificial dental plaque biofilm (Chapter 5). The elastic modulus of the gel was 17 kPa (± 12; n = 3), and the stress relaxation time was 25 seconds (± 18; n = 3), demonstrating a viscoelastic behaviour similar to that reported for real biofilms. After optimizing the technique with the gel, the mechanical properties of S. mutans biofilms were studied, the elastic modulus was 380 Pa (± 350; n = 30), and the stress relaxation time was 12 seconds (± 11; n = 10). The elastic modulus increased by increasing the sucrose percentage in the media, and decreased when the biofilms were treated with increasing concentrations of ethylene di-amine tetra acetic acid, EDTA. Treating the biofilms with different solutions of poly (ethylene glycol), PEG, resulted in behaviour similar to that previously observed for synthetic polymers. The flow field and local hydrodynamics of high velocity water microdrops impacting the interproximal (IP) space of typodont teeth, and their influence on the structure and detachment of both surrogate dental plaque and Streptococcus mutans biofilms, were studied experimentally and computationally. Water droplets of 115 μL were produced by a prototype AirFloss (PT-AirFloss) device provided by Philips Oral Healthcare, bursting water at a velocity of 60 m/s into the IP space between the maxillary central incisors. High-speed imaging, was used to characterise the PT-AirFloss microburst of pressurized air and water micodrops, and demonstrated the removal mechanism of a dental plaque biofilm substitute and the S. mutans biofilms. Using various microscopy and image analysis techniques, quantitative measurements of the removal rate and the percentage removal of biofilms from different locations in the IP space were obtained. Microcomputed Tomography (μ-CT) imaging was used to obtain 3D images of the typodont and the IP spaces. The shear stress distribution generated by the drop impacting the tooth surface was calculated by Computational Fluid Dynamics (CFD) simulations based on the finite element method (FEM). There was good agreement between experimentally measured biofilm removal and the pattern of predicted wall shear stress (τw) generated in the IP space by the microburst. High velocity water microdrops, with minimal fluid volume and time, effectively removed both the surrogate and the biofilm. The shear stress generated by the PT-AirFloss and its spatial distribution on the teeth surface played a key role in dictating the efficacy of biofilm removal. In addition, CFD models were used to predict optimal water drop or burst design with respect to more effective biofilm removal performance. Furthermore, the influence of fluid shear flow on the detachment of Streptococcus mutans biofilms inside microfluidic channels was studied using a commercially available flow-cell system. A critical biofilm detachment shear stress was estimated for the large biofilm-aggregates (CDSSagg). The CDSSagg value was used in the CFD model to predict the spatial distribution of biofilm aggregates detachment from the IP surface caused by the PT-AirFloss microburst. Next the effect of three biofilm matrix-degrading enzymes on the structure and detachment of Streptococcus mutans biofilms inside microtiter plates and on typodont teeth was studied experimentally. The enzymes used were: Bromelain (a protease), DNase, and RNase. The biofilms were treated with different enzymatic preparations, stained with Live/Dead and Crystal Violet, and the corresponding optical density (OD) and fluorescence intensity (FI) were measured by a microplate reader. The results detailed the degradation effect of each enzyme, separately and in combination. The three enzymes demonstrated different efficacies in degrading the biofilm in 6, 24 and 96 well-plates, as well as on the typodont teeth. Also, there was a large variability which could be explained by the heterogeneity of the biofilm. Using epifluorescence microscopy and image analysis, quantitative measurements of the percentage surface area coverage were obtained, and the preliminary results were consistent with the datafrom the plate reader. Furthermore, pre-coating the plates with the three enzymes did not inhibit biofilm from formation and accumulation. Lastly the use of a biocompatible copolymer of methylvinyl ether and maleic anhydride, with excellent mucosal adhesive properties and biocompatibility to improve enzymatic digestion by offering a prolonged contact of the enzymes with the teeth and oral tissues, was investigated. The rationale was to eliminate a major obstacle facing the efficacy of the enzymatic therapy which was the relatively short residence time of the enzymes at the site of administration. The adhesive copolymer could possibly enhance enzyme biofilm degradation. Combining the adhesive copolymer with the enzymes could potentially allow near total degradation of the laboratory-grown S. mutans biofilms. |
