Hervé Tassery

Functional mapping of human sound and carious enamel and dentin with Raman spectroscopy.

The goals of this trial were, first, to produce a Raman mapping of decay and sound dentin samples, through accurate analysis of the Raman band spectra variations of mineral and organic components. The second goal was to confirm the correlation between the Raman signal and the signal of a fluorescent camera, by assaying the concentration of pentosidine and natural collagen fluorescent crosslink using reverse phase high-pressure liquid chromatography. The first correlation assumed a possible relationship between the signal observed with the camera and Raman spectroscopy. The second correlation assumed an association with the Maillard reaction. Absence of a correlation for this trial was that no association could be found between Raman spectra characteristics, fluorescence variation and the HPLC assay. Our results void this absence.

In vitro investigation of fluorescence of carious dentin observed with a Soprolife® camera

ObjectivesOur aim was to determine the origin of the red fluorescence of carious dentine observed with the Soprolife® camera.MethodsWe conducted in vitro studies to evaluate the origin of the red fluorescence using acids and matrix metalloproteinase (MMP) to mimic caries and methylglycoxal (MGO) to evaluate the effect of glycation reactions on the red fluorescence. In every step of these models, we detected the changes of dentin photonic response with Soprolife® in daylight mode and in treatment mode. A Raman spectroscopy analysis was performed to determine the variations of the dentin organic during the in vitro caries processes. Raman microscopy was performed to identify change in the collagen matrix of dentine.ResultsThe red fluorescence observed in carious dentine using a Soprolife® camera corresponds to the brownish color observed using daylight. Demineralization using nitric acid induces a loss of the green fluorescence of dentine. The red fluorescence of carious dentine is resistant to acid treatment. Immersion of demineralized dentine in MGO induces a change of color from white to orange-red. This indicates that the Maillard reaction contributes to lesion coloration. Immersion of demineralized dentine in an MMP-1 solution followed by MGO treatment results in a similar red fluorescence. Raman microspectroscopy analysis reveals accumulation of AGEs product in red-colored dentine.ConclusionsOur results provide important information on the origin of the fluorescence variation of dentine observed with the Soprolife® camera. We demonstrate that the red fluorescence of carious dentine is linked to the accumulation of Advanced Glycation End products (AGE).Clinical relevanceThe study provides a new biological basis for the red fluorescence of carious dentine and reinforces the importance of the Soprolife® camera in caries diagnostics.

Wetting properties and critical micellar concentration of benzalkonium chloride mixed in sodium hypochlorite.

INTRODUCTION The purposes of the present study were to (1) assess the effect of the addition of benzalkonium chloride to sodium hypochlorite on its wetting properties, contact angle, and surface energy; (2) determine the critical micellar concentration of benzalkonium chloride in sodium hypochlorite; and (3) investigate the influence of addition of benzalkonium chloride on the free chlorine level, cytotoxicity, and antiseptic properties of the mixture. METHODS Solutions of benzalkonium chloride, with concentrations ranging from 0%-1%, were mixed in 2.4% sodium hypochlorite and tested as follows. The wetting properties were investigated by measuring the contact angle of the solutions on a nondehydrated dentin surface by using the static sessile drop method. The pending drop technique was subsequently used to determine the surface energy of the solutions. The critical micellar concentration of benzalkonium chloride mixed in sodium hypochlorite was calculated from the data. When 2.4% NaOCl was mixed with benzalkonium chloride at the critical micellar concentration, 3 parameters were tested: free chloride content, cytotoxicity, and antibacterial effects against Enterococcus faecalis. RESULTS The contact angle (P < .001) as well as the surface energy (P < .001) significantly decreased with increasing benzalkonium chloride concentrations. The critical micellar concentration of benzalkonium chloride in sodium hypochlorite was 0.008%. At this concentration, the addition of benzalkonium chloride had no effect on the free chlorine content, cytotoxicity, or antibacterial efficiency of the mixture. CONCLUSIONS The addition of benzalkonium chloride to sodium hypochlorite at the critical micellar concentration reduced the contact angle by 51.2% and the surface energy by 53.4%, without affecting the free chloride content, cytotoxicity, or antibacterial properties of the mixture.

Multiphoton imaging of the dentine-enamel junction.

Multiphoton microscopy has been used to reveal structural details of dentine and enamel at the dentin-enamel junction (DEJ) based on their 2-photon excited fluorescence (2PEF) emission and second harmonic generation (SHG). In dentine tubule 2PEF intensity varies due to protein content variation. Intertubular dentin produces both SHG and 2PEF signals. Tubules are surrounded by a thin circular zone with a lower SHG signal than the bulk dentine and the presence of collagen fibers perpendicular to the tubule longitudinal axis is indicated by strong SHG responses. The DEJ appears as a low intensity line on the 2PEF images and this was never previously reported. The SHG signal is completely absent for enamel and aprismatic enamel shows a homogeneous low 2PEF signal contrary to prismatic enamel. The SHG intensity of mantle dentine is increasing from the dentine-enamel junction in the first 12 μm indicating a progressive presence of fibrillar collagen and corresponding to the more external part of mantle dentine where matrix metallo-proteases accumulate. The high information content of multiphoton images confirms the huge potential of this method to investigate tooth structures in physiological and pathological conditions.

Molecular structural analysis of carious lesions using micro‐Raman spectroscopy

In clinical situations carious dentine tissues can be discriminated by most caries fluorescence detection tools, including a new fluorescence intra-oral camera. The objectives of this study were: (i) to analyze the Raman spectra of sound, carious, and demineralized dentine, (ii) to compare this spectral analysis with the fluorescence variation observed when using a fluorescence camera, and (iii) to evaluate the involvement of the Maillard reaction in the fluorescence variations. The first positive hypothesis tested was that the fluorescence of carious dentine obtained using a fluorescence camera and the Raman spectra variation were closely related. The second was that the variation of fluorescence could be linked with the Maillard reaction. Sound dentine, sound dentine demineralized in aqueous nitric acid solution, carious soft dentine, sound dentine demineralized in lactic acid solution, sound dentine demineralized in aqueous nitric acid solution and immersed in methylglycoxal solution, and sound dentine demineralized in aqueous nitric acid solution and immersed in methylglycoxal and glucose solutions, were studied using micro-Raman spectroscopy. Modifications in the band ratio of amide, phosphate, and carbonate were observed in the decayed and demineralized groups compared with the sound dentine group. The results indicate that a close relationship exists between the Maillard reaction and fluorescence variation.

Laboratory Studies of Nonlinear Optical Signals for Caries Detection

Multiphoton confocal microscopy and nonlinear spectroscopy are used to investigate the caries process in dentin. Although dentin is a major calcified tissue of the teeth, its organic phase comprises type I collagen fibers. Caries drive dentin demineralization and collagen denaturation. Multiphoton microscopy is a powerful imaging technique: the biological materials are transparent to infrared frequencies and can be excited to penetration depths inaccessible to 1-photon confocal microscopy. The laser excitation greatly reduces photodamage to the sole focal region, and the signal-to-noise ratio is improved significantly. The method has been used to follow pathologic processes involving collagen fibrosis or collagen destruction based on their 2-photon excited fluorescence (2PEF) emission and second harmonic generation (SHG). Combining multiphoton imaging with nonlinear spectroscopy, we demonstrate that both 2PEF and SHG intensity of human dentin are strongly modified during the tooth caries process, and we show that the ratio between SHG and 2PEF signals is a reliable parameter to follow dental caries. The ratio of the SHG/2PEF signals measured by nonlinear optical spectroscopy provides valuable information on the caries process, specifically on the degradation of the organic matrix of dentin. The goal is to bring these nonlinear optical signals to clinical application for caries diagnosis.

Multiphoton Microscopy for Caries Detection with ICDAS Classification

Dentin carious lesion is a dynamic process that involves demineralization and collagen denaturation. Collagen type I is the major protein in dentin and it has been investigated based on its optical properties. Multiphoton microscopy (MPM) is a nonlinear imaging technique that reveals the caries process using the collagen two-photon excitation fluorescence (2PEF) and its second-harmonic generation (SHG). Combining the histological and the International Caries Detection and Assessment System (ICDAS) classifications with nonlinear optical spectroscopy (NLOS), 2PEF and SHG intensities of enamel and dentin were highly altered during the caries process. It has been proven that the ratio SHG/2PEF is a relevant indicator of the organic matrix denaturation [Terrer et al.: J Dent Res 2016; 96: 574–579]. In the present study, a series of measurable signals is made to detect early stages of carious lesion according to the ICDAS classification and to explore the relationship between these measures and the ICDAS scale. Comparison of the efficiency of nonlinear optical signals for caries detection with the ICDAS classification is essential to evaluate their potential for clinical application. In our study, the use of the NLOS measured by MPM allowed us to monitor a quantitative parameter (SHG/2PEF ratio) according to the dentin carious lesion state (ICDAS and histological examination). Three coherent new groups were defined (ICDAS 0/1; ICDAS 2/3; ICDAS 4/5/6), where the carious process can be clearly described with a statistically significant decrease of the SHG/2PEF ratio.

Method to evaluate the noise of 3D intra-oral scanner

In dentistry, 3D intra-oral scanners are gaining increasing popularity essentially for the production of dental prostheses. However, there is no normalized procedure to evaluate their basic performance and enable comparisons among intra-oral scanners. The noise value highlights the trueness of a 3D intra-oral scanner and its capacity to plan prosthesis with efficient clinical precision. The aim of the present study is to develop a reproducible methodology for determining the noise of an intra-oral scanner. To this aim, and as a reference, an ultra-flat and ultra-smooth alumina wafer is used as a blank test. The roughness is calculated using an AFM (atomic force microscope) and interferometric microscope measurements to validate this ultra-flat characteristic. Then, two intra-oral scanners (Carestream CS3500 and Trios 3Shape) are used. The wafer is imaged by the two intra-oral scanners with three different angles and two different directions, 10 times for each parameter, given a total of 50 3D-meshes per intra-oral scanner. RMS (root mean square), representing the noise, is evaluated and compared for each angle/direction and each intra-oral scanner, for the whole mesh, and then in a central ROI (region of interest). In this study, we obtained RMS values ranging between 5.29 and 12.58 micrometers. No statistically significant differences were found between the mean RMS of the two intra-oral scanners, but significant differences in angulation and orientations were found between different 3D intra-oral scanners. This study shows that the evaluation of RMS can be an indicator of the value of the noise, which can be easily assessed by applying the present methodology.

Detection and Diagnosis of Carious Lesions

The early detection and diagnosis of the carious lesion are a primary consideration of the minimal intervention dentistry (MID) concept. Detection is the identification of a demineralized lesion, while diagnosis is an iterative process using further information from the patient to identify the lesion as carious in nature. Traditionally, visual, tactile, and radiographic methods are used to detect carious lesion; however, recently tactile detection, especially using a sharp explorer, is not recommended apart from the delicate detection of enamel surface integrity/roughness. Use of simple devices such as loupes for magnification of the clean surface can improve detection validity. Advances in technology have led to many different devices being released onto the market to assist in the detection and quantification of carious lesions. Fluorescence of the tooth structure using different wavelengths of light can provide information about lesion extent and area and whether dentine is involved, and several commercial systems are available such as DIAGNOdent®, QLF®, and Soprocare/Soprolife®. It is important to follow standardized procedures with all of these detection techniques, as the validity and reproducibility of the results are highly dependent on this. Electrical impedance and photothermal radiometry have been promoted as alternatives to light-based technologies; however, evidence for these devices is still somewhat limited. All of these traditional and new detection methods will be discussed in the context of minimal intervention dentistry and their potential value to the clinician and patient.