Preventing bisphosphonate induced osteonecrosis of the jaw with a polyguanidine conjugate (GuaDex): A promising new approach.

Osteonecrosis of the jaw (ONJ) is a relatively rare side effect after prolonged use of bisphosphonates, which are drugs used to treat bone resorption in osteoporosis and certain cancers. This study introduces a novel ONJ model in rats by combining exposure to bisphosphonates, oral surgery, and bacterial inoculation. Potential ONJ preventive effects of polyguanidine (GuaDex) or antibiotics were evaluated. The study consisted of twenty-four male Wistar rats were divided into four groups. Groups 1 to 3 were given weekly doses of i.v. Zoledronic acid (ZA), four weeks before and two weeks after an osteotomy procedure on their left mandibular first molar. Group 4 was a negative control. Streptococcus gordonii bacteria were introduced into the osteotomy pulp chamber and via the food for seven days. On day eight, the rats were given different treatments. Group 1 was given a GuaDex injection into the osteotomy socket, Group 2 was given an intramuscular (i.m.) injection of clindamycin, Group 3 (positive control) was given an i.m. injection of saline, and Group 4 was given an i.m. injection of saline. Blood samples were taken two weeks after the osteotomy procedure, after which the rats were euthanized. Bone healing, bone mineral density, histology, and blood status were analyzed. The results showed that Group 1 (GuaDex) had no ONJ, extensive ongoing bone regeneration, active healing activity, vascularization, and no presence of bacteria. Group 2 (clindamycin) showed early stages of ONJ, avascular areas, and bacteria. Group 3 showed stages of ONJ, inflammatory infiltrates, defective healing, and bacterial presence, and Group 4 had normal healing activity and no bacterial presence. Conclusion: ZA treatment and bacterial inoculation after tooth extraction inhibited bone remodeling/healing and induced ONJ characteristic lesions in the rats. Only GuaDex apparently prevented ONJ development, stimulated bone remodeling, and provided an antimicrobial effect.

Frequency wavelength multiplexed optoacoustic tomography.

Optoacoustics (OA) is overwhelmingly implemented in the Time Domain (TD) to achieve high signal-to-noise ratios by maximizing the excitation light energy transient. Implementations in the Frequency Domain (FD) have been proposed, but suffer from low signal-to-noise ratios and have not offered competitive advantages over time domain methods to reach high dissemination. It is therefore commonly believed that TD is the optimal way to perform optoacoustics. Here we introduce an optoacoustic concept based on pulse train illumination and frequency domain multiplexing and theoretically demonstrate the superior merits of the approach compared to the time domain. Then, using recent advances in laser diode illumination, we launch Frequency Wavelength Multiplexing Optoacoustic Tomography (FWMOT), at multiple wavelengths, and experimentally showcase how FWMOT optimizes the signal-to-noise ratios of spectral measurements over time-domain methods in phantoms and in vivo. We further find that FWMOT offers the fastest multi-spectral operation ever demonstrated in optoacoustics.

360º optoacoustic capsule endoscopy at 50 Hz for esophageal imaging.

Gastrointestinal (GI) endoscopy is a common medical diagnostic procedure used for esophageal cancer detection. Current emerging capsule optoacoustic endoscopes, however, suffer from low pulse repetition rates and slow scanning units limit attainable imaging frame rates. Consequently, motion artifacts result in inaccurate spatial mapping and misinterpretation of data. To overcome these limitations, we report a 360º, 50 Hz frame rate, distal scanning capsule optoacoustic endoscope. The translational capability of the instrument for human GI tract imaging was characterized with an Archimedean spiral phantom consisting of twelve 100 µm sutures, a stainless steel mesh with a pitch of 3 mm and an ex vivo pig esophagus sample. We estimated an imaging penetration depth of ~0.84 mm in vivo by immersing the mesh phantom in intralipid solution to simulate light scattering in human esophageal tissue and validated our findings ex vivo using pig esophagus. This proof-of-concept study demonstrates the translational potential of the proposed video-rate endoscope for human GI tract imaging.

Genetically encoded photo-switchable molecular sensors for optoacoustic and super-resolution imaging.

Reversibly photo-switchable proteins are essential for many super-resolution fluorescence microscopic and optoacoustic imaging methods. However, they have yet to be used as sensors that measure the distribution of specific analytes at the nanoscale or in the tissues of live animals. Here we constructed the prototype of a photo-switchable Ca2+ sensor based on GCaMP5G that can be switched with 405/488-nm light and describe its molecular mechanisms at the structural level, including the importance of the interaction of the core barrel structure of the fluorescent protein with the Ca2+ receptor moiety. We demonstrate super-resolution imaging of Ca2+ concentration in cultured cells and optoacoustic Ca2+ imaging in implanted tumor cells in mice under controlled Ca2+ conditions. Finally, we show the generalizability of the concept by constructing examples of photo-switching maltose and dopamine sensors based on periplasmatic binding protein and G-protein-coupled receptor-based sensors.

Speckle reduction in ultrasound endoscopy using refraction based elevational angular compounding.

Endoscopic ultrasonography (EUS) is a safe, real-time diagnostic and therapeutic tool. Speckle noise, inherent to ultrasonography, degrades the diagnostic precision of EUS. Elevational angular compounding (EAC) can provide real-time speckle noise reduction; however, EAC has never been applied to EUS because current implementations require costly and bulky arrays and are incompatible with the tight spatial constraints of hollow organs. Here we develop a radial implementation of a refraction-based elevational angular compounding technique (REACT) for EUS and demonstrate for the first time spatial compounding in a radial endoscopy. The proposed implementation was investigated in cylindrical phantoms and demonstrated superior suppression of ultrasound speckle noise and up to a two-fold improvement in signal- and contrast- ratios, compared to standard image processing techniques and averaging. The effect of elevational angular deflection on image fidelity was further investigated in a phantom with lymph node-like structures to determine the optimum elevational angular width for high speckle reduction efficiency while maintaining image fidelity. This study introduces REACT as a potential compact and low-cost solution to impart current radial echo-endoscopes with spatial compounding, which could enable accurate identification and precise sizing of lymph nodes in staging of gastrointestinal tract cancers.

In Vivo Three-Dimensional Raster Scan Optoacoustic Mesoscopy Using Frequency Domain Inversion.

Optoacoustic signals are typically reconstructed into images using inversion algorithms applied in the time-domain. However, time-domain reconstructions can be computationally intensive and therefore slow when large amounts of raw data are collected from an optoacoustic scan. Here we considered a fast weighted ω-k (FWOK) algorithm operating in the frequency domain to accelerate the inversion in raster-scan optoacoustic mesoscopy (RSOM), while seamlessly incorporating impulse response correction with minimum computational burden. We investigated the FWOK performance with RSOM measurements from phantoms and mice in vivo and obtained 360-fold speed improvement over inversions based on the back-projection algorithm in the time-domain. This previously unexplored inversion of in vivo optoacoustic data with impulse response correction in frequency domain reconstructions points to a promising strategy of accelerating optoacoustic imaging computations, toward video-rate tomography.

Interferometric optical fiber sensor for optoacoustic endomicroscopy.

Optical fiber sensors can offer robust and miniaturized detection of wideband ultrasound, yielding high sensitivity and immunity to electromagnetic interference. However, the lack of cost-effective manufacturing methods prevents the disseminated use of these sensors in biomedical applications. In this study, we developed and optimized a simple method to create optical cavities with high-quality mirrors for acoustic sensing based on micro-manipulation of UV-curable optical adhesives and electroless chemical silver deposition. This approach enables the manufacturing of ultrasound sensors based on Fabry-Pérot interferometers on optical fiber tips with minimal production costs. Characterization and high-resolution optoacoustic imaging experiments show that the manufacturing process yielded a fiber sensor with a small NEP ( 11mPa/Hz ) over a broad detection bandwidth (25 MHz), generally outperforming conventional piezoelectric based transducers. We discuss how the new manufacturing process leads to a high-performance acoustic detector that, due to low cost, can be used as a disposable sensor.

Ultra-broadband axicon transducer for optoacoustic endoscopy.

Image performance in optoacoustic endoscopy depends markedly on the design of the transducer employed. Ideally, high-resolution performance is required over an expanded depth of focus. Current optoacoustic focused transducers achieve lateral resolutions in the range of tens of microns in the mesoscopic regime, but their depth of focus is limited to hundreds of microns by the nature of their spherical geometry. We designed an ultra-broadband axicon detector with a 2 mm central aperture and investigated whether the imaging characteristics exceeded those of a spherical detector of similar size. We show a previously undocumented ability to achieve a broadband elongated pencil-beam optoacoustic sensitivity with an axicon detection geometry, providing approximately 40 µm-lateral resolution maintained over a depth of focus of 950 µm-3.8 times that of the reference spherical detector. This performance could potentially lead to optoacoustic endoscopes that can visualize optical absorption deeper and with higher resolution than any other optical endoscope today.

Improving ultrasound images with elevational angular compounding based on acoustic refraction.

Ultrasound imaging is affected by coherent noise or speckle, which reduces contrast and overall image quality and degrades the diagnostic precision of the collected images. Elevational angular compounding (EAC) is an attractive means of addressing this limitation, since it reduces speckle noise while operating in real-time. However, current EAC implementations rely on mechanically rotating a one-dimensional (1D) transducer array or electronically beam steering of two-dimensional (2D) arrays to provide different elevational imaging angles, which increases the size and cost of the systems. Here we present a novel EAC implementation based on a 1D array, which does not necessitate mechanically rotating the transducer. The proposed refraction-based elevational angular compounding technique (REACT) instead utilizes a translating cylindrical acoustic lens that steers the ultrasound beam along the elevational direction. Applying REACT to investigate phantoms and excised tissue samples demonstrated superior suppression of ultrasound speckle noise compared to previous EAC methods, with up to a two-fold improvement in signal- and contrast-to-noise ratios. The effects of elevational angular width on speckle reduction was further investigated to determine the appropriate conditions for applying EAC. This study introduces acoustic refractive elements as potential low cost solutions to noise reduction, which could be integrated into current medical ultrasound devices.

Assessment of hessian-based Frangi vesselness filter in optoacoustic imaging.

The Hessian-based Frangi vesselness filter is commonly used to enhance vasculature in optoacoustic (photoacoustic) images, but its accuracy and limitations have never been rigorously assessed. Here we validate the ability of the filter to enhance vessel-like structures in phantoms, and we introduce an experimental approach that uses measurements before and after the administration of gold nanorods (AuNRs) to examine filter performance in vivo. We evaluate the influence of contrast, filter scales, angular tomographic coverage, out-of-plane signals and light fluence on image quality, and gain insight into the performance of the filter. We observe the generation of artifactual structures that can be misinterpreted as vessels and provide recommendations to ensure appropriate use of Frangi and other vesselness filters and avoid misinterpretation of post-processed optoacoustic images.

Homogentisic acid-derived pigment as a biocompatible label for optoacoustic imaging of macrophages.

Macrophages are one of the most functionally-diverse cell types with roles in innate immunity, homeostasis and disease making them attractive targets for diagnostics and therapy. Photo- or optoacoustics could provide non-invasive, deep tissue imaging with high resolution and allow to visualize the spatiotemporal distribution of macrophages in vivo. However, present macrophage labels focus on synthetic nanomaterials, frequently limiting their ability to combine both host cell viability and functionality with strong signal generation. Here, we present a homogentisic acid-derived pigment (HDP) for biocompatible intracellular labeling of macrophages with strong optoacoustic contrast efficient enough to resolve single cells against a strong blood background. We study pigment formation during macrophage differentiation and activation, and utilize this labeling method to track migration of pro-inflammatory macrophages in vivo with whole-body imaging. We expand the sparse palette of macrophage labels for in vivo optoacoustic imaging and facilitate research on macrophage functionality and behavior.

Capsule optoacoustic endoscopy for esophageal imaging.

Detection and monitoring of esophageal cancer severity require an imaging technique sensitive enough to detect early pathological changes in the esophagus and capable of analyzing the esophagus over 360 °in a non-invasive manner. Optoacoustic endoscopy (COE) has been shown to resolve superficial vascular structure of the esophageal lumen in rats and rabbits using catheter-type probes. Although these systems can work well in small animals, they are unsuitable for larger lumens with thicker walls as required for human esophageal screening, due to their lack of position stability along the full organ circumference, sub-optimal acoustic coupling and limited signal-to-noise ratio (SNR). In this work, we introduce a novel capsule COE system that provides high-quality 360° images of the entire lumen, specifically designed for typical dimensions of human esophagus. The pill-shaped encapsulated probe consists of a novel and highly sensitive ultrasound transducer fitted with an integrated miniature pre-amplifier, which increases SNR of 10 dB by minimizing artifacts during signal transmission compared to the configuration without the preamplifier. The scanner rotates helically around the central axis of the probe to capture three-dimensional images with uniform quality. We demonstrate for the first time ex vivo volumetric vascular network images to a depth of 2 mm in swine esophageal lining using COE. Vascular information can be resolved within the mucosa and submucosa layers as confirmed by histology of samples stained with hematoxylin and eosin and with antibody against vascular marker CD31. COE creates new opportunities for optoacoustic screening of esophageal cancer in humans.

Assessing surface characteristics of eroded dentine with optical coherence tomography: a preliminary in vitro validation study.

We conducted the first pilot study to investigate the use of the attenuation coefficient from an optical coherence tomography (OCT) backscattered signal as a measure of surface roughness changes in eroded dentine at an early stage of the erosion process. Ten human premolar root samples were subjected to citric acid treatment before scanning by OCT. The extracted relative attenuation coefficient (µR) from backscattered OCT signals was shown to increase with the duration of acid challenge. Validated against roughness measurements (rSa) from scanning electron microscopy scans, µR is significantly correlated with rSa indicative of severity of erosion (p<0.01, r=0.9195). We conclude that the OCT attenuation coefficient of the immediate subsurface in eroded dentine is a potential surrogate measure for its surface roughness. However, further work should be performed to study how it relates to the surface and immediate subsurface changes effected by other mechanical wear before it could unequivocally be used as a surrogate measurement for surface roughness.

Endogenous TNFα orchestrates the trafficking of neutrophils into and within lymphatic vessels during acute inflammation.

Neutrophils are recognised to play a pivotal role at the interface between innate and acquired immunities following their recruitment to inflamed tissues and lymphoid organs. While neutrophil trafficking through blood vessels has been extensively studied, the molecular mechanisms regulating their migration into the lymphatic system are still poorly understood. Here, we have analysed neutrophil-lymphatic vessel interactions in real time and in vivo using intravital confocal microscopy applied to inflamed cremaster muscles. We show that antigen sensitisation of the tissues induces a rapid but transient entry of tissue-infiltrated neutrophils into lymphatic vessels and subsequent crawling along the luminal side of the lymphatic endothelium. Interestingly, using mice deficient in both TNF receptors p55 and p75, chimeric animals and anti-TNFα antibody blockade we demonstrate that tissue-release of TNFα governs both neutrophil migration through the lymphatic endothelium and luminal crawling. Mechanistically, we show that TNFα primes directly the neutrophils to enter the lymphatic vessels in a strictly CCR7-dependent manner; and induces ICAM-1 up-regulation on lymphatic vessels, allowing neutrophils to crawl along the lumen of the lymphatic endothelium in an ICAM-1/MAC-1-dependent manner. Collectively, our findings demonstrate a new role for TNFα as a key regulator of neutrophil trafficking into and within lymphatic system in vivo.

Histological validation of near-infrared reflectance multispectral imaging technique for caries detection and quantification.

Near infrared (NIR) multispectral imaging is a novel noninvasive technique that maps and quantifies dental caries. The technique has the ability to reduce the confounding effect of stain present on teeth. The aim of this study was to develop and validate a quantitative NIR multispectral imaging system for caries detection and assessment against a histological reference standard. The proposed technique is based on spectral imaging at specific wavelengths in the range from 1000 to 1700 nm. A total of 112 extracted teeth (molars and premolars) were used and images of occlusal surfaces at different wavelengths were acquired. Three spectral reflectance images were combined to generate a quantitative lesion map of the tooth. The maximum value of the map at the corresponding histological section was used as the NIR caries score. The NIR caries score significantly correlated with the histological reference standard (Spearman's Coefficient=0.774, p<0.01). Caries detection sensitivities and specificities of 72% and 91% for sound areas, 36% and 79% for lesions on the enamel, and 82% and 69% for lesions in dentin were found. These results suggest that NIR spectral imaging is a novel and promising method for the detection, quantification, and mapping of dental caries.

Quantitative light fluorescence (QLF) and polarized white light (PWL) assessments of dental fluorosis in an epidemiological setting.

BACKGROUND: To determine if a novel dual camera imaging system employing both polarized white light (PWL) and quantitative light induced fluorescence imaging (QLF) is appropriate for measuring enamel fluorosis in an epidemiological setting. The use of remote and objective scoring systems is of importance in fluorosis assessments due to the potential risk of examiner bias using clinical methods. METHODS: Subjects were recruited from a panel previously characterized for fluorosis and caries to ensure a range of fluorosis presentation. A total of 164 children, aged 11 years (±1.3) participated following consent. Each child was examined using the novel imaging system, a traditional digital SLR camera, and clinically using the Dean's and Thylstrup and Fejerskov (TF) Indices on the upper central and lateral incisors. Polarized white light and SLR images were scored for both Dean's and TF indices by raters and fluorescence images were automatically scored using software. RESULTS: Data from 164 children were available with a good distribution of fluorosis severity. The automated software analysis of QLF images demonstrated significant correlations with the clinical examinations for both Dean's and TF index. Agreement (measured by weighted Kappa's) between examiners scoring clinically, from polarized photographs and from SLR images ranged from 0.56 to 0.92. CONCLUSIONS: The study suggests that the use of a digital imaging system to capture images for either automated software analysis, or remote assessment by raters is suitable for epidemiological work. The use of recorded images enables study archiving, assessment by multiple examiners, remote assessment and objectivity due to the blinding of subject status.

Near-infrared hyperspectral imaging of teeth for dental caries detection.

Near-infrared (NIR) is preferred for caries detection compared to visible light imaging because it exhibits low absorption by stain and deeper penetration into teeth. Hyperspectral images from 1000 to 2500 nm have been obtained for a total of 12 extracted teeth (premolars and molars) with different degrees of natural lesion. Analysis of the reflectance spectra suggests that light scattering by porous enamel and absorption by water in dentin can be used to quantify the lesion severity and generate a NIR caries score. Teeth were ground for histological examination after the measurements. The NIR caries score obtained correlates significantly (Spearman's correlation of 0.89, p<0.01) with the corresponding histological score. Results yield a sensitivity of >99% and a specificity of 87.5% for enamel lesions and a sensitivity of 80% and a specificity >99% for dentine lesions. The nature of the technique offers significant advantages, including the ability to map the lesion distribution rather than obtaining single-point measurements, it is also noninvasive, noncontact, and stain insensitive. These results suggest that NIR spectral imaging is a potential clinical technique for quantitative caries diagnosis and can determine the presence of occlusal enamel and dentin lesions.

In vivo quantification of gingival inflammation using spectral imaging.

Erythema is a reaction of the skin and oral soft tissues commonly associated with inflammation and an increase in blood flow. Diffuse reflection spectroscopy is a powerful tool for the assessment of skin inflammation where erythema has been linked to the relative concentration of oxygenated hemoglobin and blood perfusion. Here we demonstrate the applicability of a spectral imaging method for the quantification of gingival inflammation by looking at the gingival margin and papillary tip erythema. We present a longitudinal study on 22 healthy volunteers divided in two groups. The first was allowed to have normal oral hygiene and the second was subjected to an induced gingivitis for two weeks by cessation of oral hygiene. The spectral reflectance ratio at 615 and 460 nm, R(615)R(460), was proposed as a method to quantify and map the erythema spatial distribution. These wavelengths represent spectral absorption crossovers observed between oxygenated and deoxygenated hemoglobin. The spectral method presented shows a significant separation (p<0.01) between the groups when gingivitis was induced and correlates significantly (p<0.05) with the clinical gingival index scores. We believe that these investigations could contribute to the development of functional imaging methods for periodontal disease detection and monitoring.

Laser Doppler imaging through tissues phantoms by using self-mixing interferometry with a laser diode.

Laser Doppler imaging has been widely used for the evaluation of cutaneous blood flow. We report on how the self-mixing interferometry configuration with a laser diode is explored for what is believed to be the first time to generate flow maps. The experiment was carried out by sensing the laser intensity power spectrum at each pixel as the laser was scanned over a model that mimics the properties of skin and circulating blood.

Dynamic light scattering by using self-mixing interferometry with a laser diode.

The power spectrum of the laser intensity is studied when optical feedback in a laser diode is used as a sensing configuration for dynamic light-scattering experiments. We present a theory that relates the power spectrum obtained from standard dynamic light-scattering theory to the intensity power spectrum of the laser. This theory provides a concise description of this sensing technique, also known as self-mixing interferometry, when it is applied to Doppler shift and line-broadening measurements of the backscattered field.