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Bone material contains a hierarchical network of micro- and nano-cavities and channels, known as the lacuna-canalicular network (LCN), that is thought to play an important role in mechanobiology and turnover. The LCN comprises micrometer-sized lacunae, voids that house osteocytes, and submicrometer-sized canaliculi that connect bone cells. Characterization of this network in three dimensions is crucial for many bone studies. To quantify X-ray Zernike phase-contrast nanotomography data, deep learning is used to isolate and assess porosity in artifact-laden tomographies of zebrafish bones. A technical solution is proposed to overcome the halo and shade-off domains in order to reliably obtain the distribution and morphology of the LCN in the tomographic data. Convolutional neural network (CNN) models are utilized with increasing numbers of images, repeatedly validated by `error loss' and `accuracy' metrics. U-Net and Sensor3D CNN models were trained on data obtained from two different synchrotron Zernike phase-contrast transmission X-ray microscopes, the ANATOMIX beamline at SOLEIL (Paris, France) and the P05 beamline at PETRA III (Hamburg, Germany). The Sensor3D CNN model with a smaller batch size of 32 and a training data size of 70 images showed the best performance (accuracy 0.983 and error loss 0.032). The analysis procedures, validated by comparison with human-identified ground-truth images, correctly identified the voids within the bone matrix. This proposed approach may have further application to classify structures in volumetric images that contain non-linear artifacts that degrade image quality and hinder feature identification.
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Aprendizado Profundo , Animais , Humanos , Artefatos , Porosidade , Peixe-Zebra , Osso e Ossos/diagnóstico por imagem , Processamento de Imagem Assistida por Computador/métodosRESUMO
Diabetes is a major concern of our society as it affects one person out of 11 around the world. Elastic fiber alterations due to diabetes increase the stiffness of large arteries, but the structural effects of these alterations are poorly known. To address this issue, we used synchrotron X-ray microcomputed tomography with in-line phase contrast to image in three dimensions C57Bl6J (control) and db/db (diabetic) mice with a resolution of 650 nm/voxel and a field size of 1.3 mm3. Having previously shown in younger WT and db/db mouse cohorts that elastic lamellae contain an internal supporting lattice, here we show that in older db/db mice the elastic lamellae lose this scaffold. We coupled this label-free method with automated image analysis to demonstrate that the elastic lamellae from the arterial wall are structurally altered and become 11% smoother (286,665 measurements). This alteration suggests a link between the loss of the 3D lattice-like network and the waviness of the elastic lamellae. Therefore, waviness measurement appears to be a measurable elasticity indicator and the 3D lattice-like network appears to be at the origin of the existence of this waviness. Both could be suitable indicators of the overall elasticity of the aorta.
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Diabetes Mellitus , Síncrotrons , Idoso , Animais , Aorta/diagnóstico por imagem , Tecido Elástico , Elasticidade , Humanos , Camundongos , Microtomografia por Raio-XRESUMO
Background: Magnesium (Mg) is one of the most promising materials for human use in surgery due to material characteristics such as its elastic modulus as well as its resorbable and regenerative properties. In this study, HF-coated and uncoated novel bioresorbable magnesium fixation screws for maxillofacial and dental surgical applications were investigated in vitro and in vivo to evaluate the biocompatibility of the HF coating. Methods: Mg alloy screws that had either undergone a surface treatment with hydrofluoric-acid (HF) or left untreated were investigated. In vitro investigation included XTT, BrdU and LDH in accordance with the DIN ISO 10993-5/-12. In vivo, the screws were implanted into the tibia of rabbits. After 3 and 6 weeks, degradation, local tissue reactions and bony integration were analyzed histopathologically and histomorphometrically. Additionally, SEM/EDX analysis and synchrotron phase-contrast microtomography (µCT) measurements were conducted. The in vitro analyses revealed that the Mg screws are cytocompatible, with improved results when the surface had been passivated with HF. In vivo, the HF-treated Mg screws implanted showed a reduction in gas formation, slower biodegradation and a better bony integration in comparison to the untreated Mg screws. Histopathologically, the HF-passivated screws induced a layer of macrophages as part of its biodegradation process, whereas the untreated screws caused a slight fibrous tissue reaction. SEM/EDX analysis showed that both screws formed a similar layer of calcium phosphates on their surfaces and were surrounded by bone. Furthermore, the µCT revealed the presence of a metallic core of the screws, a faster absorbing corrosion front and a slow absorbing region of corroded magnesium. Conclusions: Overall, the HF-passivated Mg fixation screws showed significantly better biocompatibility in vitro and in vivo compared to the untreated screws.
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Regeneração Óssea/efeitos dos fármacos , Parafusos Ósseos/efeitos adversos , Regeneração Tecidual Guiada , Magnésio/farmacologia , Animais , Regeneração Óssea/genética , Linhagem Celular , Materiais Revestidos Biocompatíveis/química , Materiais Revestidos Biocompatíveis/farmacologia , Humanos , Ácido Fluorídrico/química , Ácido Fluorídrico/farmacologia , Magnésio/efeitos adversos , Teste de Materiais , Camundongos , Procedimentos Cirúrgicos Ortognáticos/tendências , Osteoblastos/efeitos dos fármacosRESUMO
Double-sided Fresnel zone plates are diffractive lenses used for high-resolution hard X-ray microscopy. The double-sided structures have significantly higher aspect ratios compared with single-sided components and hence enable more efficient imaging. The zone plates discussed in this paper are fabricated on each side of a thin support membrane, and the alignment of the zone plates with respect to each other is critical. Here, a simple and reliable way of quantifying misalignments by recording efficiency maps and measuring the absolute diffraction efficiency of the zone plates as a function of tilting angle in two directions is presented. The measurements are performed in a setup based on a tungsten-anode microfocus X-ray tube, providing an X-ray energy of 8.4â keV through differential measurements with a Cu and an Ni filter. This study investigates the sources of the misalignments and concludes that they can be avoided by decreasing the structure heights on both sides of the membrane and by pre-programming size differences between the front- and back-side zone plates.
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OBJECTIVES: An increasing number of CAD/CAM (computer-aided design/computer-aided manufacturing) hybrid materials have been introduced to the dental market in recent years. In addition, CAD/CAM hybrid materials for additive manufacturing (AM) are becoming more attractive in digital dentistry. Studies on material microstructures using micro-computed tomography (µ-CT) combined with scanning electron microscopy (SEM) have only been available to a limited extent so far. METHODS: One CAD/CAM three-dimensional- (3D-) printable hybrid material (VarseoSmile Crown plus) and two CAD/CAM millable hybrid materials (Vita Enamic; Voco Grandio), as well as one direct composite material (Ceram.x duo), were included in the present study. Cylindrical samples with a diameter of 2 mm were produced from each material and investigated by means of synchrotron radiation µ-CT at a voxel size of 0.65 µm. Different samples from the same materials, obtained by cutting and polishing, were investigated by SEM. RESULTS: The 3D-printed hybrid material showed some agglomerations and a more irregular distribution of fillers, as well as a visible layered macrostructure and a few spherical pores due to the printing process. The CAD/CAM millable hybrid materials revealed a more homogenous distribution of ceramic particles. The direct composite material showed multiple air bubbles and microstructural irregularities based on manual processing. SIGNIFICANCE: The µ-CT and SEM analysis of the materials revealed different microstructures even though they belong to the same class of materials. It could be shown that µ-CT and SEM imaging are valuable tools to understand microstructure and related mechanical properties of materials.
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Desenho Assistido por Computador , Materiais Dentários , Teste de Materiais , Microscopia Eletrônica de Varredura , Propriedades de Superfície , Microtomografia por Raio-X , Materiais Dentários/química , Resinas Compostas/química , Impressão Tridimensional , Cerâmica/químicaRESUMO
Cobalt-chromium-molybdenum (CoCrMo) alloys are routinely used in arthroplasty. CoCrMo wear particles and ions derived from arthroplasty implants lead to macrophage-driven adverse local tissue reactions, which have been linked to an increased risk of periprosthetic joint infection after revision arthroplasty. While metal-induced cytotoxicity is well characterized in human macrophages, direct effects on their functionality have remained elusive. Synchrotron radiation X-ray microtomography and X-ray fluorescence mapping indicated that peri-implant tissues harvested during aseptic revision of different arthroplasty implants are exposed to Co and Cr in situ. Confocal laser scanning microscopy revealed that macrophage influx is predominant in patient tissue. While in vitro exposure to Cr3+ had only minor effects on monocytes/macrophage phenotype, pathologic concentrations of Co2+ significantly impaired both, monocyte/macrophage phenotype and functionality. High concentrations of Co2+ led to a shift in macrophage subsets and loss of surface markers, including CD14 and CD16. Both Co2+ and Cr3+ impaired macrophage responses to Staphylococcus aureus infection, and particularly, Co2+-exposed macrophages showed decreased phagocytic activity. These findings demonstrate the immunosuppressive effects of locally elevated metal ions on the innate immune response and support further investigations, including studies exploring whether Co2+ and Cr3+ or CoCrMo alloys per se expose the patients to a higher risk of infections post-revision arthroplasty.
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Cobalto , Infecções Estafilocócicas , Humanos , Cobalto/toxicidade , Cromo/toxicidade , Staphylococcus aureus , Macrófagos/patologia , Íons/farmacologia , Ligas , Infecções Estafilocócicas/patologiaRESUMO
Deformation of the first crystal of an X-ray monochromator under the heat load of a high-power beam, commonly referred to as `heat bump', is a challenge frequently faced at synchrotron beamlines. Here, quantitative measurements of the deformations of an externally water-cooled silicon (111) double-crystal monochromator tuned to a photon energy of 17.6 keV are reported. These measurements were made using two-dimensional hard X-ray grating interferometry, a technique that enables in situ at-wavelength wavefront investigations with high angular sensitivity. The observed crystal deformations were of the order of 100 nm in the meridional and 5 nm in the sagittal direction, which lead to wavefront slope errors of up to 4 µrad in the meridional and a few hundred nanoradians in the sagittal direction.
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Potential applications of grating-based X-ray phase-contrast imaging are investigated in various fields due to its compatibility with laboratory X-ray sources. So far the method was mainly restricted to X-ray energies below 40 keV, which is too low to examine dense or thick objects, but a routine operation at higher energies is on the brink of realisation. In this study, imaging results obtained at 82 keV are presented. These comprise a test object consisting of well-defined materials for a quantitative analysis and a tooth to translate the findings to a biomedical sample. Measured linear attenuation coefficients ? and electron densities ?e are in good agreement with theoretical values. Improved contrast-to-noise ratios were found in phase contrast compared to attenuation contrast. The combination of both contrast modalities further enables to simultaneously assess information on density and composition of materials with effective atomic numbers Z? > 8. In our biomedical example, we demonstrate the possibility to detect differences in mass density and calcium concentration within teeth.
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Intensificação de Imagem Radiográfica/instrumentação , Tomografia Computadorizada por Raios X/instrumentação , Difração de Raios X/instrumentação , Desenho de Equipamento , Análise de Falha de EquipamentoRESUMO
OBJECTIVE: Limited contrast between healthy and tumour tissue is a limiting factor in mammography and CT of the breast. Phase-contrast computed tomography (PC-CT) provides improved soft-tissue contrast compared with absorption-based techniques. In this study, we assessed the technical feasibility of grating-based PC-CT imaging of the breast for characterisation of ductal carcinoma in situ (DCIS). METHODS: Grating-based PC-CT was performed on one breast specimen containing an invasive ductal carcinoma and DCIS using monochromatic radiation of 23 keV. Phase-contrast and absorption-based images were compared qualitatively and quantitatively with histopathology in a blinded fashion. RESULTS: Grating-based PC-CT showed improved differentiation of soft-tissue components. Circular structures of high phase-shift contrast corresponding to the walls of the dilated ductuli of the DCIS were visualised with a contrast-to-noise ratio (CNR) of 9.6 using PC-CT but were not detectable on absorption-based images (CNR = 0.27). The high phase-shift structures of the dilated ductuli were identifiable in the PC-CT volume data set allowing for 3D characterisation of DCIS. CONCLUSIONS: Our results indicate that unlike conventional CT, grating-based PC-CT may allow the differentiation between invasive carcinoma and intraductal carcinoma and healthy breast tissue and provide 3D visualisation of DCIS.
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Neoplasias da Mama/diagnóstico por imagem , Carcinoma Ductal de Mama/diagnóstico por imagem , Carcinoma Intraductal não Infiltrante/diagnóstico por imagem , Interpretação de Imagem Radiográfica Assistida por Computador , Tomografia Computadorizada por Raios X/métodos , Idoso , Biópsia por Agulha , Neoplasias da Mama/patologia , Neoplasias da Mama/cirurgia , Carcinoma Ductal de Mama/patologia , Carcinoma Ductal de Mama/cirurgia , Carcinoma Intraductal não Infiltrante/patologia , Carcinoma Intraductal não Infiltrante/cirurgia , Meios de Contraste , Diagnóstico Diferencial , Detecção Precoce de Câncer/métodos , Feminino , Humanos , Imuno-Histoquímica , Mamografia/métodos , Mastectomia/métodos , Intensificação de Imagem Radiográfica/métodos , Manejo de EspécimesRESUMO
Nephrotoxicity is an important drug safety aspect to be assessed during drug discovery and development. To study renal toxicity, in vitro cell-based assays are often used. Unfortunately, translating the results of such cell assays to vertebrates including human remains challenging. Therefore, we aim to evaluate whether zebrafish larvae (ZFL) could serve as a vertebrate screening model to detect gentamicin-induced changes of kidney glomeruli and proximal tubules. To validate the model, we compared the results of ZFL with those obtained from kidney biopsies of gentamicin-treated mice. We used transgenic zebrafish lines expressing enhanced green fluorescent proteins in the glomerulus to visualize glomerular damage. Synchrotron radiation-based computed tomography (SRµCT) is a label-free approach providing three-dimensional representations of renal structures with micrometre resolution. Clinically used gentamicin concentrations induce nephrotoxicity and affect glomerular and proximal tubular morphology. Findings were confirmed in mice and ZFL. There was a strong correlation between fluorescent signals in ZFL, SRµCT- derived descriptors of glomerular and proximal tubular morphology and the histological analysis of mouse kidney biopsies. A combination of SRµCT and confocal microscopy provides unprecedented insights into anatomical structures of the zebrafish kidney. Based on our findings, we suggest to use ZFL as a predictive vertebrate screening model to study drug-induced nephrotoxicity and to bridge the gap between cell culture-based test systems and experiments in mammals.
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Nefropatias , Peixe-Zebra , Humanos , Animais , Camundongos , Gentamicinas/toxicidade , Larva , Rim/diagnóstico por imagem , Rim/patologia , Glomérulos Renais/patologia , Nefropatias/patologia , MamíferosRESUMO
In musculoskeletal surgery, the treatment of large bone defects is challenging and can require the use of bone graft substitutes to restore mechanical stability and promote host-mediated regeneration. The use of bone allografts is well-established in many bone regenerative procedures, but is associated with low rates of ingrowth due to pre-therapeutic graft processing. Cold physical plasma (CPP), a partially ionized gas that simultaneously generates reactive oxygen (O2) and nitrogen (N2) species, is suggested to be advantageous in biomedical implant processing. CPP is a promising tool in allograft processing for improving surface characteristics of bone allografts towards enhanced cellularization and osteoconduction. However, a preclinical assessment regarding the feasibility of pre-therapeutic processing of allogeneic bone grafts with CPP has not yet been performed. Thus, this pilot study aimed to analyze the bone morphology of CPP processed allografts using synchrotron radiation-based microcomputed tomography (SR-µCT) and to analyze the effects of CPP processing on human bone cell viability and function. The analyzes, including co-registration of pre- and post-treatment SR-µCT scans, revealed that the main bone morphological properties (total volume, mineralized volume, surface area, and porosity) remained unaffected by CPP treatment if compared to allografts not treated with CPP. Varying effects on cellular metabolic activity and alkaline phosphatase activity were found in response to different gas mixtures and treatment durations employed for CPP application. It was found that 3 min CPP treatment using a He + 0.1% N2 gas mixture led to the most favourable outcome regarding a significant increase in bone cell viability and alkaline phosphatase activity. This study highlights the promising potential of pre-therapeuthic bone allograft processing by CPP prior to intraoperative application and emphasizes the need for gas source and treatment time optimization for specific applications.
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Amyloid-ß (Aß) plaques from Alzheimer's Disease (AD) can be visualized ex vivo in label-free brain samples using synchrotron X-ray phase-contrast tomography (XPCT). However, for XPCT to be useful as a screening method for amyloid pathology, it is essential to understand which factors drive the detection of Aß plaques. The current study was designed to test the hypothesis that Aß-related contrast in XPCT could be caused by Aß fibrils and/or by metals trapped in the plaques. Fibrillar and elemental compositions of Aß plaques were probed in brain samples from different types of AD patients and AD models to establish a relationship between XPCT contrast and Aß plaque characteristics. XPCT, micro-Fourier-Transform Infrared spectroscopy and micro-X-Ray Fluorescence spectroscopy were conducted on human samples (one genetic and one sporadic case) and on four transgenic rodent strains (mouse: APPPS1, ArcAß, J20; rat: TgF344). Aß plaques from the genetic AD patient were visible using XPCT, and had higher ß-sheet content and higher metal levels than those from the sporadic AD patient, which remained undetected by XPCT. Aß plaques in J20 mice and TgF344 rats appeared hyperdense on XPCT images, while they were hypodense with a hyperdense core in the case of APPPS1 and ArcAß mice. In all four transgenic strains, ß-sheet content was similar, while metal levels were highly variable: J20 (zinc and iron) and TgF344 (copper) strains showed greater metal accumulation than APPPS1 and ArcAß mice. Hence, a hyperdense contrast formation of Aß plaques in XPCT images was associated with biometal entrapment within plaques. STATEMENT OF SIGNIFICANCE: The role of metals in Alzheimer's disease (AD) has been a subject of continuous interest. It was already known that amyloid-ß plaques (Aß), the earliest hallmark of AD, tend to trap endogenous biometals like zinc, iron and copper. Here we show that this metal accumulation is the main reason why Aß plaques are detected with a new technique called X-ray phase contrast tomography (XPCT). XPCT enables to map the distribution of Aß plaques in the whole excised brain without labeling. In this work we describe a unique collection of four transgenic models of AD, together with a human sporadic and a rare genetic case of AD, thus exploring the full spectrum of amyloid contrast in XPCT.
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Doença de Alzheimer , Oligoelementos , Humanos , Camundongos , Animais , Ratos , Doença de Alzheimer/diagnóstico por imagem , Doença de Alzheimer/patologia , Cobre/química , Raios X , Camundongos Transgênicos , Peptídeos beta-Amiloides/metabolismo , Metais , Zinco/química , Ferro , Encéfalo/metabolismo , Amiloide , Placa Amiloide/diagnóstico por imagem , Placa Amiloide/química , Modelos Animais de DoençasRESUMO
The visualization of the vascular network in tumors down to the smallest vessels requires high spatial resolution and reasonable contrast. Stained corrosion casts of the microvasculature network guarantee superior X-ray absorption contrast and highest reproduction fidelity. Tomography of a centimeter-size tumor, however, is unfeasible at the spatial resolution needed to reveal the smallest vessels. Therefore, local tomography has been performed to visualize the smallest capillaries within the region of interest. These three-dimensional data show the detailed morphology, but the reconstructed absorption coefficients obtained in local tomography differ substantially from the absorption coefficients retrieved from the less detailed global tomography data. This paper deals with the adaptation of local tomography data using the global data and considers two-parameter histogram matching of the radiographs, sinogram extension, and multi-parameter cupping correction. It is demonstrated that two-parameter histogram matching of the radiographs already provides reasonable agreement. The change of the lens in front of the detector's camera, however, significantly affects the obtained local X-ray absorption coefficients in the tomograms predominantly owing to the dissimilar point-spread functions of the two configurations used, and much less to the fact that one of the data sets was acquired in a local geometry.
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Neoplasias do Colo/irrigação sanguínea , Neoplasias do Colo/diagnóstico por imagem , Imageamento Tridimensional/métodos , Animais , Capilares/anatomia & histologia , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neovascularização Patológica , Síncrotrons , Tomografia Computadorizada por Raios X/métodos , Microtomografia por Raio-X/métodosRESUMO
A key issue in developing strategies against diseases such as cancer is the analysis of the vessel tree in comparison to the healthy one. In the search for parameters that might be characteristic for tumor capillaries we study the vascularization in mice for cancerous and healthy tissues using synchrotron radiation-based micro computed tomography in absorption and phase contrast modes. Our investigations are based on absorption tomograms of casted healthy and cancerous tissues as well as a phase tomogram of a fixated tumor. We demonstrate how the voxel-based tomography data can be vectorized to assess the capillary networks quantitatively. The processing includes segmentation, skeletonization, and vectorization to finally extract the vessel parameters. The mean diameter of capillaries in healthy and cancerous tissues corresponds to (8.0±1.1) µm and (3.9±1.1) µm, respectively. Further evaluated parameters show marginal or no differences between capillaries in healthy and cancerous tissues, namely fractal dimension 2.3±0.3 vs. 2.3±0.2, tortuosity (SOAM) 0.18 rad/µm vs. 0.24 rad/µm and vessel length 20 µm vs. 17 µm. The bifurcation angles exhibit a narrow distribution around 115°. Furthermore, we show that phase tomography is a powerful alternative to absorption tomography of casts for the vessel visualization omitting any invasive specimen preparation procedure.
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Capilares/patologia , Neoplasias/patologia , Absorção , Animais , Linhagem Celular Tumoral , Fractais , Humanos , Processamento de Imagem Assistida por Computador/métodos , Imageamento Tridimensional , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Microcirculação , Microscopia de Contraste de Fase/métodos , Transplante de Neoplasias , Fótons , Microtomografia por Raio-X/métodosRESUMO
Synchrotron X-rays micro-computed tomography was applied to visualize and quantify 3D ice crystal changes into a model sponge cake after freezing and subsequent frozen storage. Model sponge cake samples were submitted to two different freezing rates (fast: 17.2 °C min-1 and slow: 0.3 °C min-1), then stored at constant and fluctuating temperatures over a two weeks period. 3D images were acquired at frozen state thanks to a thermostated cell (CellStat) and processed using a grey level based segmentation method. Image analysis revealed that the ice volume fraction is conserved during storage but ice crystal size and location change whatever the freezing rate and the storage conditions. Maximum local thicknesses increase both inside (from 20 µm to 50 µm) and outside (from 47 µm to 70 µm) the matrix during the fourteen days storage period. Both specific surface areas between starch and ice (SSAice/starch) and between air and ice (SSAair/ice) also evolve with storage duration: SSAice/starch decreases up to - 30 % while SSAair/ice increases up to + 13 % depending on the freezing rates and the storage conditions. These results highlighted that, during storage, ice crystals evolve according to two different mechanisms depending on the freezing rate: fast freezing leads to a local redistribution of water both within the starch matrix and within the pores, while slow freezing results in both local redistribution within the starch matrix and water migration towards the pores. In addition, stable storage temperatures favor local water redistribution whereas water migration from the starch matrix towards the pores was greater in the case of fluctuating storage temperatures. This study shows that freezing and frozen storage conditions have a synergistic effect on the microstructure evolution of sponge cake due to recrystallization phenomena.
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Gelo , Síncrotrons , Microtomografia por Raio-X , Congelamento , Raios X , Amido , ÁguaRESUMO
BACKGROUND: Fixation and embedding of post mortem brain tissue is a pre-requisite for both gold-standard conventional histology and X-ray virtual histology. This process alters the morphology and density of the brain microanatomy. NEW METHOD: To quantify these changes, we employed synchrotron radiation-based hard X-ray tomography with 3 µm voxel length to visualize the same mouse brain after fixation in 4% formalin, immersion in ethanol solutions (50%, 70%, 80%, 90%, and 100%), xylene, and finally after embedding in a paraffin block. The volumetric data were non-rigidly registered to the initial formalin-fixed state to align the microanatomy within the entire mouse brain. RESULTS: Volumetric strain fields were used to characterize local shrinkage, which was found to depend on the anatomical region and distance to external surface. X-ray contrast was altered and enhanced by preparation-induced inter-tissue density changes. The preparation step can be selected to highlight specific anatomical features. For example, fiber tract contrast is amplified in 100% ethanol. COMPARISON WITH EXISTING METHODS: Our method provides volumetric strain fields, unlike approaches based on feature-to-feature or volume measurements. Volumetric strain fields are produced by non-rigid registration, which is less labor-intensive and observer-dependent than volume change measurements based on manual segmentations. X-ray microtomography provides spatial resolution at least an order of magnitude higher than magnetic resonance microscopy, allowing for analysis of morphology and density changes within the brain's microanatomy. CONCLUSION: Our approach belongs to three-dimensional virtual histology with isotropic micrometer spatial resolution and therefore complements atlases based on a combination of magnetic resonance microscopy and optical micrographs of serial histological sections.
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Encéfalo , Formaldeído , Animais , Encéfalo/diagnóstico por imagem , Camundongos , Inclusão em Parafina , Síncrotrons , Microtomografia por Raio-X/métodosRESUMO
Purpose: Synchrotron radiation-based tomography yields microanatomical features in human and animal tissues without physical slicing. Recent advances in instrumentation have made laboratory-based phase tomography feasible. We compared the performance of three cutting-edge laboratory systems benchmarked by synchrotron radiation-based tomography for three specimens. As an additional criterion, the user-friendliness of the three microtomography systems was considered. Approach: The three tomography systems-SkyScan 2214 (Bruker-microCT, Kontich, Belgium), Exciscope prototype (Stockholm, Sweden), and Xradia 620 Versa (Zeiss, Oberkochen, Germany)-were given 36 h to measure three medically relevant specimens, namely, zebrafish larva, archaeological human tooth, and porcine nerve. The obtained datasets were registered to the benchmark synchrotron radiation-based tomography from the same specimens and selected ones to the SkyScan 1275 and phoenix nanotom m® laboratory systems to characterize development over the last decade. Results: Next-generation laboratory-based microtomography almost reached the quality achieved by synchrotron-radiation facilities with respect to spatial and density resolution, as indicated by the visualization of the medically relevant microanatomical features. The SkyScan 2214 system and the Exciscope prototype demonstrated the complementarity of phase information by imaging the eyes of the zebrafish larva. The 3 - µ m thin annual layers in the tooth cementum were identified using Xradia 620 Versa. Conclusions: SkyScan 2214 was the simplest system and was well-suited to visualizing the wealth of anatomical features in the zebrafish larva. Data from the Exciscope prototype with the high photon flux from the liquid metal source showed the spiral nature of the myelin sheaths in the porcine nerve. Xradia 620 Versa, with detector optics as typically installed for synchrotron tomography beamlines, enabled the three-dimensional visualization of the zebrafish larva with comparable quality to the synchrotron data and the annual layers in the tooth cementum.
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We present in this Letter a type of quadriwave lateral shearing interferometer for x-ray phase imaging. This device is based on a phase chessboard, and we take advantage of the large spectrum of the source to produce interferograms with a propagation-invariant contrast. Such a grating has been created for hard x-ray interferometry and experimentally tested on a synchrotron beamline at Soleil.
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BACKGROUND: Micrometer-resolution neuroimaging with gold-standard conventional histology requires tissue fixation and embedding. The exchange of solvents for the creation of sectionable paraffin blocks modifies tissue density and generates non-uniform brain shrinkage. NEW METHOD: We employed synchrotron radiation-based X-ray microtomography for slicing- and label-free virtual histology of the mouse brain at different stages of the standard preparation protocol from formalin fixation via ascending ethanol solutions and xylene to paraffin embedding. Segmentation of anatomical regions allowed us to quantify non-uniform tissue shrinkage. Global and local changes in X-ray absorption gave insight into contrast enhancement for virtual histology. RESULTS: The volume of the entire mouse brain was 60%, 56%, and 40% of that in formalin for, respectively, 100% ethanol, xylene, and paraffin. The volume changes of anatomical regions such as the hippocampus, anterior commissure, and ventricles differ from the global volume change. X-ray absorption of the full brain decreased, while local absorption differences increased, resulting in enhanced contrast for virtual histology. These trends were also observed with laboratory microtomography measurements. COMPARISON WITH EXISTING METHODS: Microtomography provided sub-10 µm spatial resolution with sufficient density resolution to resolve anatomical structures at each step of the embedding protocol. The spatial resolution of conventional computed tomography and magnetic resonance microscopy is an order of magnitude lower and both do not match the contrast of microtomography over the entire embedding protocol. Unlike feature-to-feature or total volume measurements, our approach allows for calculation of volume change based on segmentation. CONCLUSION: We present isotropic micrometer-resolution imaging to quantify morphology and composition changes in a mouse brain during the standard histological preparation. The proposed method can be employed to identify the most appropriate embedding medium for anatomical feature visualization, to reveal the basis for the dramatic X-ray contrast enhancement observed in numerous embedded tissues, and to quantify morphological changes during tissue fixation and embedding.
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Formaldeído , Imageamento Tridimensional , Animais , Encéfalo/diagnóstico por imagem , Camundongos , Inclusão em Parafina , Microtomografia por Raio-XRESUMO
A study of the coherence and wavefront properties of a pseudo-channel-cut monochromator in comparison with a double-crystal monochromator is presented. Using a double-grating interferometer designed for the hard X-ray regime, the complex coherence factor was measured and the wavefront distortions at the sample position were analyzed. A transverse coherence length was found in the vertical direction that was a factor of two larger for the channel-cut monochromator owing to its higher mechanical stability. The wavefront distortions after different optical elements in the beam, such as monochromators and mirrors, were also quantified. This work is particularly relevant for coherent diffraction imaging experiments with synchrotron sources.