The Study of the Caudal Vertebrae of Thick-Toed Geckos after a Prolonged Space Flight by X-ray Phase-Contrast Micro-CT.

The proximal caudal vertebrae and notochord in thick-toed geckos (TG) (Chondrodactylus turneri, Gray, 1864) were investigated after a 30-day space flight onboard the biosatellite Bion-M1. This region has not been explored in previous studies. Our research focused on finding sites most affected by demineralization caused by microgravity (G0). We used X-ray phase-contrast tomography to study TG samples without invasive prior preparation to clarify our previous findings on the resistance of TG's bones to demineralization in G0. The results of the present study confirmed that geckos are capable of preserving bone mass after flight, as neither cortical nor trabecular bone volume fraction showed statistically significant changes after flight. On the other hand, we observed a clear decrease in the mineralization of the notochordal septum and a substantial rise in intercentrum volume following the flight. To monitor TG's mineral metabolism in G0, we propose to measure the volume of mineralized tissue in the notochordal septum. This technique holds promise as a sensitive approach to track the demineralization process in G0, given that the volume of calcification within the septum is limited, making it easy to detect even slight changes in mineral content.

Crystal Analyzer Based Multispectral Microtomography Using CCD-Sensor.

To solve the problems of spectral tomography, an X-ray optical scheme was proposed, using a crystal analyzer in Laue geometry between the sample and the detector, which allowed for the selection of predetermined pairs of wavelengths from the incident polychromatic radiation to obtain projection images. On a laboratory X-ray microtomography setup, an experiment was carried out for the first time where a mixture of micro-granules of sodium chloride NaCl, silver behenate AgC22H43O2, and lithium niobate LiNbO3 was used as a test sample to identify their spatial arrangement. The elements were chosen based on the presence of absorption edges in two of the elements in the energy range of the polychromatic spectrum of the probing radiation. The method of projection distortion correction was used to preprocess the obtained projections. To interpret the obtained reconstruction results, the segmentation method based on the analysis of joint histograms was used. This allowed us to identify each of the three substances. To compare the results obtained, additional "reference" tomographic measurements were performed: one in polychromatic and two in monochromatic (MoKα-, MoKß-lines) modes. It took three times less time for the tomographic experiment with the crystal analyzer, while the reconstruction accuracy was comparable to that of the "reference" tomography.

Micromorphology of pineal gland calcification in age-related neurodegenerative diseases.

BACKGROUND: The formation of concrements in human pineal gland (PG) is a physiological process and, according to many researchers, is associated with the involution of PG structures. The majority of scientific publications concern progressive calcification of PG, leaving out studies on the destruction of already formed calcified concrements. Our study fills the gap in knowledge about calcified zones destruction in PG in normal aging and neuropathological conditions, which has not been addressed until now. PURPOSE: Our objective is to gain insight into human PG tissue impairment in both normal aging and neurodegenerative conditions. X-ray phase-contrast tomography (XPCT) allowed us to study PG tissue degeneration at high spatial resolution and, for the first time, to examine the damaged PG concrements in detail. Our research finding could potentially enhance the understanding of the PG involvement in the process of aging as well as in Alzheimer's disease (AD) and vascular dementia (VD). METHODS: The research was carried out on human PG autopsy material in normal aging, VD, and AD conditions. Laboratory-based micro-computed tomography (micro-CT) was used to collect and evaluate samples of native, uncut, and unstained PG with different degrees of pineal calcification. The detailed high-resolution 3D images of the selected PGs were produced using synchrotron-based XPCT. Histology and immunohistochemistry of soft PG tissue confirmed XPCT results. RESULTS: We performed via micro-CT the evaluation of the morphometric parameters of PG such as total sample volume, calcified concrements volume, and percentage of concrements in the total volume of the sample. XPCT imaging revealed high-resolution details of age-related PG alteration. In particular, we noted signs of moderate degradation of concrements in some PGs from elderly donors. In addition, our analysis revealed noticeable degenerative change in both concrements and soft tissue of PGs with neuropathology. In particular, we observed a hollow core and separated layers as well as deep ragged cracks in PG concrements of AD and VD samples. In parenchyma of some samples, we detected wide pinealocyte-free fluid-filled areas adjacent to the calcified zones. CONCLUSION: The present work provides the basis for future scientific research focused on the dynamic nature of PG calcium deposits and PG soft tissue in normal aging and neurodegenerative diseases.

Vacuum-free femtosecond fiber laser microplasma X-ray source for radiography.

Radiographic imaging using X-rays is a tool for basic research and applications in industry, materials science, and medical diagnostics. In this article, we present a novel approach for the generation of X-rays using a vacuum-free microplasma by femtosecond fiber laser. By tightly focusing a laser pulse onto a micrometer-sized solid density near-surface plasma from a rotating copper target, we demonstrate the generation of Cu K-photons (8-9 keV) with high yield ∼ 1.6 × 109 phot/s/2π, and with a source size diameter of approximately 10 microns. Femtosecond fiber laser allows working with a high repetition rate (∼2 MHz) and moderate energy levels (10-40 µJ), ensuring the effective quasi-continuous generation of X-ray photons. Furthermore, we introduce a hybrid scheme that combines the tightly focusing laser-plasma X-ray generator with an online control unit for microplasma size source based on the back-reflected second harmonic generated in the laser-induced microplasma. The compactness and high performance of this vacuum-free femtosecond fiber laser microplasma X-ray source makes it a promising solution for advanced radiographic applications. Our preliminary results on the creation of a microfocus X-ray source provide insights into the feasibility and potential of this innovative approach.

Deep Learning-Based Segmentation of Post-Mortem Human's Olfactory Bulb Structures in X-ray Phase-Contrast Tomography.

The human olfactory bulb (OB) has a laminar structure. The segregation of cell populations in the OB image poses a significant challenge because of indistinct boundaries of the layers. Standard 3D visualization tools usually have a low resolution and cannot provide the high accuracy required for morphometric analysis. X-ray phase contrast tomography (XPCT) offers sufficient resolution and contrast to identify single cells in large volumes of the brain. The numerous microanatomical structures detectable in XPCT image of the OB, however, greatly complicate the manual delineation of OB neuronal cell layers. To address the challenging problem of fully automated segmentation of XPCT images of human OB morphological layers, we propose a new pipeline for tomographic data processing. Convolutional neural networks (CNN) were used to segment XPCT image of native unstained human OB. Virtual segmentation of the whole OB and an accurate delineation of each layer in a healthy non-demented OB is mandatory as the first step for assessing OB morphological changes in smell impairment research. In this framework, we proposed an effective tool that could help to shed light on OB layer-specific degeneration in patients with olfactory disorder.

Assessment of dental implant surface stability at the nanoscale level.

OBJECTIVES: To study the oxide layer stability of certified dental implants of system "P", made based on TiO2 alloy with carbon coating. To perform a comparative statistical analysis of the obtained data with the available data for the dental implants of systems "A" and "B". METHODS: X-ray microtomography and X-ray fluorescence analysis were used to study soft tissue biopsy specimens. Supernatants were studied by dynamic light scattering and transmission electron microscopy when simulating free emission of nanoscale metal oxide particles from the surface of dental implants as well as when simulating physical loading. A comparative analysis of three parameters of nanoscale particles was performed by statistical data analysis. The surface of the "P" system dental implant with surface treatment was analyzed by scanning electron microscopy. RESULTS: Both free emission of nanoscale oxide layer particles and yield of nano- and microscale particles during simulation of physical load were confirmed. Statistically significant differences were noted in a comparative analysis of the size and frequency of occurrence of these particles in the supernatants obtained from the surfaces of three dental implant systems. The elemental composition of the particles and the composition and structure of the "P" system dental implants themselves were analyzed. SIGNIFICANCE: The developed method of dynamic light scattering can be used to compare the stability of the oxide layer of standardized medical products manufactured on the basis of the TiO2 alloy.

Micro-CT Study of Mongolian Gerbil Humeral Bone After Prolonged Spaceflight Based on a New Algorithm for Delimitation of Long-Bone Regions.

The Mongolian gerbil displays unique physiological and anatomical features that make this species an attractive object for biological experiments in space. However, until recently, the Mongolian gerbil has remained a novel, mostly unstudied animal model in investigating bone loss in weightlessness (G0). After 12 days of orbital Foton-M3 mission, the humerus of Mongolian gerbils has been studied here via micro-computed tomography (micro-CT) to quantify bone morphometric parameters. The samples from the flight group, delayed synchronous ground-control group, and basal control group were investigated, and main morphometric parameters were reported in the article. The accurate selection of a region of interest is an essential step for a correct assessment of bone parameters. We proposed a new, easy and efficient method for delimiting the bone's basic regions in the humerus. It is based on quantitative estimation of X-ray attenuation in the cortical bone as a function of humerus bone length. The micro-CT analysis of the basic bone regions revealed a difference in bone morphometric parameters between the flight and control gerbils. The most significant bone loss was observed in the cortical part of the proximal humeral zone in the flight group. No statistically significant changes of volume fraction in the cancellous tissue of proximal and distal epiphyses and metaphyses were observed. A statistically significant increase in both cancellous bone volume and bone X-ray attenuation in the flight group was detected in the proximal part of the diaphyses. We assume that enhanced calcium deposition in the diaphyseal cancellous tissue occurred due to a bone response to G0 conditions.

Monitored Tomographic Reconstruction-An Advanced Tool to Study the 3D Morphology of Nanomaterials.

Detailed and accurate three-dimensional (3D) information about the morphology of hierarchically structured materials is derived from multi-scale X-ray computed tomography (XCT) and subsequent 3D data reconstruction. High-resolution X-ray microscopy and nano-XCT are suitable techniques to nondestructively study nanomaterials, including porous or skeleton materials. However, laboratory nano-XCT studies are very time-consuming. To reduce the time-to-data by more than an order of magnitude, we propose taking advantage of a monitored tomographic reconstruction. The benefit of this new protocol for 3D imaging is that the data acquisition for each projection is interspersed by image reconstruction. We demonstrate this new approach for nano-XCT data of a novel transition-metal-based materials system: MoNi4 electrocatalysts anchored on MoO2 cuboids aligned on Ni foam (MoNi4/MoO2@Ni). Quantitative data that describe the 3D morphology of this hierarchically structured system with an advanced electrocatalytically active nanomaterial are needed to tailor performance and durability of the electrocatalyst system. We present the framework for monitored tomographic reconstruction, construct three stopping rules for various reconstruction quality metrics and provide their experimental evaluation.

Wide-Ranging Multitool Study of Structure and Porosity of PLGA Scaffolds for Tissue Engineering.

In this study, the nanoscale transformation of the polylactic-co-glycolic acid (PLGA) internal structure, before and after its supercritical carbon dioxide (sc-CO2) swelling and plasticization, followed by foaming after a CO2 pressure drop, was studied by small-angle X-ray scattering (SAXS) for the first time. A comparative analysis of the internal structure data and porosity measurements for PLGA scaffolds, produced by sc-CO2 processing, on a scale ranging from 0.02 to 1000 µm, was performed by SAXS, helium pycnometry (HP), mercury intrusion porosimetry (MIP) and both "lab-source" and synchrotron X-ray microtomography (micro-CT). This approach opens up possibilities for the wide-scale evaluation, computer modeling, and prediction of the physical and mechanical properties of PLGA scaffolds, as well as their biodegradation behavior in the body. Hence, this study targets optimizing the process parameters of PLGA scaffold fabrication for specific biomedical applications.

Infrared Laser Effect on Healthy and Ossified Costal Cartilage: The Development of Stable Load-Bearing Autoimplants.

BACKGROUND AND OBJECTIVE: The laser-induced stress relaxation provides new prospects to obtain stable long fragments of costal cartilage for autoimplantation avoiding the risk of spontaneous deformation and poor engraftment. However, the age-related alterations of cartilage may sufficiently influence its interaction with infrared (IR) laser radiation and disrupt the effectiveness and safety of the technique. The aim of the work is to study the influence of the structural quality of costal cartilage on its interaction with IR laser and efficiency of obtaining of curved implants for trachea surgery. STUDY DESIGN/MATERIALS AND METHODS: Healthy costal cartilage was taken from pigs and human. Ossified costal cartilage was taken from humans of age 65 ± 7. The cartilage slices with a mean thickness of 3 mm were mechanically curved and processed to stress relaxation by laser irradiation with the wavelength 1.56 µm. The structure and mineral content were studied by X-ray microtomography and element analysis. The optical measurements included the study of the propagation of IR radiation, speckle interferometry, and IR radiometry. RESULTS: The aged cartilage demonstrates a high level of heterogeneity in structure and properties and decreased water content. The presence of dense inclusions consisting of amorphous calcined volumes makes the tissue more fragile and less elastic. The IR radiation propagation intensity for aged cartilage is at least twice higher than that for healthy cartilage. The thermal-induced motion of scatterers in aged cartilage is slower. X-ray microtomography showed the cartilage-like and the bone-like structures within the ossified samples. CONCLUSIONS: The main challenge for laser reshaping of aged cartilage is the presence of ossifications. However, the new stable curvature can be obtained with adjustment of laser power. To obtain the satisfying stable curvature of an implant the ossified volumes should be avoided The laser-induced stress-relaxation mechanism for aged cartilage can be particularly different from that of healthy tissue and the optimal laser regimes should be specified. Lasers Surg. Med. © 2020 Wiley Periodicals, Inc.

Enhanced Tomographic Sensing Multimodality with a Crystal Analyzer.

This article demonstrates how a combination of well-known tools-a standard 2D detector (CCD (charge-coupled device) camera) and a crystal analyzer-can improve the multimodality of X-ray imaging and tomographic sensing. The use of a crystal analyzer allowed two characteristic lines of the molybdenum anode-Kα and Kß-to be separated from the polychromatic radiation of the conventional X-ray tube. Thus, as a result of one measurement, three radiographic projections (images) were simultaneously recorded. The projection images at different wavelengths were separated in space and registered independently for further processing, which is of interest for the spectral tomography method. A projective transformation to compensate for the geometric distortions that occur during asymmetric diffraction was used. The first experimental results presented here appear promising.

Investigation of the human pineal gland 3D organization by X-ray phase contrast tomography.

Pineal gland (PG) is a part of the human brain epithalamus that plays an important role in sleep, circadian rhythm, immunity, and reproduction. The calcium deposits and lesions in PG interfere with normal function of the organ and can be associated with different health disorders including serious neurological diseases. At the moment, the detailed mechanisms of PG calcifications and PG lesions formation as well as their involvement in pathological processes are not fully understood. The deep and comprehensive study of the structure of the uncut human PG with histological details, poses a stiff challenge to most imaging techniques, due to low spatial resolution, low visibility or to exceedingly aggressive sample preparation. Here, we investigate the whole uncut and unstained human post-mortem PGs by X-ray phase contrast tomography (XPCT). XPCT is an advanced 3D imaging technique, that permits to study of both soft and calcified tissue of a sample at different scales: from the whole organ to cell structure. In our research we simultaneously resolved 3D structure of parenchyma, vascular network and calcifications. Moreover, we distinguished structural details of intact and degenerated PG tissue. We discriminated calcifications with different structure, pinealocytes nuclei and the glial cells processes. All results were validated by histology. Our research clear demonstrated that XPCT is a potential tool for the high resolution 3D imaging of PG morphological features. This technique opens a new perspective to investigate PG dysfunction and understand the mechanisms of onset and progression of diseases involving the pineal gland.

Dual-energy crystal-analyzer scheme for spectral tomography.

In the present work, a method for adjusting a crystal analyzer to separate two characteristic lines from the spectrum of a conventional X-ray tube for simultaneous registration of tomographic projections is proposed. The experimental implementation of this method using radiation of a molybdenum anode (Kα1, Kß lines) and a silicon Si(111) crystal analyzer in Laue geometry is presented. Projection images at different wavelengths are separated in space and can be recorded independently for further processing. Potential uses of this scheme are briefly discussed.

Soft X-ray diffraction patterns measured by a LiF detector with sub-micrometre resolution and an ultimate dynamic range.

The unique diagnostic possibilities of X-ray diffraction, small X-ray scattering and phase-contrast imaging techniques applied with high-intensity coherent X-ray synchrotron and X-ray free-electron laser radiation can only be fully realized if a sufficient dynamic range and/or spatial resolution of the detector is available. In this work, it is demonstrated that the use of lithium fluoride (LiF) as a photoluminescence (PL) imaging detector allows measuring of an X-ray diffraction image with a dynamic range of ∼107 within the sub-micrometre spatial resolution. At the PETRA III facility, the diffraction pattern created behind a circular aperture with a diameter of 5 µm irradiated by a beam with a photon energy of 500 eV was recorded on a LiF crystal. In the diffraction pattern, the accumulated dose was varied from 1.7 × 105 J cm-3 in the central maximum to 2 × 10-2 J cm-3 in the 16th maximum of diffraction fringes. The period of the last fringe was measured with 0.8 µm width. The PL response of the LiF crystal being used as a detector on the irradiation dose of 500 eV photons was evaluated. For the particular model of laser-scanning confocal microscope Carl Zeiss LSM700, used for the readout of the PL signal, the calibration dependencies on the intensity of photopumping (excitation) radiation (λ = 488 nm) and the gain have been obtained.

High resolution 3D visualization of the spinal cord in a post-mortem murine model.

A crucial issue in the development of therapies to treat pathologies of the central nervous system is represented by the availability of non-invasive methods to study the three-dimensional morphology of spinal cord, with a resolution able to characterize its complex vascular and neuronal organization. X-ray phase contrast micro-tomography enables a high-quality, 3D visualization of both the vascular and neuronal network simultaneously without the need of contrast agents, destructive sample preparations or sectioning. Until now, high resolution investigations of the post-mortem spinal cord in murine models have mostly been performed in spinal cords removed from the spinal canal. We present here post-mortem phase contrast micro-tomography images reconstructed using advanced computational tools to obtain high-resolution and high-contrast 3D images of the fixed spinal cord without removing the bones and preserving the richness of micro-details available when measuring exposed spinal cords. We believe that it represents a significant step toward the in-vivo application.

Reptiles in Space Missions: Results and Perspectives.

Reptiles are a rare model object for space research. However, some reptile species demonstrate effective adaptation to spaceflight conditions. The main scope of this review is a comparative analysis of reptile experimental exposure in weightlessness, demonstrating the advantages and shortcomings of this model. The description of the known reptile experiments using turtles and geckos in the space and parabolic flight experiments is provided. Behavior, skeletal bones (morphology, histology, and X-ray microtomography), internal organs, and the nervous system (morphology, histology, and immunohistochemistry) are studied in the spaceflight experiments to date, while molecular and physiological results are restricted. Therefore, the results are discussed in the scope of molecular data collected from mammalian (mainly rodents) specimens and cell cultures in the parabolic and orbital flights and simulated microgravity. The published data are compared with the results of the gecko model studies after the 12-44.5-day spaceflights with special reference to the unique peculiarities of the gecko model for the orbital experiments. The complex study of thick-toed geckos after three spaceflights, in which all geckos survived and demonstrated effective adaptation to spaceflight conditions, was performed. However, future investigations are needed to study molecular mechanisms of gecko adaptation in space.

Start-to-end simulation of single-particle imaging using ultra-short pulses at the European X-ray Free-Electron Laser.

Single-particle imaging with X-ray free-electron lasers (XFELs) has the potential to provide structural information at atomic resolution for non-crystalline biomolecules. This potential exists because ultra-short intense pulses can produce interpretable diffraction data notwithstanding radiation damage. This paper explores the impact of pulse duration on the interpretability of diffraction data using comprehensive and realistic simulations of an imaging experiment at the European X-ray Free-Electron Laser. It is found that the optimal pulse duration for molecules with a few thousand atoms at 5 keV lies between 3 and 9 fs.

WavePropaGator: interactive framework for X-ray free-electron laser optics design and simulations.

This article describes the WavePropaGator (WPG) package, a new interactive software framework for coherent and partially coherent X-ray wavefront propagation simulations. The package has been developed at European XFEL for users at the existing and emerging free-electron laser (FEL) facilities, as well as at the third-generation synchrotron sources and future diffraction-limited storage rings. The WPG addresses the needs of beamline scientists and user groups to facilitate the design, optimization and improvement of X-ray optics to meet their experimental requirements. The package uses the Synchrotron Radiation Workshop (SRW) C/C++ library and its Python binding for numerical wavefront propagation simulations. The framework runs reliably under Linux, Microsoft Windows 7 and Apple Mac OS X and is distributed under an open-source license. The available tools allow for varying source parameters and optics layouts and visualizing the results interactively. The wavefront history structure can be used for tracking changes in every particular wavefront during propagation. The batch propagation mode enables processing of multiple wavefronts in workflow mode. The paper presents a general description of the package and gives some recent application examples, including modeling of full X-ray FEL beamlines and start-to-end simulation of experiments.

A comprehensive simulation framework for imaging single particles and biomolecules at the European X-ray Free-Electron Laser.

The advent of newer, brighter, and more coherent X-ray sources, such as X-ray Free-Electron Lasers (XFELs), represents a tremendous growth in the potential to apply coherent X-rays to determine the structure of materials from the micron-scale down to the Angstrom-scale. There is a significant need for a multi-physics simulation framework to perform source-to-detector simulations for a single particle imaging experiment, including (i) the multidimensional simulation of the X-ray source; (ii) simulation of the wave-optics propagation of the coherent XFEL beams; (iii) atomistic modelling of photon-material interactions; (iv) simulation of the time-dependent diffraction process, including incoherent scattering; (v) assembling noisy and incomplete diffraction intensities into a three-dimensional data set using the Expansion-Maximisation-Compression (EMC) algorithm and (vi) phase retrieval to obtain structural information. We demonstrate the framework by simulating a single-particle experiment for a nitrogenase iron protein using parameters of the SPB/SFX instrument of the European XFEL. This exercise demonstrably yields interpretable consequences for structure determination that are crucial yet currently unavailable for experiment design.

Monochromatic computed microtomography using laboratory and synchrotron sources and X-ray fluorescence analysis for comprehensive analysis of structural changes in bones.

A combination of X-ray tomography at different wavelengths and X-ray fluorescence analysis was applied in the study of two types of bone tissue changes: prolonged presence in microgravity conditions and age-related bone growth. The proximal tail vertebrae of geckos were selected for investigation because they do not bear the supporting load in locomotion, which allows them to be considered as an independent indicator of gravitational influence. For the vertebrae of geckos no significant differences were revealed in the elemental composition of the flight samples and the synchronous control samples. In addition, the gecko bone tissue samples from the jaw apparatus, spine and shoulder girdle were measured. The dynamics of structural changes in the bone tissue growth was studied using samples of a human fetal hand. The hands of human fetuses of 11-15 weeks were studied. Autonomous zones of calcium accumulation were found not only in individual fingers but in each of the investigated phalanges. The results obtained are discussed.