Phase-contrast neutron imaging compared with wave propagation and McStas simulations.

Propagation-based phase contrast, for example in the form of edge enhancement contrast, is well established within X-ray imaging but is not widely used in neutron imaging. This technique can help increase the contrast of low-attenuation samples but may confuse quantitative absorption measurements. Therefore, it is important to understand the experimental parameters that cause and amplify or dampen this effect in order to optimize future experiments properly. Two simulation approaches have been investigated, a wave-based simulation and a particle-based simulation conducted in McStas [Willendrup & Lefmann (2020). J. Neutron Res. 22, 1-16], and they are compared with experimental data. The experiment was done on a sample of metal foils with weakly and strongly neutron absorbing layers, which were measured while varying the rotation angle and propagation distance from the sample. The experimental data show multiple signals: attenuation, phase contrast and reflection. The wave model reproduces the sample attenuation and the phase peaks but it does not reproduce the behavior of these peaks as a function of rotation angle. The McStas simulation agrees better with the experimental data, as it reproduces attenuation, phase peaks and reflection, as well as the change in these signals as a function of rotation angle and distance. This suggests that the McStas simulation approach, where the particle description of the neutron facilitates the incorporation of multiple effects, is the most convenient way of modeling edge enhancement in neutron imaging.

Correlative study of liquid in human bone by 3D neutron microscopy and lab-based X-ray µCT.

Liquid plays an important role in bone that has a complex 3D hierarchical pore structure. However, liquid (water) is difficult to discern from e.g. an organic matrix by X-ray imaging. Therefore, we use a correlative approach using both high resolution X-ray and neutron imaging. Human femoral bone with liquid adsorbed into some of the pores was imaged with both the Neutron Microscope at the ICON beamline, SINQ at PSI, and by lab-based µCT using 2.7 µm voxel size. Segmentation of the two datasets showed that, even though the liquid was clearly distinguishable in the neutron data and not in the X-ray data, it remained challenging to segment it from bone due to overlaps of peaks in the gray level histograms. In consequence, segmentations from X-ray and neutron data varied significantly. To address this issue, the segmented X-ray porosities was overlaid on the neutron data, making it possible to localize the liquid in the vascular porosities of the bone sample and use the neutron attenuation to identify it as H2O. The contrast in the neutron images was lowered slightly between the bone and the liquid compared to the bone and the air. This correlative study shows that the complementary use of X-rays and neutrons is very favorable, since H2O is very distinct in the neutron data, while D2O, H2O, and organic matter can barely be distinguished from air in the X-ray data.

Polychromatic neutron phase-contrast imaging of weakly absorbing samples enabled by phase retrieval.

The use of a phase-retrieval technique for propagation-based phase-contrast neutron imaging with a polychromatic beam is demonstrated. This enables imaging of samples with low absorption contrast and/or improving the signal-to-noise ratio to facilitate e.g. time-resolved measurements. A metal sample, designed to be close to a phase pure object, and a bone sample with canals partially filled with D2O were used for demonstrating the technique. These samples were imaged with a polychromatic neutron beam followed by phase retrieval. For both samples the signal-to-noise ratios were significantly improved and, in the case of the bone sample, the phase retrieval allowed for separation of bone and D2O, which is important for example for in situ flow experiments. The use of deuteration contrast avoids the use of chemical contrast enhancement and makes neutron imaging an interesting complementary method to X-ray imaging of bone.

Opportunities for biomineralization research using multiscale computed X-ray tomography as exemplified by bone imaging.

Biominerals typically have complex hierarchical structures traversing many length scales. This makes their structural characterization complicated, since it requires 3D techniques that can probe full specimens at down to nanometer-resolution, a combination that is difficult - if not impossible - to achieve simultaneously. One challenging example is bone, a mineralized tissue with a highly complex architecture that is replete with a network of cells. X-ray computed tomography techniques enable multiscale structural characterization through the combination of various equipment and emerge as promising tools for characterizing biominerals. Using bone as an example, we discuss how combining different X-ray imaging instruments allow characterizing bone structures from the nano- to the organ-scale. In particular, we compare and contrast human and rodent bone, emphasize the importance of the osteocyte lacuno-canalicular network in bone, and finally illustrate how combining synchrotron X-ray imaging with laboratory instrumentation for computed tomography is especially helpful for multiscale characterization of biominerals.

Genetic Ablation of Osteopontin in Osteomalacic Hyp Mice Partially Rescues the Deficient Mineralization Without Correcting Hypophosphatemia.

PHEX is predominantly expressed by bone and tooth-forming cells, and its inactivating mutations in X-linked hypophosphatemia (XLH) lead to renal phosphate wasting and severe hypomineralization of bones and teeth. Also present in XLH are hallmark hypomineralized periosteocytic lesions (POLs, halos) that persist despite stable correction of serum phosphate (Pi ) that improves bulk bone mineralization. In XLH, mineralization-inhibiting osteopontin (OPN, a substrate for PHEX) accumulates in the extracellular matrix of bone. To investigate how OPN functions in Hyp mice (a model for XLH), double-null (Hyp;Opn-/- ) mice were generated. Undecalcified histomorphometry performed on lumbar vertebrae revealed that Hyp;Opn-/- mice had significantly reduced osteoid area/bone area (OV/BV) and osteoid thickness of trabecular bone as compared to Hyp mice, despite being as hypophosphatemic as Hyp littermate controls. However, tibias examined by synchrotron radiation micro-CT showed that mineral lacunar volumes remained abnormally enlarged in these double-null mice. When Hyp;Opn-/- mice were fed a high-Pi diet, serum Pi concentration increased, and OV/BV and osteoid thickness normalized, yet mineral lacunar area remained abnormally enlarged. Enpp1 and Ankh gene expression were increased in double-null mice fed a high-Pi diet, potentially indicating a role for elevated inhibitory pyrophosphate (PPi ) in the absence of OPN. To further investigate the persistence of POLs in Hyp mice despite stable correction of serum Pi , immunohistochemistry for OPN on Hyp mice fed a high-Pi diet showed elevated OPN in the osteocyte pericellular lacunar matrix as compared to Hyp mice fed a control diet. This suggests that POLs persisting in Hyp mice despite correction of serum Pi may be attributable to the well-known upregulation of mineralization-inhibiting OPN by Pi , and its accumulation in the osteocyte pericellular matrix. This study shows that OPN contributes to osteomalacia in Hyp mice, and that genetic ablation of OPN in Hyp mice improves the mineralization phenotype independent of systemic Pi -regulating factors. © 2020 American Society for Bone and Mineral Research.

Bone Biomineral Properties Vary across Human Osteonal Bone.

The biomineralization of bone remains a puzzle. During Haversian remodeling in the dense human cortical bone, osteoclasts excavate a tunnel that is then filled in by osteoblasts with layers of bone of varying fibril orientations, resulting in a lamellar motif. Such bone represents an excellent possibility to increase our understanding of bone as a material as well as bone biomineralization by studying spatio/temporal variations in the biomineral across an osteon. To this end, fluorescence computed tomography and diffraction scattering computed tomography with sub-micrometer resolution is applied to obtain position resolved fluorescence spectra and diffraction patterns in a 3D volume. The microstructural properties of the apatite biomineral are not homogeneous but depend critically on the time point at which it was laid down. This indicates that the nature of bone biomineral is highly dependent on the microenvironment during bone formation and remodeling.