Regional variations in discrete collagen fibre mechanics within intact intervertebral disc resolved using synchrotron computed tomography and digital volume correlation

Many soft tissues, such as the intervertebral disc (IVD), have a hierarchical fibrous composite structure which suffers from regional damage. We hypothesise that these tissue regions have distinct, inherent fibre structure and structural response upon loading. Here we used synchrotron computed tomography (sCT) to resolve collagen fibre bundles (∼5μm width) in 3D throughout an intact native rat lumbar IVD under increasing compressive load. Using intact samples meant that tissue boundaries (such as endplate-disc or nucleus-annulus) and residual strain were preserved; this is vital for characterising both the inherent structure and structural changes upon loading in tissue regions functioning in a near-native environment. Nano-scale displacement measurements along >10,000 individual fibres were tracked, and fibre orientation, curvature and strain changes were compared between the posterior-lateral region and the anterior region. These methods can be widely applied to other soft tissues, to identify fibre structures which cause tissue regions to be more susceptible to injury and degeneration. Our results demonstrate for the first time that highly-localised changes in fibre orientation, curvature and strain indicate differences in regional strain transfer and mechanical function (e.g. tissue compliance). This included decreased fibre reorientation at higher loads, specific tissue morphology which reduced capacity for flexibility and high strain at the disc-endplate boundary

Towards a mechanistic understanding of particle shrinkage during biomass pyrolysis via synchrotron X-ray microtomography and in-situ radiography

Accurate modelling of particle shrinkage during biomass pyrolysis is key to the production of biochars with specific morphologies. Such biochars represent sustainable solutions to a variety of adsorption-dependent environmental remediation challenges. Modelling of particle shrinkage during biomass pyrolysis has heretofore been based solely on theory and ex-situ experimental data. Here we present the first in-situ phase-contrast X-ray imaging study of biomass pyrolysis. A novel reactor was developed to enable operando synchrotron radiography of fixed beds of pyrolysing biomass. Almond shell particles experienced more bulk shrinkage and less change in porosity than did walnut shell particles during pyrolysis, despite their similar composition. Alkaline pretreatment was found to reduce this difference in feedstock behaviour. Ex-situ synchrotron X-ray microtomography was performed to study the effects of pyrolysis on pore morphology. Pyrolysis led to a redistribution of pores away from particle surfaces, meaning newly formed surface area may be less accessible to adsorbates

Towards a mechanistic understanding of particle shrinkage during biomass pyrolysis via synchrotron X-ray microtomography and in-situ radiography.

Accurate modelling of particle shrinkage during biomass pyrolysis is key to the production of biochars with specific morphologies. Such biochars represent sustainable solutions to a variety of adsorption-dependent environmental remediation challenges. Modelling of particle shrinkage during biomass pyrolysis has heretofore been based solely on theory and ex-situ experimental data. Here we present the first in-situ phase-contrast X-ray imaging study of biomass pyrolysis. A novel reactor was developed to enable operando synchrotron radiography of fixed beds of pyrolysing biomass. Almond shell particles experienced more bulk shrinkage and less change in porosity than did walnut shell particles during pyrolysis, despite their similar composition. Alkaline pretreatment was found to reduce this difference in feedstock behaviour. Ex-situ synchrotron X-ray microtomography was performed to study the effects of pyrolysis on pore morphology. Pyrolysis led to a redistribution of pores away from particle surfaces, meaning newly formed surface area may be less accessible to adsorbates

Operando and High-throughput multicscale-tomography

We report about multiscale tomography with high throughput at the Diamond beamline I13L. The beamline has the purpose of multi-scale and operando imaging and consists of two independent branchlines operating in real and reciprocal space. The imaging branch -called Diamond-Manchester branchline- hosts micro-tomography, grating interferometry and a full-field microscope. For rapid recording a broad spectrum of the undulator radiation is used either with band-passing the light with a combination of a filter and a deflecting mirror or using a multilayer monochromator. For all the methods similar recording times can be achieved, with typical scanning times of some minutes and covering the resolution range from microns to the 100nm range. Most recently a robot arm has been installed to increase the throughput to 300 samples per day. The system is now implemented for user operation in remote operation mode for the micro-tomography setup and can be expanded to the two other experiments. The instrumental capabilities are applied on various topics such as the study of biodiversity of insects or the structural variations of electrode materials in batteries. Fast recording with dedicated sample environments (not using the sample changing robot) enables operando studies in many areas, the charging/discharging cycles on batteries, the degradation of teeth enamel under various conditions or loading brine sandstone mixtures with CO2, to name some examples. For imaging with highest spatial resolution we managed to improve significantly the recording speed of ptycho-tomography, which is now in the order of hours and will be reduced further. We demonstrated in the past 2-D recording with 10kHz and expand the instrumental capability with specific hardware dependent triggering and scanning schemes. We expand the research program for multi-scale imaging across both branchlines (imaging and coherence branchlines) with first studies such as batteries, brain research, concrete

Preservation of bone tissue integrity with temperature control for in situ SR-MicroCT experiments

© 2018 by the authors. Digital volume correlation (DVC), combined with in situ synchrotron microcomputed tomography (SR-microCT) mechanics, allows for 3D full-field strain measurement in bone at the tissue level. However, long exposures to SR radiation are known to induce bone damage, and reliable experimental protocols able to preserve tissue properties are still lacking. This study aims to propose a proof-of-concept methodology to retain bone tissue integrity, based on residual strain determination using DVC, by decreasing the environmental temperature during in situ SR-microCT testing. Compact and trabecular bone specimens underwent five consecutive full tomographic data collections either at room temperature or 0 °C. Lowering the temperature seemed to reduce microdamage in trabecular bone but had minimal effect on compact bone. A consistent temperature gradient was measured at each exposure period, and its prolonged effect over time may induce localised collagen denaturation and subsequent damage. DVC provided useful information on irradiation-induced microcrack initiation and propagation. Future work is necessary to apply these findings to in situ SR-microCT mechanical tests, and to establish protocols aiming to minimise the SR irradiation-induced damage of bone

Four-dimensional imaging and quantification of viscous flow sintering within a 3D printed bioactive glass scaffold using synchrotron X-ray tomography

Bioglass® was the first material to form a stable chemical bond with human tissue. Since its discovery, a key goal was to produce three-dimensional (3D) porous scaffolds which can host and guide tissue repair, in particular, regeneration of long bone defects resulting from trauma or disease. Producing 3D scaffolds from bioactive glasses is challenging because of crystallization events that occur while the glass particles densify at high temperatures. Bioactive glasses such as the 13–93 composition can be sintered by viscous flow sintering at temperatures above the glass transition onset (T_{g}) and below the crystallization temperature (T_{c}). There is, however, very little literature on viscous flow sintering of bioactive glasses, and none of which focuses on the viscous flow sintering of glass scaffolds in four dimensions (4D) (3D + time). Here, high-resolution synchrotron-sourced X-ray computed tomography (sCT) was used to capture and quantify viscous flow sintering of an additively manufactured bioactive glass scaffold in 4D. In situ sCT allowed the simultaneous quantification of individual particle (local) structural changes and the scaffold's (global) dimensional changes during the sintering cycle. Densification, calculated as change in surface area, occurred in three distinct stages, confirming classical sintering theory. Importantly, our observations show for the first time that the local and global contributions to densification are significantly different at each of these stages: local sintering dominates stages 1 and 2, while global sintering is more prevalent in stage 3. During stage 1, small particles coalesced to larger particles because of their higher driving force for viscous flow at lower temperatures, while large angular particles became less faceted (angular regions had a local small radius of curvature). A transition in the rate of sintering was then observed in which significant viscous flow occurred, resulting in large reduction of surface area, total strut volume, and interparticle porosity because the majority of the printed particles coalesced to become continuous struts (stage 2). Transition from stage 2 to stage 3 was distinctly obvious when interparticle pores became isolated and closed, while the sintering rate significantly reduced. During stage 3, at the local scale, isolated pores either became more spherical or reduced in size and disappeared depending on their initial morphology. During stage 3, sintering of the scaffolds continued at the strut level, with interstrut porosity reducing, while globally the strut diameter increased in size, suggesting overall shrinkage of the scaffold with the flow of material via the strut contacts. This study provides novel insights into viscous flow in a complex non-idealized construct, where, locally, particles are not spherical and are of a range of sizes, leading to a random distribution of interparticle porosity, while globally, predesigned porosity between the struts exists to allow the construct to support tissue growth. This is the first time that the three stages of densification have been captured at the local and global scales simultaneously. The insights provided here should accelerate the development of 3D bioactive glass scaffolds

Effect of SR-microCT radiation on the mechanical integrity of trabecular bone using in situ mechanical testing and digital volume correlation

The use of synchrotron radiation micro-computed tomography (SR-microCT) is becoming increasingly popular for studying the relationship between microstructure and bone mechanics subjected to in situ mechanical testing. However, it is well known that the effect of SR X-ray radiation can considerably alter the mechanical properties of bone tissue. Digital volume correlation (DVC) has been extensively used to compute full-field strain distributions in bone specimens subjected to step-wise mechanical loading, but tissue damage from sequential SR-microCT scans has not been previously addressed. Therefore, the aim of this study is to examine the influence of SR irradiation-induced microdamage on the apparent elastic properties of trabecular bone using DVC applied to in situ SR-microCT tomograms obtained with different exposure times. Results showed how DVC was able to identify high local strain levels (> 10,000 µε) corresponding to visible microcracks at high irradiation doses (~ 230 kGy), despite the apparent elastic properties remained unaltered. Microcracks were not detected and bone plasticity was preserved for low irradiation doses (~ 33 kGy), although image quality and consequently, DVC performance were reduced. DVC results suggested some local deterioration of tissue that might have resulted from mechanical strain concentration further enhanced by some level of local irradiation even for low accumulated dose

Synchrotron tomography of intervertebral disc deformation quantified by digital volume correlation reveals microstructural influence on strain patterns

The intervertebral disc (IVD) has a complex and multiscale extracellular matrix structure which provides unique mechanical properties to withstand physiological loading. Low back pain has been linked to degeneration of the disc but reparative treatments are not currently available. Characterising the disc’s 3D microstructure and its response in a physiologically relevant loading environment is required to improve understanding of degeneration and to develop new reparative treatments. In this study, techniques for imaging the native IVD, measuring internal deformation and mapping volumetric strain were applied to an in situ compressed ex vivo rat lumbar spine segment. Synchrotron X-ray micro-tomography (synchrotron CT) was used to resolve IVD structures at microscale resolution. These image data enabled 3D quantification of collagen bundle orientation and measurement of local displacement in the annulus fibrosus between sequential scans using digital volume correlation (DVC). The volumetric strain mapped from synchrotron CT provided a detailed insight into the micromechanics of native IVD tissue. The DVC findings showed that there was no slipping at lamella boundaries, and local strain patterns were of a similar distribution to the previously reported elastic network with some heterogeneous areas and maximum strain direction aligned with bundle orientation, suggesting bundle stretching and sliding. This method has the potential to bridge the gap between measures of macro-mechanical properties and the local 3D micro-mechanical environment experienced by cells. This is the first evaluation of strain at the micro scale level in the intact IVD and provides a quantitative framework for future IVD degeneration mechanics studies and testing of tissue engineered IVD replacements

Fast Multi-scale imaging using the Beamline I13L at the Diamond Light Source

The DIAMOND beamline I13L is dedicated to multi-scale and multi-modal imaging in real and reciprocal space. The beamline consists of two independently operating experimental stations, located at a distance of more than 200 m from the source. The Imaging Branch performs micro-tomography with in-line phase contrast in the 6-30 keV energy range. In addition, a grating interferometry setup and a full-field microscope for nano-tomography are currently implemented. Other techniques providing high-resolution three-dimensional information, in particular coherent X-ray diffraction, are hosted on the Coherence Branch. All imaging methods are tested to operate with large energy bandwidths and therefore shorter exposure times. To this end, two options are currently used: the so-called ‘pink-beam’ mode using a reflecting mirror and X-ray filters and monochromatic mode using a multilayer monochromator. The operation mode enables science for in-situ and operando studies across a wide range of scientific areas