44 45 D I A M O N D L I G H T S O U R C E A N N U A L R E V I E W 2 0 2 2 / 2 3 D I A M O N D L I G H T S O U R C E A N N U A L R E V I E W 2 0 2 2 / 2 3 Detailedmeasurement of spinal discmechanics provides insight into common back pain Health &Wellbeing – Life Sciences & Biotech The leading cause of years lived with disability is from people suffering with lower back pain. It severely affects the quality of life of all global populations. The spine (intervertebral) discs are the soft tissue in the spinal column. When injured or degenerated, they are responsible for the majority of these lower back pain cases. Currently, only symptomatic treatment or spinal surgery is available; therefore, understanding the mechanical pathogenesis of degeneration is crucial to developing non-invasive therapies. Tissues are developed, maintained, and repaired through the actions of cells using signals from their micromechanical environment, but the ultimate function of large orthopaedic structures such as the spine is mechanical – providing both strength and flexibility to the core of our bodies. This system is elegant but also prone to degradation and injury. The development of better medical approaches to spine problems will continue to be elusive without a proper understanding of tissue mechanics at the level of the cell. Despite a wealth of clinical evidence, we do not yet know whether there is a regional distinction in collagen fibre architecture and whether deformation under mechanical load is region-specific. Therefore, researchers used synchrotron Computed Tomography (sCT) on Diamond’s I13-2 beamline to investigate collagen fibre bundles in 3D throughout an intact native rat lumbar IVD under increasing compressive load. Spinal discs are large structures and must be kept intact to preserve mechanical characteristics. Still, the outer disc material is an intricate collagen fibre composite requiring high-resolution (micron scale) visualisation. Furthermore, unlike the adjacent vertebrae, spinal discs are unmineralised (with no bone tissue), so standard X-ray methods are unsuited. Add to this the need for compression loading to simulate spine function, and a combination of I13-2 beamline capabilities becomes essential: soft tissue imaging (phase contrast), applying load whilst imaging ( in situ tomography), and imaging volumes large enough for animal spine models (millimetres to centimetres). By directly examining the functional response of intact spinal discs, in high enough detail to observe the critical collagen fibres, this study contributed to the understanding of disc degradation and failure. Portions of the disc that fail most frequently (posterior-lateral) have an inherently different fibre architecture from regions less prone to failure (anterior) and have a lower capacity to respond to compressive loading. The level of detail in these observations, with tens of thousands of individual fibres visualised and measured within each region, is essential to a full understanding of disc mechanics. The structure and function patterns revealed in this study suggest a compromise between tissue flexibility and strength characteristics that is important to acknowledge when preventive and treatment therapies for disc pathologies are developed. They also provide a roadmap for attempts to restore spinal tissues to a more functional state. Being able to performdetailed characterisation of numerous individual collagen fibres from in situ imaging of intact tissues and structures will enhance our understanding of tissue biomechanics. This technique will allow us to discover the structures that cause some tissue regions to be more susceptible to degeneration. The study outlines a powerful methodology to evaluate the performance and guide the development of new treatments as they emerge. Related publication: Disney, CM. et al . Regional variations in discrete collagen fibre mechanics within intact intervertebral disc resolved using synchrotron computed tomography and digital volume correlation. Acta Biomaterialia 138 , 361-374 (2022). DOI: 10.1016/j.actbio.2021.10.012 Funding acknowledgement: EPSRC & MRC Centre for Doctoral Training (CDT) Regenerative Medicine (EP/ L014904/1) studentship EPSRC Doctoral Prize Fellowship (EP/R513131/1). UK-MRC ImagingBioPro grant (MR/R025673/1) Corresponding authors: Catherine Disney, Diamond Light Source,
[email protected] Brian Bay, Oregon State University,
[email protected] Imaging andMicroscopy Group Beamline I13-2 A: Three-dimensional imaging of a whole rat spine segment imaged at I13 where soft tissue microstructure could be resolved. Spine segments were compressed during measurements to record structural changes during mechanical function. B: Regional comparisons can be made within the same sample to determine structural-mechanical aspects that could lead to degeneration. C: Tissue biomechanics such as deformation measured as fibre strain and morphological changes was evaluated at three structural levels of region (i), lamella (ii) and individual fibre (iii). (Left column before the sample was compressed, right column after the sample was compressed). High throughput ptychographic tomography at the I13 Coherence Branch Technique Development – Materials Science X-ray Ptychography is an imaging technique that scans a sample through a coherent X-ray beam to collect diffraction information from overlapping regions before reconstructing them into a high- resolution image. This technique produces quantitative phase images with the highest possible spatial resolutions, going well beyond the conventional limitations of the available X-ray optics, and has wide- reaching applications across the physical and life sciences. However, scanning techniques have inherent overheads, with the need to move sample stages and capture motor positions and detector frames. The data collection time for ptychographic tomography data was typically 24 hours, with only a fraction of that time actually exposing the detector. The problem was that the data collection overheads, caused by the time required for motor settling and file writing, were limiting the throughput of the imaging method. As a result, Ptychography has remained significantly slower than direct methods such as Transmission X-ray Microscopy (TXM) and Micro Computed Tomography (micro-CT), which limits its application to the wider scientific community. Diamond scientists have taken steps to reduce the bottlenecks on the I13-1 beamline, making it the fastest ptychography beamline in the world. Faster scans enable the study of more dynamic and in-operando processes, increase the throughput of experiments and offer users real-time feedback. Starting with a hardware scanning protocol created at I24, they began investigating continuous scanning strategies. Once in a continuous scanning modality, the scanning speed relates directly to the data collection rate. The fastest detector available for these measurements could run at up to 9kHz, but the motion controller at the beamline was limited to 800Hz. They investigated novel up-triggering and file-writing strategies that would permit rapid data collection. The experiments and developments were performed and applied to the I13-1 coherence branch. The coherence branch performs multiscale and multimodal ptychographic tomography. This work opens up the powerful imaging capabilities available at the branchline to a much wider scientific community. The approach they developed and implemented allowed for a 20µm 3 volume of battery material to be scanned at the nanoscale in under three hours. The work is continuing and is allowing for samples previously too large (>100µm 3 ) to be scanned in hours and smaller samples to be scanned in minutes. A greater throughput of nanoscale ptychographic tomography allows us to study larger volumes of materials. In addition, statistically relevant data will be recorded. For example, understanding the functionality of brain tissue, requires imaging at the length scale of the synapses (10s nm) and connecting these pathways along several hundreds of microns. Also, studying smaller samples in operando at these length scales is now possible. That is exciting in many applications, for example, tracking the structural evolution within individual primary particles to understand battery degradation as the active material undergoes charging cycles. A major milestone has been achieved by significantly accelerating the recording speed for tomography with ultimate X-ray resolution. As a result, significant numbers of samples can be investigated in three dimensions with resolution on the nanometre length scale. Related publication: Batey, D. et al . High-speed X-ray ptychographic tomography. Scientific Reports 12 , 1-6 (2022). DOI: 10.1038/s41598-022-11292-8 Corresponding author: Darren Batey, Diamond Light Source,
[email protected] 2 kHz ptychographic tomography scan reconstruction. (a) Reconstructed phase projection of NMC particle. (b) 2D slice of the 3D reconstructed volume. (c) 3D view of the reconstructed volume. (d) Fourier shell correlation for the 3D volume (resolution 250 nm). Imaging andMicroscopy Group Beamline I13-1