Endostructural and periosteal growth of the human humerus.

The growth and development of long bones are of considerable interests in the fields of comparative anatomy and palaeoanthropology, as evolutionary changes and adaptations to specific physical activity patterns are expected to be revealed during bone ontogeny. Traditionally, the cross-sectional geometry of long bones has been examined at discrete locations usually placed at set intervals or fixed percentage distances along the midline axis of the bone shaft. More recently, the technique of morphometric mapping has enabled the continuous analysis of shape variation along the shaft. Here we extend this technique to the full sequence of late fetal and postnatal development of the humeral shaft in a modern human population sample, with the aim of establishing the shape changes during growth and their relationship with the development of the arm musculature and activity patterns. A sample of modern human humeri from individuals of age ranging from 24 weeks in utero to 18 years was imaged using microtomography at multiple resolutions and custom Matlab scripts. Standard biomechanical properties, cortical thickness, surface curvature, and pseudo-landmarks were extracted along radial vectors spaced at intervals of 1° at each 0.5% longitudinal increment measured along the shaft axis. Heat maps were also generated for cortical thickness and surface curvature. The results demonstrate that a whole bone approach to analysis of cross-sectional geometry is more desirable where possible, as there is a continuous pattern of variation along the shaft. It is also possible to discriminate very young individuals and adolescents from other groups by relative cortical thickness, and also by periosteal surface curvature.

Fluid/Fiber Interactions and the Conductivity of Inkjet Printed Ag on Textile Substrates.

X-ray tomographic reconstruction reveals that the distribution of Ag after inkjet printing and sintering a nanoparticle conducting ink on a woven polyester textile substrate is strongly controlled by the fiber surface properties and fabric architecture. Capillarity confines the transport of the ink predominantly within the warp or weft yarns of the fabric and there is little transport of ink between the yarns. Changing the fiber surface energy through the Scotchgard treatment leads to an increase in the contact angle, reducing ink transport along the fibers and an increase in conductance. A similar effect is seen when printed drop spacing is reduced, increasing the local Ag concentration. Electrical conductivity is strongly influenced by the fiber density in each yarn direction and, in this case, the different densities lead to different electrical conductance values. Through the use of image segmentation, it is possible to identify a low level of electrical interconnection between the warp and weft yarns. Conductance within a yarn is shown to depend on Ag concentration via a percolation mechanism and this is confirmed by a simple model relating the volume of the largest interconnected Ag object present to the measured conductance. These results illustrate the complexity of the interaction between conductive inks and fibrous substrates and that concepts, such as sheet resistance, used to characterize printed conductors on solid substrates are not applicable to textile substrates.

Refinement of pore size at sub-angstrom precision in robust metal-organic frameworks for separation of xylenes.

The demand for xylenes is projected to increase over the coming decades. The separation of xylene isomers, particularly p- and m-xylenes, is vital for the production of numerous polymers and materials. However, current state-of-the-art separation is based upon fractional crystallisation at 220 K which is highly energy intensive. Here, we report the discrimination of xylene isomers via refinement of the pore size in a series of porous metal-organic frameworks, MFM-300, at sub-angstrom precision leading to the optimal kinetic separation of all three xylene isomers at room temperature. The exceptional performance of MFM-300 for xylene separation is confirmed by dynamic ternary breakthrough experiments. In-depth structural and vibrational investigations using synchrotron X-ray diffraction and terahertz spectroscopy define the underlying host-guest interactions that give rise to the observed selectivity (p-xylene < o-xylene < m-xylene) and separation factors of 4.6-18 for p- and m-xylenes.

3D Imaging of Indentation Damage in Bone.

Bone is a complex material comprising high stiffness, but brittle, crystalline bio-apatite combined with compliant, but tough, collagen fibres. It can accommodate significant deformation, and the bone microstructure inhibits crack propagation such that micro-cracks can be quickly repaired. Catastrophic failure (bone fracture) is a major cause of morbidity, particularly in aging populations, either through a succession of small fractures or because a traumatic event is sufficiently large to overcome the individual crack blunting/shielding mechanisms. Indentation methods provide a convenient way of characterising the mechanical properties of bone. It is important to be able to visualise the interactions between the bone microstructure and the damage events in three dimensions (3D) to better understand the nature of the damage processes that occur in bone and the relevance of indentation tests in evaluating bone resilience and strength. For the first time, time-lapse laboratory X-ray computed tomography (CT) has been used to establish a time-evolving picture of bone deformation/plasticity and cracking. The sites of both crack initiation and termination as well as the interconnectivity of cracks and pores have been visualised and identified in 2D and 3D.

Four-Dimensional Imaging of Soft Tissue and Implanted Biomaterial Mechanics: A Barbed Suture Case Study for Tendon Repair.

Timely, recent developments in X-ray microcomputed tomography (XµCT) imaging such as increased resolution and improved sample preparation enable nondestructive time-lapse imaging of polymeric biomaterials when implanted in soft tissue, which we demonstrate herein. Imaging the full three-dimensional (3D) structure of an implanted biomaterial provides new opportunities to assess the micromechanics of the interface between the implant and tissues and how this changes over time as force is applied in load-bearing musculoskeletal applications. In this paper, we present a case study demonstrating in situ XµCT and finite element analysis, using a dynamically loaded barbed suture repair for its novel use in tendon tissue. The aim of this study was to identify the distribution of stress in the suture and tendon as load is applied. The data gained demonstrate a clear 3D visualization of microscale features in both the tissue and implant in wet conditions. XµCT imaging has revealed, for the first time, pores around the suture, preventing full engagement of all the barbs with the tendon tissue. Subsequent finite element analysis reveals the localized stress and strain, which are not evenly distributed along the suture, or throughout the tissue. This case study demonstrates for the first time a powerful in situ mechanical imaging tool, which could be readily adapted by other laboratories to interrogate and optimize the interface between the implanted biomaterials and the soft tissue.

Morphological Substrates for Atrial Arrhythmogenesis in a Heart With Atrioventricular Septal Defect.

Due to advances in corrective surgery, congenital heart disease has an ever growing patient population. Atrial arrhythmias are frequently observed pre- and post-surgical correction. Pharmaceutical antiarrhythmic therapy is not always effective, therefore many symptomatic patients undergo catheter ablation therapy. In patients with atrioventricular septal defects (AVSD), ablation therapy itself has mixed success; arrhythmogenic recurrences are common, and because of the anatomical displacement of the atrioventricular node, 3-degree heart block post-ablation is a real concern. In order to develop optimal and safe ablation strategies, the field of congenital cardiac electrophysiology must combine knowledge from clinical electrophysiology with a thorough understanding of the anatomical substrates for arrhythmias. Using image-based analysis and multi-cellular mathematical modeling of electrical activation, we show how the anatomical alterations characteristic of an AVSD serve as arrhythmogenic substrates. Using ex-vivo contrast enhanced micro-computed tomography we imaged post-mortem the heart of a 5 month old male with AVSD at an isometric spatial resolution of 38 µm. Morphological analysis revealed the 3D disposition of the cardiac conduction system for the first time in an intact heart with this human congenital malformation. We observed displacement of the compact atrioventricular node inferiorly to the ostium of the coronary sinus. Myocyte orientation analysis revealed that the normal arrangement of the major atrial muscle bundles was preserved but was modified in the septal region. Models of electrical activation suggest the disposition of the myocytes within the atrial muscle bundles associated with the "fast pathway," together with the displaced atrioventricular node, serve as potential substrates for re-entry and possibly atrial fibrillation. This study used archived human hearts, showing them to be a valuable resource for the mathematical modeling community, and opening new possibilities for the investigations of arrhythmogenesis and ablation strategies in the congenitally malformed heart.

A study of the progression of damage in an axially loaded Branta leucopsis femur using X-ray computed tomography and digital image correlation.

This paper uses X-ray computed tomography to track the mechanical response of a vertebrate (Barnacle goose) long bone subjected to an axial compressive load, which is increased gradually until failure. A loading rig was mounted in an X-ray computed tomography system so that a time-lapse sequence of three-dimensional (3D) images of the bone's internal (cancellous or trabecular) structure could be recorded during loading. Five distinct types of deformation mechanism were observed in the cancellous part of the bone. These were (i) cracking, (ii) thinning (iii) tearing of cell walls and struts, (iv) notch formation, (v) necking and (vi) buckling. The results highlight that bone experiences brittle (notch formation and cracking), ductile (thinning, tearing and necking) and elastic (buckling) modes of deformation. Progressive deformation, leading to cracking was studied in detail using digital image correlation. The resulting strain maps were consistent with mechanisms occurring at a finer-length scale. This paper is the first to capture time-lapse 3D images of a whole long bone subject to loading until failure. The results serve as a unique reference for researchers interested in how bone responds to loading. For those using computer modelling, the study not only provides qualitative information for verification and validation of their simulations but also highlights that constitutive models for bone need to take into account a number of different deformation mechanisms.

Towards understanding the influence of porosity on mechanical and fracture behaviour of quasi-brittle materials: experiments and modelling.

In this work, porosity-property relationships of quasi-brittle materials are explored through a combined experimental and numerical approach. In the experimental part, hemihyrate gypsum plaster powder ( CaSO 4 · 1 / 2 H 2 O ) and expanded spherical polystyrene beads (1.5-2.0 mm dia.) have been mixed to form a model material with controlled additions of porosity. The expanded polystyrene beads represent pores within the bulk due to their light weight and low strength compared with plaster. Varying the addition of infill allows the production of a material with different percentages of porosity: 0, 10, 20, 30 and 31 vol%. The size and location of these pores have been characterised by 3D X-ray computed tomography. Beams of the size of 20 × 20 × 150  mm were cast and loaded under four-point bending to obtain the mechanical characteristics of each porosity level. The elastic modulus and flexural strength are found to decrease with increased porosity. Fractography studies have been undertaken to identify the role of the pores on the fracture path. Based on the known porosity, a 3D model of each microstructure has been built and the deformation and fracture was computed using a lattice-based multi-scale finite element model. This model predicted similar trends as the experimental results and was able to quantify the fractured sites. The results from this model material experimental data and the lattice model predictions are discussed with respect to the role of porosity on the deformation and fracture of quasi-brittle materials.

Optimal Contrast Agent Staining of Ligaments and Tendons for X-Ray Computed Tomography.

X-ray computed tomography has become an important tool for studying the microstructures of biological soft tissues, such as ligaments and tendons. Due to the low X-ray attenuation of such tissues, chemical contrast agents are often necessary to enhance contrast during scanning. In this article, the effects of using three different contrast agents--iodine potassium iodide solution, phosphotungstic acid and phosphomolybdic acid--are evaluated and compared. Porcine anterior cruciate ligaments, patellar tendons, medial collateral ligaments and lateral collateral ligaments were used as the basis of the study. Three samples of each of the four ligament/tendon types were each assigned a different contrast agent (giving a total of twelve samples), and the progression of that agent through the tissue was monitored by performing a scan every day for a total period of five days (giving a total of sixty scans). Since the samples were unstained on day one, they had been stained for a total of four days by the time of the final scans. The relative contrast enhancement and tissue deformation were measured. It was observed that the iodine potassium iodide solution penetrated the samples fastest and caused the least sample shrinkage on average (although significant deformation was observed by the time of the final scans), whereas the phosphomolybdic acid caused the greatest sample shrinkage. Equations describing the observed behaviour of the contrast agents, which can be used to predict optimal staining times for ligament and tendon X-ray computed tomography, are presented.

Congestive Heart Failure Leads to Prolongation of the PR Interval and Atrioventricular Junction Enlargement and Ion Channel Remodelling in the Rabbit.

Heart failure is a major killer worldwide. Atrioventricular conduction block is common in heart failure; it is associated with worse outcomes and can lead to syncope and bradycardic death. We examine the effect of heart failure on anatomical and ion channel remodelling in the rabbit atrioventricular junction (AVJ). Heart failure was induced in New Zealand rabbits by disruption of the aortic valve and banding of the abdominal aorta resulting in volume and pressure overload. Laser micro-dissection and real-time polymerase chain reaction (RT-PCR) were employed to investigate the effects of heart failure on ion channel remodelling in four regions of the rabbit AVJ and in septal tissues. Investigation of the AVJ anatomy was performed using micro-computed tomography (micro-CT). Heart failure animals developed first degree heart block. Heart failure caused ventricular myocardial volume increase with a 35% elongation of the AVJ. There was downregulation of HCN1 and Cx43 mRNA transcripts across all regions and downregulation of Cav1.3 in the transitional tissue. Cx40 mRNA was significantly downregulated in the atrial septum and AVJ tissues but not in the ventricular septum. mRNA abundance for ANP, CLCN2 and Navß1 was increased with heart failure; Nav1.1 was increased in the inferior nodal extension/compact node area. Heart failure in the rabbit leads to prolongation of the PR interval and this is accompanied by downregulation of HCN1, Cav1.3, Cx40 and Cx43 mRNAs and anatomical enlargement of the entire heart and AVJ.

Interpreting pathologies in extant and extinct archosaurs using micro-CT.

Palaeopathology offers unique insight to the healing strategies of extinct organisms, permitting questions concerning bone physiology to be answered in greater depth. Unfortunately, most palaeopathological studies are confined to external morphological interpretations due to the destructive nature of traditional methods of study. This limits the degree of reliable diagnosis and interpretation possible. X-ray MicroTomography (micro-CT, XMT) provides a non-destructive means of analysing the internal three-dimensional structure of pathologies in both extant and extinct individuals, at higher resolutions than possible with medical scanners. In this study, we present external and internal descriptions of pathologies in extant and extinct archosaurs using XMT. This work demonstrates that the combination of external/internal diagnosis that X-ray microtomography facilitates is crucial when differentiating between pathological conditions. Furthermore, we show that the use of comparative species, both through direct analysis and from the literature, provides key information for diagnosing between vertebrate groups in the typical pathological conditions and physiological processes. Micro-CT imaging, combined with comparative observations of extant species, provides more detailed and reliable interpretation of palaeopathologies. Micro-CT is an increasingly accessible tool, which will provide key insights for correctly interpreting vertebrate pathologies in the future.

X-ray computed tomography of the anterior cruciate ligament and patellar tendon.

The effect of phosphotungstic acid (PTA) and iodine solution (IKI) staining was investigated as a method of enhancing contrast in the X-ray computed tomography of porcine anterior cruciate ligaments (ACL) and patellar tendons (PT). We show that PTA enhanced surface contrast, but was ineffective at penetrating samples, whereas IKI penetrated more effectively and enhanced contrast after 70 hours of staining. Contrast enhancement was compared when using laboratory and synchrotron based X-ray sources. Using the laboratory source, PT fascicles were tracked and their alignment was measured. Individual ACL fascicles could not be identified, but identifiable features were evident that were tracked. Higher resolution scans of fascicle bundles from the PT and ACL were obtained using synchrotron imaging techniques. These scans exhibited greater contrast between the fascicles and matrix in the PT sample, facilitating the identification of the fascicle edges; however, it was still not possible to detect individual fascicles in the ACL.

Optimal iodine staining of cardiac tissue for X-ray computed tomography.

X-ray computed tomography (XCT) has been shown to be an effective imaging technique for a variety of materials. Due to the relatively low differential attenuation of X-rays in biological tissue, a high density contrast agent is often required to obtain optimal contrast. The contrast agent, iodine potassium iodide ([Formula: see text]), has been used in several biological studies to augment the use of XCT scanning. Recently I2KI was used in XCT scans of animal hearts to study cardiac structure and to generate 3D anatomical computer models. However, to date there has been no thorough study into the optimal use of I2KI as a contrast agent in cardiac muscle with respect to the staining times required, which has been shown to impact significantly upon the quality of results. In this study we address this issue by systematically scanning samples at various stages of the staining process. To achieve this, mouse hearts were stained for up to 58 hours and scanned at regular intervals of 6-7 hours throughout this process. Optimal staining was found to depend upon the thickness of the tissue; a simple empirical exponential relationship was derived to allow calculation of the required staining time for cardiac samples of an arbitrary size.

Micro-CT evaluation of the effectiveness of the combined use of rotary and hand instrumentation in removal of Resilon.

This study compares the effectiveness of ProTaper rotary files with ProTaper retreatment and K-files in the removal of Resilon or gutta percha (GP) from canals filled either by cold lateral condensation or thermal obturation using micro-CT. Ninety-six teeth were prepared using ProTaper files and allocated into four groups (n=24): Group-1 was filled with GP/AH-Plus and Group-2 with Resilon/RealSeal using cold lateral condensation. Group-3 was filled with GP/AH-Plus and Group-4 with Resilon/RealSeal using System B and Obtura II. The roots were scanned by micro-CT. Each group was divided into two subgroups (n=12): A, retreated using ProTaper files and B, using ProTaper retreatment and K-files. The roots were scanned to calculate the volume of the remaining material. With thermal obturation, roots filled with Resilon had significantly more remaining material than GP. Obturation using thermal technique resulted in significantly less remaining material than cold condensation except Resilon retreated using ProTaper retreatment and K-files.

Metamorphosis revealed: time-lapse three-dimensional imaging inside a living chrysalis.

Studies of model insects have greatly increased our understanding of animal development. Yet, they are limited in scope to this small pool of model species: a small number of representatives for a hyperdiverse group with highly varied developmental processes. One factor behind this narrow scope is the challenging nature of traditional methods of study, such as histology and dissection, which can preclude quantitative analysis and do not allow the development of a single individual to be followed. Here, we use high-resolution X-ray computed tomography (CT) to overcome these issues, and three-dimensionally image numerous lepidopteran pupae throughout their development. The resulting models are presented in the electronic supplementary material, as are figures and videos, documenting a single individual throughout development. They provide new insight and details of lepidopteran metamorphosis, and allow the measurement of tracheal and gut volume. Furthermore, this study demonstrates early and rapid development of the tracheae, which become visible in scans just 12 h after pupation. This suggests that there is less remodelling of the tracheal system than previously expected, and is methodologically important because the tracheal system is an often-understudied character system in development. In the future, this form of time-lapse CT-scanning could allow faster and more detailed developmental studies on a wider range of taxa than is presently possible.

Application of micro-computed tomography with iodine staining to cardiac imaging, segmentation, and computational model development.

Micro-computed tomography (micro-CT) has been widely used to generate high-resolution 3-D tissue images from small animals nondestructively, especially for mineralized skeletal tissues. However, its application to the analysis of soft cardiovascular tissues has been limited by poor inter-tissue contrast. Recent ex vivo studies have shown that contrast between muscular and connective tissue in micro-CT images can be enhanced by staining with iodine. In the present study, we apply this novel technique for imaging of cardiovascular structures in canine hearts. We optimize the method to obtain high-resolution X-ray micro-CT images of the canine atria and its distinctive regions-including the Bachmann's bundle, atrioventricular node, pulmonary arteries and veins-with clear inter-tissue contrast. The imaging results are used to reconstruct and segment the detailed 3-D geometry of the atria. Structure tensor analysis shows that the arrangement of atrial fibers can also be characterized using the enhanced micro-CT images, as iodine preferentially accumulates within the muscular fibers rather than in connective tissues. This novel technique can be particularly useful in nondestructive imaging of 3-D cardiac architectures from large animals and humans, due to the combination of relatively high speed ( ~ 1 h/per scan of the large canine heart) and high voxel resolution (36 µm) provided. In summary, contrast micro-CT facilitates fast and nondestructive imaging and segmenting of detailed 3-D cardiovascular geometries, as well as measuring fiber orientation, which are crucial in constructing biophysically detailed computational cardiac models.