Mechanics of cortical bone: exploring the micro- and nano-scale

This PhD aimed at characterising mechanical, structural and compositional properties of cortical bone at the micro- and nano-scale employing various techniques applied to mouse models of ageing and disease. Nanoindentation was used to probe bone at the micro-scale. Nanoindentation properties of the same control mouse bones were measured using a range of preparation, testing and analysis options. This was the first time that such systematic study was conducted. It was concluded that although the measured values were influenced by experimental and analysis choices, nanoindentation was capable of capturing relative trends in bone’s mechanical properties. Nanoindentation was then coupled to a poroelastic approach to measure age-related changes in mouse bone permeability. Permeability is key to understanding fluid flow in bone, which may indicate how bone cells sense changes in the mechanical environment. These first permeability measurements in mouse bone demonstrated that the permeability caused by fluid flowing through bone’s lacunar-canalicular porosity decreases with age. Porosity is expected to also affect bone’s ability to resist fracture at the micro-scale. The influence of intra-cortical porosity on crack propagation was explored via extended finite element methods. A novel technique was suggested to propagate cracks through holes and applied to 2D models of the porosity of osteogenesis imperfecta mouse bone. Results showed that vascular canals affect crack propagation and might contribute to the brittleness of osteogenesis impefecta bone. Skeletal pathologies often cause alterations in bone’s building blocks leading to deteriorated whole-bone toughness. Mineral properties of brittle and ductile mouse bone (models of osteogenesis imperfecta and rickets respectively) were evaluated. Results revealed that deviations in size, composition and organisation of bone mineral reduce bone’s mechanical integrity both in brittle and ductile pathologic bone. The outcomes of this thesis provide a deeper understanding of bone material, which is required for future improvements in treatments for skeletal diseases.Open Acces

Analytical model for the prediction of permeability of triply periodic minimal surfaces

Triply periodic minimal surfaces (TPMS) are mathematically defined cellular structures whose geometry can be quickly adapted to target desired mechanical response (structural and fluid). This has made them desirable for a wide range of bioengineering applications; especially as bioinspired materials for bone replacement. The main objective of this study was to develop a novel analytical framework which would enable calculating permeability of TPMS structures based on the desired architecture, pore size and porosity. To achieve this, computer-aided designs of three TPMS structures (Fisher-Koch S, Gyroid and Schwarz P) were generated with varying cell size and porosity levels. Computational Fluid Dynamics (CFD) was used to calculate permeability for all models under laminar flow conditions. Permeability values were then used to fit an analytical model dependent on geometry parameters only. Results showed that permeability of the three architectures increased with porosity at different rates, highlighting the importance of pore distribution and architecture. The computed values of permeability fitted well with the suggested analytical model (R2>0.99, p<0.001). In conclusion, the novel analytical framework presented in the current study enables predicting permeability values of TPMS structures based on geometrical parameters within a difference <5%. This model, which could be combined with existing structural analytical models, could open new possibilities for the smart optimisation of TPMS structures for biomedical applications where structural and fluid flow properties need to be optimised

Studies of chain substitution caused sub-fibril level differences in stiffness and ultrastructure of wildtype and oim/oim collagen fibers using multifrequency-AFM and molecular modeling

Molecular alteration in type I collagen, i.e., substituting the α2 chain with α1 chain in tropocollagen molecule, can cause osteogenesis imperfecta (OI), a brittle bone disease, which can be represented by a mouse model (oim/oim). In this work, we use dual-frequency Atomic Force Microscopy (AFM) and incorporated with molecular modeling to quantify the ultrastructure and stiffness of the individual native collagen fibers from wildtype (+/+) and oim/oim diseased mice humeri. Our work presents direct experimental evidences that the +/+ fibers have highly organized and compact ultrastructure and corresponding ordered stiffness distribution. In contrast, oim/oim fibers have ordered but loosely packed ultrastructure with uncorrelated stiffness distribution, as well as local defects. The molecular model also demonstrates the structural and molecular packing differences between +/+ and oim/oim collagens. The molecular mutation significantly altered sub-fibril structure and mechanical property of collagen fibers. This study can give the new insight for the mechanisms and treatment of the brittle bone disease

Towards a radiation free numerical modelling framework to predict spring assisted correction of scaphocephaly

Sagittal Craniosynostosis (SC) is a congenital craniofacial malformation, involving premature sagittal suture ossification; spring-assisted cranioplasty (SAC) – insertion of metallic distractors for skull reshaping – is an established method for treating SC. Surgical outcomes are predictable using numerical modelling, however published methods rely on computed tomography (CT) scans availability, which are not routinely performed. We investigated a simplified method, based on radiation-free 3D stereophotogrammetry scans.Eight SAC patients (age 5.1 ± 0.4 months) with preoperative CT and 3D stereophotogrammetry scans were included. Information on osteotomies, spring model and post-operative spring opening were recorded. For each patient, two preoperative models (PREOP) were created: i) CT model and ii) S model, created by processing patient specific 3D surface scans using population averaged skin and skull thickness and suture locations. Each model was imported into ANSYS Mechanical (Analysis System Inc., Canonsburg, PA) to simulate spring expansion. Spring expansion and cranial index (CI - skull width over length) at times equivalent to immediate postop (POSTOP) and follow up (FU) were extracted and compared with in-vivo measurements.Overall expansion patterns were very similar for the 2 models at both POSTOP and FU. Both models had comparable outcomes when predicting spring expansion. Spring induced CI increase was similar, with a difference of 1.2%±0.8% for POSTOP and 1.6%±0.6% for FU.This work shows that a simplified model created from the head surface shape yields acceptable results in terms of spring expansion prediction. Further modelling refinements will allow the use of this predictive tool during preoperative planning

Additively manufactured lattice structures with controlled transverse isotropy for orthopedic porous implants

Additively manufactured lattice structures enable the design of tissue scaffolds with tailored mechanical properties, which can be implemented in porous biomaterials. The adaptation of bone to physiological loads results in anisotropic bone tissue properties which are optimized for site-specific loads; therefore, some bone sites are stiffer and stronger along the principal load direction compared to other orientations. In this work, a semi-analytical model was developed for the design of transversely isotropic lattice structures that can mimic the anisotropy characteristics of different types of bone tissue. Several design possibilities were explored, and a particular unit cell, which was best suited for additive manufacturing was further analyzed. The design of the unit cell was parameterized and in-silico analysis was performed via Finite Element Analysis. The structures were manufactured additively in metal and tested under compressive loads in different orientations. Finite element analysis showed good correlation with the semi-analytical model, especially for elastic constants with low relative densities. The anisotropy measured experimentally showed a variable accuracy, highlighting the deviations from designs to additively manufactured parts. Overall, the proposed model enables to exploit the anisotropy of lattice structures to design lighter scaffolds with higher porosity and increased permeability by aligning the scaffold with the principal direction of the load

Meeting high precision requirements of additively manufactured components through hybrid manufacturing

A hybrid approach combining the laser powder bed fusion (LPBF) process and post-processing operations through 5-axis milling was employed to manufacture a Ti6Al4V aerospace component. From the design step, the requirements and needs in all the stages of the Hybrid Additive Manufacturing process were taken into account. A numerical simulation of distortions promoted by residual stresses during the additive process was employed to consider material allowance. The status of the as-built and post-processed component was analysed through scanning and CMM inspection and roughness measurements. The 3D scanned model of the as-built LPBF-ed component was used to understand the distortion behaviour of the component and compared to the numerical simulation. Finally, 5-axis milling operations were conducted in some critical surfaces in order to improve surface quality and dimensional accuracy of the as-built com- ponent. The inspection of the as-built and post-processed component showed the improvement achieved through the proposed hybrid approach. The work aims to provide the baselines needed to enable the metal Hybrid Additive Manufacturing of components with complex geometries where mandatory precision is required by integrating high accuracy machining operations as post-processing technique

In silico assessment of the bone regeneration potential of complex porous scaffolds

Mechanical environment plays a crucial role in regulating bone regeneration in bone defects. Assessing the mechanobiological behavior of patient-specific orthopedic scaffolds in-silico could help guide optimal scaffold designs, as well as intra- and post-operative strategies to enhance bone regeneration and improve implant longevity. Additively manufactured porous scaffolds, and specifically triply periodic minimal surfaces (TPMS), have shown promising structural properties to act as bone substitutes, yet their ability to induce mechanobiologially-driven bone regeneration has not been elucidated. The aim of this study is to i) explore the bone regeneration potential of TPMS scaffolds made of different stiffness biocompatible materials, to ii) analyze the influence of pre-seeding the scaffolds and increasing the post-operative resting period, and to iii) assess the influence of patient-specific parameters, such as age and mechanosensitivity, on outcomes. To perform this study, an in silico model of a goat tibia is used. The bone ingrowth within the scaffold pores was simulated with a mechano-driven model of bone regeneration. Results showed that the scaffold's architectural properties affect cellular diffusion and strain distribution, resulting in variations in the regenerated bone volume and distribution. The softer material improved the bone ingrowth. An initial resting period improved the bone ingrowth but not enough to reach the scaffold's core. However, this was achieved with the implantation of a pre-seeded scaffold. Physiological parameters like age and health of the patient also influence the bone regeneration outcome, though to a lesser extent than the scaffold design. This analysis demonstrates the importance of the scaffold's geometry and its material, and highlights the potential of using mechanobiological patient-specific models in the design process for bone substitutes

Meeting high precision requirements of additively manufactured components through hybrid manufacturing

A hybrid approach combining the laser powder bed fusion (LPBF) process and post-processing operations through 5-axis milling was employed to manufacture a Ti6Al4V aerospace component. From the design step, the requirements and needs in all the stages of the Hybrid Additive Manufacturing process were taken into account. A numerical simulation of distortions promoted by residual stresses during the additive process was employed to consider material allowance. The status of the as-built and post-processed component was analysed through scanning and CMM inspection and roughness measurements. The 3D scanned model of the as-built LPBF-ed component was used to understand the distortion behaviour of the component and compared to the numerical simulation. Finally, 5-axis milling operations were conducted in some critical surfaces in order to improve surface quality and dimensional accuracy of the as-built com-ponent. The inspection of the as-built and post-processed component showed the improvement achieved through the proposed hybrid approach. The work aims to provide the baselines needed to enable the metal Hybrid Additive Manufacturing of components with complex geometries where mandatory precision is required by integrating high accuracy machining operations as post-processing technique.(c) 2022 The Author(s). This is an open access article under the CC BY license (http://creativecommons.org/ licenses/by/4.0/)

Three-dimensionalenvironment and vascularization induce osteogenic maturation of human adipose-derived stem cells comparable to that of bone-derived progenitors

While human adipose-derived stem cells (hADSCs) are known to possess osteogenic differentiation potential, the bone tissues formed are generally considered rudimentary and immature compared with those made by bone-derived precursor cells such as human bone marrow-derived mesenchymal stem cells (hBMSCs) and less commonly studied human calvarium osteoprogenitor cells (hOPs). Traditional differentiation protocols have tended to focus on osteoinduction of hADSCs through the addition of osteogenic differentiation media or use of stimulatory bioactive scaffolds which have not resulted in mature bone formation. Here, we tested the hypothesis that by reproducing the physical as well as biochemical bone microenvironment through the use of three-dimensional (3D) culture and vascularization we could enhance osteogenic maturation in hADSCs. In addition to biomolecular characterization, we performed structural analysis through extracellular collagen alignment and mineral density in our bone tissue engineered samples to evaluate osteogenic maturation. We further compared bone formed by hADSCs, hBMSCs, and hOPs against mature human pediatric calvarial bone, yet not extensively investigated. Although bone generated by all three cell types was still less mature than native pediatric bone, a fibrin-based 3D microenvironment together with vascularization boosted osteogenic maturation of hADSC making it similar to that of bone-derived osteoprogenitors. This demonstrates the important role of vascularization and 3D culture in driving osteogenic maturation of cells easily available but constitutively less committed to this lineage and suggests a crucial avenue for recreating the bone microenvironment for tissue engineering of mature craniofacial bone tissues from pediatric hADSCs, as well as hBMSCs and hOPs

Lack of association of cranial lacunae with intracranial hypertension in children with Crouzon syndrome and Apert syndrome: a 3D morphometric quantitative analysis

Purpose Cranial lacunae (foci of attenuated calvarial bone) are CT equivalents ofBcopper beating seen on plain skull radio-graphs in children with craniosynostosis. The qualitative presence of copper beating has not been found to be useful for the diagnosis of intracranial hypertension (IH) in these patients. 3D morphometric analysis (3DMA) allows a more systematic and quantitative assessment of calvarial attenuation. We used 3DMA to examine the relationship between cranial lacunae and IH in children with Crouzon and Apert syndromic craniosynostosis