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Osteoporosis is one of the most common skeletal diseases, but current therapies are limited to generalized antiresorptive or anabolic interventions, which do not target regions that would benefit from improvements to skeletal health. To improve the evaluation of treatment plans, we used a spatio-temporal multiscale approach that combines longitudinal in vivo micro-computed tomography (micro-CT) and in silico subject-specific finite element modeling to quantitatively map bone adaptation changes due to disease and treatment at high resolution. Our findings show time and region-dependent modifications in bone remodelling following one and two sets of mechanical loading and/or pharmacological interventions. The multiscale results highlighted that the distal section was unaffected by mechanical loading alone but the proximal tibia had the greatest gain from positive interactions of combined therapies. Mechanical loading abated the catabolic effect of PTH, but the main benefit of combined treatments occurred from the additive interactions of the two therapies in periosteal apposition. These results provide detailed insight into the efficacy of combined treatments, facilitating the optimisation of dosage and treatment duration in preclinical mouse studies, and the development of novel interventions for skeletal diseases. Statement of Significance Combined mechanical loading and pharmacotherapy have the potential to slow osteoporosis-induced bone loss but current therapies do not target the regions in need of strengthening. We show for the first time spatial region-dependant interactions between PTH and mechanical loading treatment in OVX mouse tibiae, highlighting local regions in the tibia that benefitted from separate and combined treatments. Combined experimental-computational analysis also detailed the lasting period of each treatment per location in the tibia, the extent of positive (or negative) interactions of the combined therapies, and the impact of each treatment on the regulation of bone adaptation spatio-temporally. This approach can be used to create hypothesis about the interactions of different treatments to optimise the design of biomaterials and medical interventions.

This preliminary study quantified tibia cartilage thickness (Cart.Th), subchondral bone plate thickness (SBPl.Th) and subchondral trabecular bone (STB) microarchitecture in subjects with varus- or valgus- malaligned knees diagnosed with end-stage knee osteoarthritis (OA) and compared them to controls (non-OA). Tibial plateaus from 25 subjects with knee-OA (undergoing knee arthroplasty) and 15 cadavers (controls) were micro-CT scanned (17 µm/voxel). Joint alignment was classified radiographically for OA subjects (varus-aligned n = 18, valgus-aligned n = 7). Cart.Th, SBPl.Th, STB bone volume fraction (BV/TV) and their medial-to-lateral ratios were analyzed in anteromedial, anterolateral, posteromedial and posterolateral subregions. Varus-OA and valgus-OA were compared to controls. Compared to controls (1.19-1.54 mm), Cart.Th in varus-OA was significantly lower anteromedially (0.58 mm, -59%) and higher laterally (2.19-2.47 mm, +60-63%); in valgus-OA, Cart.Th was significantly higher posteromedially (1.86 mm, +56%). Control medial-to-lateral Cart.Th ratios were around unity (0.8-1.1), in varus-OA significantly below (0.2-0.6) and in valgus-OA slightly above (1.0-1.3) controls. SBPl.Th and BV/TV were significantly higher medially in varus-OA (0.58-0.72 mm and 37-44%, respectively) and laterally in valgus-OA (0.60-0.61 mm and 32-37%), compared to controls (0.26-0.47 mm and 18-37%). In varus-OA, the medial-to-lateral SBPl.Th and BV/TV ratios were above unity (1.4-2.4) and controls (0.8-2.1); in valgus-OA they were closer to unity (0.8-1.1) and below controls. Varus- and valgus-OA tibia differ significantly from controls in Cart.Th, SBPl.Th and STB microarchitecture depending on joint alignment, suggesting structural changes in OA may reflect differences in medial-to-lateral load distribution upon the tibial plateau. Here we identified an inverse relationship between cartilage thickness and underlying subchondral bone, suggesting a whole-joint response in OA to daily stimuli.

Osteoporosis disrupts the healthy remodelling process in bone and affects its mechanical properties. Mechanical loading has been shown to be effective in stimulating bone formation to mitigate initial bone loss. However, no study has investigated the effects of repeated mechanical loading, with a pause of one week in between, in the mouse tibia with oestrogen deficiency. This study uses a combined experimental and computational approach, through longitudinal monitoring with micro-computed tomography, to evaluate the effects of loading on bone adaptation in the tibiae of ovariectomised (OVX) C57BL/6 mice from 14 to 22 weeks of age. Micro-FE models coupled with bone adaptation algorithms were used to estimate changes in local tissue strains due to OVX and mechanical loading, and to quantify the relationship between local strain and remodelling. The first in vivo mechanical loading increased apposition, by 50-150%, while resorption decreased by 50-60%. Both endosteal and periosteal resorption increased despite the second mechanical loading, and periosteal resorption was up to 70% higher than that after the first loading. This was found to correlate with an initial decrease in average strain energy density after the first loading, which was lower and more localised after the second loading. Predictions of bone adaptation showed that between 50 and 90% of the load-induced bone apposition is linearly strain driven at the organ-level, but resorption is more biologically driven at the local level. The results imply that a systematic increase in peak load or loading rate may be required to achieve a similar bone adaptation rate in specific regions of interests.

Combined treatment with PTH(1-34) and mechanical loading confers increased structural benefits to bone. However, it remains unclear how this longitudinal adaptation affects the bone mechanics. This study quantified the individual and combined longitudinal effects of PTH(1-34) and loading on micro-finite element (microFE) model estimates of tibia stiffness and strength in ovariectomised mice. C57BL/6 mice were ovariectomised at 14-weeks-old and treated either with injections of PTH(1-34), compressive tibia loading or both interventions concurrently. Right tibiae were in vivo microCT-scanned every two weeks from 14 until 24-weeks-old. MicroCT images were rigidly registered to reference tibia and the cortical organ (whole bone) and tissue-level (midshaft) morphometric properties and bone mineral content were quantified. MicroCT images were converted into voxel-based homogeneous, linear elastic microFE models to quantify bone stiffness and strength under uniaxial compression in vivo. Both stiffness and strength were higher with co-treatment than with individual therapies, consistent with increased benefits with the tibia bone mineral content and cortical area – properties strongly associated with the tibia mechanics. To our knowledge, this is the first study to longitudinally assess the mechanical effects of two bone anabolics, PTH(1-34) and tibia loading, in a mouse model of osteoporosis.

Biomechanical factors (e.g., joint loading) have a significant role in the progression of osteoarthritis (OA). However, some relationships between in vivo joint loading indices and tibial cartilage thickness are conflicting. This study investigated relationships between pre-operative in vivo external knee joint moments, joint alignment and regional tibial cartilage thickness using micro-CT in subjects with end-stage knee OA. Tibial plateaus from 25 patients that underwent knee replacement for OA were micro-CT scanned (17 µm/voxel). Prior to surgery, subjects underwent gait analysis to calculate external knee moments. The mechanical axis deviation (MAD) was obtained from pre-operative radiographs. Cartilage thickness (Cart.Th) was analyzed from micro-CT images, in anteromedial, anterolateral, posteromedial and posterolateral subregions of interest. Medial-to-lateral Cart.Th ratios were also explored. Relationships between Cart.Th and joint loading indices were examined using Pearson's correlations. Significant correlations were found between Cart.Th and joint loading indices, positive anteromedially with the first peak knee adduction moment (r = 0.55, p < 0.01) and external rotation moment (ERM; r = 0.52, p < 0.01), and negative with MAD (r = -0.76, p < 0.001). In the lateral regions, these correlations had opposite signs. The medial-to-lateral Cart.Th ratio correlated strongly with ERM (r = 0.63, p = 0.001) and MAD (r = -0.75, p < 0.001). Joint loading indices correlated with regional cartilage thickness values and their medial-to-lateral ratios in end-stage knee OA subjects, with higher regional loads corresponding to thinner cartilage. These relationships have the opposite sign compared to the subchondral bone microarchitecture found in our previous study on the same specimens, which may suggest a complementary bone-cartilage interplay in response to loading.

In preclinical mouse models, a synergistic anabolic response to PTH(1–34) and tibia loading was shown. Whether combined treatment improves bone properties with oestrogen deficiency, a cardinal feature of osteoporosis, remains unknown. This study quantified the individual and combined longitudinal effects of PTH(1–34) and loading on the bone morphometric and densitometric properties in ovariectomised mice. C57BL/6 mice were ovariectomised at 14-weeks-old and treated either with injections of PTH(1–34); compressive loading of the right tibia; both interventions concurrently; or both interventions on alternating weeks. Right tibiae were microCT-scanned from 14 until 24-weeks-old. Trabecular metaphyseal and cortical midshaft morphometric properties, and bone mineral content (BMC) in 40 different regions of the tibia were measured. Mice treated only with loading showed the highest trabecular bone volume fraction at week 22. Cortical thickness was higher with co-treatment than in the mice treated with PTH alone. In the mid-diaphysis, increases in BMC were significantly higher with loading than PTH. In ovariectomised mice, the osteogenic benefits of co-treatment on the trabecular bone were lower than loading alone. However, combined interventions had increased, albeit regionally-dependent, benefits to cortical bone. Increased benefits were largest in the mid-diaphysis and postero-laterally, regions subjected to higher strains under compressive loads.

Oestrogen deficiency-related bone loss in the ovariectomized (OVX) mouse is a common model for osteoporosis. However, a comprehensive in vivo assessment of intervention-related changes in multiple bone properties, and in multiple mouse strains, is required in order to identify an appropriate model for future evaluation of novel anti-osteoporotic therapies. The aim of this study was to evaluate the effect of OVX on the morphometric and densitometric properties measured in the microCT images and the mechanical properties estimated with finite element models of the tibia in two mouse strains, C57BL/6 and BALB/c.\ud \ud \ud \ud 14-weeks-old female C57BL/6 and BALB/c mice were divided into two groups per strain: (1) ovariectomized, (2) non-operated control. The right tibia was scanned at baseline (14 weeks) and then every two weeks thereafter, until 24-weeks-old, using in vivo microCT. Changes in trabecular and cortical bone morphometry, spatiotemporal changes in densitometric properties and in mechanical properties (from micro-finite element (μFE) analysis) were computed. Differences between OVX and non-operated controls were evaluated by ANCOVA, adjusted for 14-weeks baseline.\ud \ud \ud \ud In morphometry, trabecular bone mass was significantly reduced in both C57BL/6 and BALB/c from four weeks following surgery. Though the OVX-effect was transient in BALB/c as bone mass reached skeletal homeostasis. OVX inhibited the age-related thickening of cortical bone only in C57BL/6. In both strains, increments in bone mineral content were significantly lower with OVX only in the proximal tibia, with intervention-related differences increasing with time. OVX had no effect on μFE estimates of stiffness nor failure load in either strain.\ud \ud \ud \ud The results of this study show strain-, time- and region-(trabecular or cortical) dependent changes in morphometric and densitometric properties. These findings highlight the importance of choosing an appropriate mouse model and time points for research of treatments against accelerated bone resorption.

Tibial subchondral bone plays an important role in knee osteoarthritis (OA). Microarchitectural characterisation of subchondral bone plate (SBP), underlying trabecular bone (STB) and relationships between microarchitecture in these compartments, however, is limited. The aim of this study was to characterise the spatial distribution of SBP thickness and porosity and STB microarchitecture, and relationships between compartments, in OA tibiae of varying joint alignment. Twenty-five tibial plateaus from end-stage knee-OA patients, with varus (n= 17) or non-varus (n= 8) alignment were micro-CT scanned (17µm/voxel). SBP and STB microarchitecture was quantified via a systematic mapping in 22 volumes of interest per knee (11 medial, 11 lateral). Significant within-condylar and between-condylar (medial vs. lateral) differences (p

Objective: To study, in end-stage knee osteoarthritis (OA) patients, relationships between indices of in vivo dynamic knee joint loads obtained pre-operatively using gait analysis, static knee alignment, and the subchondral trabecular bone (STB) microarchitecture of their excised tibial plateau quantified with 3D micro-CT. Design: Twenty-five knee OA patients scheduled for total knee arthroplasty underwent pre-operative gait analysis. Mechanical axis deviation (MAD) was determined radiographically. Following surgery, excised tibial plateaus were micro-CT-scanned and STB microarchitecture analysed in four subregions (anteromedial, posteromedial, anterolateral, posterolateral). Regional differences in STB microarchitecture and relationships between joint loading and microarchitecture were examined. Results: STB microarchitecture differed among subregions (P < 0.001), anteromedially exhibiting highest bone volume fraction (BV/TV) and lowest structure model index (SMI). Anteromedial BV/TV and SMI correlated strongest with the peak external rotation moment (ERM; r = -0.74, r = 0.67, P < 0.01), despite ERM being the lowest (by factor of 10) of the moments considered, with majority of ERM measures below accuracy thresholds; medial-to-lateral BV/TV ratios correlated with ERM, MAD, knee adduction moment (KAM) and internal rotation moment (|r|-range: 0.54-0.74). When controlling for walking speed, KAM and MAD, the ERM explained additional 11-30% of the variations in anteromedial BV/TV and medial-to-lateral BV/TV ratio (R 2 = 0.59, R 2 = 0.69, P < 0.01). Conclusions: This preliminary study suggests significant associations between tibial plateau STB microarchitecture and knee joint loading indices in end-stage knee OA patients. Particularly, anteromedial BV/TV correlates strongest with ERM, whereas medial-to-lateral BV/TV ratio correlates strongest with indicators of medial-to-lateral joint loading (MAD, KAM) and rotational moments. However, associations with ERM should be interpreted with caution. Refereed/Peer-reviewed

Objectives: To (1) stratify patient subgroups according to their distinct walking gait patterns in end-stage knee osteoarthritis (OA); (2) compare measures of joint loading and proximal tibia subchondral trabecular bone (STB) microarchitecture among these gait subgroups. Design: Twenty-five knee OA patients undergoing total knee arthroplasty (TKA) had pre-operative gait analysis. Following surgery, excised tibial plateaus were micro-CT-scanned and STB microarchitecture analysed in four tibial condylar regions of interest. Peak knee moments were input to k-means cluster analysis, to identify subgroups with homogeneous gait patterns. Joint loading and STB microarchitecture parameters were compared among gait subgroups (Kruskal-Wallis, Bonferroni-corrected Mann-Whitney U tests). Results: Three gait subgroups were revealed: biphasics (n = 7), flexors (n = 9), counter-rotators (n = 9). Peak knee adduction moment (KAM) and KAM impulse were significantly higher (P < 0.05) in biphasics than in flexors and counter-rotators (KAM = -0.65, -0.40 and -0.21 Nm/kg, respectively), suggesting a higher medial-to-lateral tibiofemoral load ratio in biphasics. Interestingly, STB medial-to-lateral bone volume fraction (BV/TV) ratio was also significantly higher (more than double) in biphasics and flexors than in counter-rotators (2.24, 2.00 and 1.00, respectively), whereas in biphasics it was only 10% higher than in flexors and not significantly so. Conclusions: Within the confines of the limited sample size, data suggests that different mechanisms between the biphasic and flexor gait subroups may generate comparable loads upon the tibial plateau and corresponding bony responses, despite significantly lower KAM indices in flexors. Hence, in flexor gait OA patients, conservative treatments designed to reduce KAM, may not be appropriate. Understanding joint loading among walking gait patterns and relationships to bone microarchitecture may aid at identifying/improving management of persons at risk for developing knee OA. Refereed/Peer-reviewed

Digital volume correlation (DVC) provides experimental measurements of displacements and strains throughout the interior of porous materials such as trabecular bone. It can provide full-field continuum- and tissue-level measurements, desirable for validation of finite element models, by comparing image volumes from subsequent µCT scans of a sample in unloaded and loaded states. Since the first application of DVC for measurement of strain in bone tissue, subsequent reports of its application to trabecular bone cores up to whole bones have appeared within the literature. An “optimal” set of procedures capable of precise and accurate measurements of strain, however, still remains unclear, and a systematic review focussing explicitly on the increasing number of DVC algorithms applied to bone or structurally similar materials is currently unavailable. This review investigates the effects of individual parameters reported within individual studies, allowing to make recommendations for suggesting algorithms capable of achieving high accuracy and precision in displacement and strain measurements. These recommendations suggest use of subsets that are sufficiently large to encompass unique datasets (e.g. subsets of 500 µm edge length when applied to human trabecular bone cores, such as cores 10 mm in height and 5 mm in diameter, scanned at 15 µm voxel size), a shape function that uses full affine transformations (translation, rotation, normal strain and shear strain), the robust normalized cross-correlation coefficient objective function, and high-order interpolation schemes. As these employ computationally burdensome algorithms, researchers need to determine whether they have the necessary computational resources or time to adopt such strategies. As each algorithm is suitable for parallel programming however, the adoption of high precision techniques may become more prevalent in the future.