Steven K. Boyd

Bone strength at the distal radius can be estimated from high-resolution peripheral quantitative computed tomography and the finite element method

Bone strength is a fundamental contributor to fracture risk, and with the recent development of in vivo 3D bone micro-architecture measurements by high-resolution peripheral quantitative computed tomography, the finite element (FE) analysis may provide a means to assess patient bone strength in the distal radius. The purpose of this study was to determine an appropriate FE procedure to estimate bone strength by comparison with experimental data. Models based on a homogeneous tissue modulus or a modulus scaled according to computed tomography attenuation were assessed, and these were solved by linear and non-linear FE analyses to estimate strength. The distal radius from fresh, human cadaver forearms (5 male/5 female, ages 55 to 93) was dissected free and four 9.1 mm sections were cut beginning at the subchondral plate to provide 40 test specimens. The sections were scanned using an in vivo protocol providing 3D image data with an 82 microm voxel size. All specimens were mechanically tested in uniaxial compression, and elastic and yield properties were determined. Linear FE analyses were performed on all specimens (N=40), and non-linear analyses using an asymmetric, bilinear yield strain criteria were performed on a sub-sample (N=10) corresponding to the normal clinical measurement site. Experimentally determined apparent elastic properties correlated highly with ultimate stress (R2=0.977, p<0.05, N=31) for the 31 specimens tested to failure. Subsequently, a linear FE analysis estimating apparent elastic properties also correlated highly with failure, and the correlation was higher when moduli were determined from scaled CT-attenuation values than a homogeneous modulus (R2=0.983 vs. R2=0.972, p<0.05, N=31). A non-linear analysis based on tensile and compressive yield strains of 0.0295 and 0.0493 for homogeneous models, and 0.0127 and 0.0212 for scaled models directly estimated ultimate stress, and correlated highly (R2=0.951 vs. R2=0.937, p<0.05, N=5). The linear relation between stiffness and strength may be unique to radius compressive loading. It supports the use of a linear FE analysis to determine bone strength by regression equations established here. Scaled tissue modulus models performed better than homogeneous modulus models, and the advantage of a scaled model is its potential to account for mineralization changes. The combined numerical-experimental procedure for FE model validation on the patient micro-CT technology demonstrated that bone strength can be estimated non-invasively, and this may provide important insight into fracture risk in patient populations.

Reproducibility of direct quantitative measures of cortical bone microarchitecture of the distal radius and tibia by HR-pQCT☆

Quantitative cortical microarchitectural end points are important for understanding structure-function relations in the context of fracture risk and therapeutic efficacy. This technique study details new image-processing methods to automatically segment and directly quantify cortical density, geometry, and microarchitecture from HR-pQCT images of the distal radius and tibia. An automated segmentation technique was developed to identify the periosteal and endosteal margins of the distal radius and tibia and detect intracortical pore space morphologically consistent with Haversian canals. The reproducibility of direct quantitative cortical bone indices based on this method was assessed in a pooled data set of 56 subjects with two repeat acquisitions for each site. The in vivo precision error was characterized using root mean square coefficient of variation (RMSCV%) from which the least significant change (LSC) was calculated. Bland-Altman plots were used to characterize bias in the precision estimates. The reproducibility of cortical density and cross-sectional area measures was high (RMSCV <1% and <1.5%, respectively) with good agreement between young and elder medians. The LSC for cortical porosity (Ct.Po) was somewhat smaller in the radius (0.58%) compared with the distal tibia (0.84%) and significantly different between young and elder medians in the distal tibia (LSC: 0.75% vs. 0.92%, p<0.001). The LSC for pore diameter and distribution (Po.Dm and Po.Dm.SD) ranged between 15 and 23 microm. Bland-Altman analysis revealed moderate bias for integral measures of area and volume but not for density or microarchitecture. This study indicates that HR-pQCT measures of cortical bone density and architecture can be measured in vivo with high reproducibility and limited bias across a biologically relevant range of values. The results of this study provide informative data for the design of future clinical studies of bone quality.

Postmenopausal women with osteopenia have higher cortical porosity and thinner cortices at the distal radius and tibia than women with normal aBMD: an in vivo HR-pQCT study

Increases in cortical porosity (Ct.Po) and decreases in cortical thickness (Ct.Th) are associated with increased bone fragility. The purpose of this study was to validate an autosegmentation method for high‐resolution peripheral quantitative computed tomography (HR‐pQCT) scans to measure Ct.Po and Ct.Th and use it to compare Ct.Po and Ct.Th between pre‐ and postmenopausal women with normal, osteopenic, and osteoporotic areal bone mineral density (aBMD). The Ct.Po and Ct.Th measurements were validated using cadaver forearms (n = 10) and micro‐computed tomography (µCT) as the gold standard. The analysis was applied to distal radius and tibia HR‐pQCT scans from a subset of participants from the Calgary, Alberta, cohort of the Canadian Multicentre Osteoporosis Study (n = 280, 18 to 90 years). Analysis of covariance compared Ct.Po and Ct.Th outcomes between 63 normal premenopausal (dual‐energy X‐ray absorptiometry femoral neck T‐score > −1), 87 normal postmenopausal, 121 osteopenic postmenopausal, and 9 osteoporotic postmenopausal women. Linear regression analysis and Bland‐Altman plots were used to assess the agreement between the HR‐pQCT and µCT measurements, resulting in r2 values of 0.80 for Ct.Po and 0.98 for Ct.Th. At both sites, Ct.Po was higher in postmenopausal (all groups) than in premenopausal women (3.2% to 12.9%, p < .001). Ct.Th was not significantly different between normal premenopausal and postmenopausal women at either site; however, both osteopenic and osteoporotic women had thinner (−12.8% to −30.3%, p < .01), more porous (2.1% to 8.1%, p < .001) cortices than normal postmenopausal women. Our method offers promise as a valuable tool to measure Ct.Po and Ct.Th in vivo and investigate associations among cortical bone structure, age, and disease status.

Age-related patterns of trabecular and cortical bone loss differ between sexes and skeletal sites: a population-based HR-pQCT study.

In this cross‐sectional study, we aimed to predict age‐related changes in bone microarchitecture and strength at the distal radius (DR) and distal tibia (DT) in 644 Canadian adults (n = 442 women and 202 men) aged 20 to 99 years. We performed a standard morphologic analysis of the DR and DT with high‐resolution peripheral quantitative computed tomography (pQCT) and used finite‐element analysis (FEA) to estimate bone strength (failure load) and the load distribution. We also calculated a DR load‐to‐strength ratio as an estimate of forearm fracture risk. Total bone area, which was 33% larger in young men at both sites, changed similarly with age in women and men at the DT but increased 17% more in men than in women at the DR (p < .001). Trabecular number and thickness (Tb.Th) were 7% to 20% higher in young men than in young women at both sites, and with the exception of Tb.Th at the DR, which declined more with age in men (−16%) than in women (−2%, p < .01), the age‐related decline in these outcomes was similar in women and in men. In the cortex, porosity (Ct.Po) was 31% to 44% lower in young women than in young men but increased 92% to 176% more with age in women than in men (p < .001). The DR cortex carried 14% more load in young women than in young men, and the percentage of load carried by the DR cortex did not change with age in women but declined by 17% in men (p < .01). FEA‐estimated bone strength was 34% to 47% greater in young men, but the predicted change with age was similar in both sexes. In contrast, the load‐to‐strength ratio increased 27% more in women than in men with age (p < .01). These results highlight important site‐ and sex‐specific differences in patterns of age‐related bone loss. In particular, the trends for less periosteal expansion, more porous cortices, and a greater percentage of load carried by the DR cortex in women may underpin sex differences in forearm fracture risk.

Microarchitectural deterioration of cortical and trabecular bone: Differing effects of denosumab and alendronate

The intensity of bone remodeling is a critical determinant of the decay of cortical and trabecular microstructure after menopause. Denosumab suppresses remodeling more than alendronate, leading to greater gains in areal bone mineral density (aBMD). These greater gains may reflect differing effects of each drug on bone microarchitecture and strength. In a phase 2 double‐blind pilot study, 247 postmenopausal women were randomized to denosumab (60 mg subcutaneous 6 monthly), alendronate (70 mg oral weekly), or placebo for 12 months. All received daily calcium and vitamin D. Morphologic changes were assessed using high‐resolution peripheral quantitative computed tomography (HR‐pQCT) at the distal radius and distal tibia and QCT at the distal radius. Denosumab decreased serum C‐telopeptide more rapidly and markedly than alendronate. In the placebo arm, total, cortical, and trabecular BMD and cortical thickness decreased (−2.1% to −0.8%) at the distal radius after 12 months. Alendronate prevented the decline (−0.6% to 2.4%, p = .051 to <.001 versus placebo), whereas denosumab prevented the decline or improved these variables (0.3% to 3.4%, p < .001 versus placebo). Changes in total and cortical BMD were greater with denosumab than with alendronate (p ≤ .024). Similar changes in these parameters were observed at the tibia. The polar moment of inertia also increased more in the denosumab than alendronate or placebo groups (p < .001). Adverse events did not differ by group. These data suggest that structural decay owing to bone remodeling and progression of bone fragility may be prevented more effectively with denosumab.

Improved reproducibility of high-resolution peripheral quantitative computed tomography for measurement of bone quality

A human high-resolution peripheral quantitative computed tomography scanner (HR-pQCT) (XtremeCT, Scanco Medical, Switzerland) capable of measuring three important indicators of bone quality (micro-architectural morphology, mineralization and mechanical stiffness) has been developed. The goal of this study was to evaluate the reproducibility of male and female HR-pQCT in vivo measurements, and elucidate the causes of error in these measurements through a comparison with in vitro measurements. The best possible short-term reproducibility was found using a set of 10 in vitro measurements without repositioning, and a set of 10 with repositioning. Subsequently, in vivo measurements were performed on 15 male and 15 female subjects at baseline and follow-ups of 1 week and 4 months to determine the short- and long-term reproducibility of the system. In addition to the 2D area matching method used in the standard evaluation protocol, a custom developed 3D registration method was used to find the common region between repeated scans. The best possible reproducibility without movement artifacts and repositioning error was less than 0.5%, while the reproducibility with repositioning error was less than 1.5%. The in vivo reproducibility of density (<1%), morphological (<4.5%) and stiffness (<3.5) measurements was consistently poorer than the reproducibility of cadaver measurements, presumably due to small movement artifacts and repositioning errors. Using 3D image registration, repositioning error was reduced on average by 23% and 8% for measurements of the radius and tibia sites, respectively. This study has provided bounds for the reproducibility of HR-pQCT to monitor bone quality longitudinally, and a basis for clinical study design to determine detectable changes.

The Lysyl Oxidase Inhibitor, β-Aminopropionitrile, Diminishes the Metastatic Colonization Potential of Circulating Breast Cancer Cells

Lysyl oxidase (LOX), an extracellular matrix remodeling enzyme, appears to have a role in promoting breast cancer cell motility and invasiveness. In addition, increased LOX expression has been correlated with decreases in both metastases-free, and overall survival in breast cancer patients. With this background, we studied the ability of β-aminopropionitrile (BAPN), an irreversible inhibitor of LOX, to regulate the metastatic colonization potential of the human breast cancer cell line, MDA-MB-231. BAPN was administered daily to mice starting either 1 day prior, on the same day as, or 7 days after intracardiac injection of luciferase expressing MDA-MB-231-Luc2 cells. Development of metastases was monitored by in vivo bioluminescence imaging, and tumor-induced osteolysis was assessed by micro-computed tomography (μCT). We found that BAPN administration was able to reduce the frequency of metastases. Thus, when BAPN treatment was initiated the day before, or on the same day as the intra-cardiac injection of tumor cells, the number of metastases was decreased by 44%, and 27%, and whole-body photon emission rates (reflective of total tumor burden) were diminished by 78%, and 45%, respectively. In contrast, BAPN had no effect on the growth of established metastases. Our findings suggest that LOX activity is required during extravasation and/or initial tissue colonization by circulating MDA-MB-231 cells, lending support to the idea that LOX inhibition might be useful in metastasis prevention.

Mechanical and Architectural Bone Adaptation in Early Stage Experimental Osteoarthritis

The purpose of this study was to quantify mechanical and architectural changes to knee joint periarticular subchondral cancellous bone in early stage experimental osteoarthritis (OA). Unilateral anterior cruciate ligament transection (ACLX) was performed on 10 dogs that were assigned randomly to two groups: 3 weeks or 12 weeks post‐ACLX. Cylindrical bone cores excised from the medial condyle of the distal femur after death were scanned using high‐resolution microcomputed tomography (μCT) and subsequently failed under unconstrained uniaxial compression. The apparent‐level elastic modulus was less in the ACLX femur compared with the contralateral control, and the decrease was significant (−45%; p < 0.05) by 12‐weeks post‐ACLX. A finite element (FE) analysis based on μCT data simulated the uniaxial compression tests on a specimen‐by‐specimen basis to determine tissue modulus. No change in tissue modulus was detected, and a single tissue modulus of 5100 MPa (95% CI, ±600 MPa) explained the apparent‐level modulus changes observed in the disease‐related bone adaptation. The three‐dimensional (3D) connectivity was evaluated from the original μCT data to quantify architectural alterations in contrast to tissue alterations. Significantly increased connectivity (through plate perforations) occurred as early as 3 weeks post‐ACLX and was as high as 127% by 12 weeks post‐ACLX in the distal femur. These measured changes indicated that architectural adaptation predominated over tissue modulus changes affecting apparent‐level elastic modulus in the early stage of experimental OA and suggests that to maintain normal cancellous bone after a traumatic injury, early intervention should focus on preventing the substantial architectural alterations.

Radiation effects on bone architecture in mice and rats resulting from in vivo micro-computed tomography scanning.

Recently established techniques for performing in vivo micro-computed tomography (micro-CT) provide the capability of monitoring bone changes in a living animal at various points in time. However, radiation exposure from repeated micro-CT scans may have an effect on skeletal growth in normal or disease-model animals. The purpose of this study is to test a high resolution (approximately 10 microm) in vivo micro-CT protocol on mice and rats used for bone research to understand the impact of micro-CT radiation exposure on bone architecture. Ovariectomy (OVX) or sham-OVX surgery was performed on groups (n=6-8/group) of 12-week-old C3H/HeJ, C57BL/6J, and BALB/cByJ mice, and one strain of rat (Wistar, retired breeders). The right proximal tibiae were scanned at weekly intervals while the contralateral left limbs were not scanned until the endpoint of the protocol. Trabecular and cortical bone morphology was compared between radiated and non-radiated limbs at the endpoint to quantify the radiation effect. No effects of radiation were observed in OVX or sham rats. Lower trabecular bone volume was observed in the radiated limbs (-8 to -20% relative to non-radiated limb) of all mice groups except sham BALB/cByJ mice and normal control C57BL/6J mice, however, the observed effects were much less than the observed effects of ovariectomy ( approximately 40-50% total bone volume reduction, depending on mouse strain), and no interactions between radiation and OVX treatment were observed (p>0.2). Using an internal non-radiated control within each animal is a potential method to elucidate the effect of radiation exposure for any in vivo protocol. Thus, although in vivo micro-CT is a valuable tool for bone-related research, the impact of radiation in skeletally immature mice should be considered, particularly for strains with low bone volume at the measured site.

Cortical porosity is higher in boys compared with girls at the distal radius and distal tibia during pubertal growth: An HR-pQCT study

The aim of this study was to determine the sex‐ and maturity‐related differences in bone microstructure and estimated bone strength at the distal radius and distal tibia in children and adolescents. We used high‐resolution pQCT to measure standard morphological parameters in addition to cortical porosity (Ct.Po) and estimated bone strength by finite element analysis. Participants ranged in age from 9 to 22 years (n = 212 girls and n = 186 boys) who were scanned annually for either one (11%) or two (89%) years at the radius and for one (15%), two (39%), or three (46%) years at the tibia. Participants were grouped by the method of Tanner into prepubertal, early pubertal, peripubertal, and postpubertal groups. At the radius, peri‐ and postpubertal girls had higher cortical density (Ct.BMD; 9.4% and 7.4%, respectively) and lower Ct.Po (–118% and–56%, respectively) compared with peri‐ and postpubertal boys (all p < 0.001). Peri‐ and postpubertal boys had higher trabecular bone volume ratios (p < 0.001) and larger cortical cross‐sectional areas (p < 0.05, p < 0.001) when compared with girls. Based upon the load‐to‐strength ratio (failure load/estimated fall force), boys had lower risk of fracture than girls at every stage except during early puberty. Trends at the tibia were similar to the radius with differences between boys and girls in Ct.Po (p < 0.01) and failure load (p < 0.01) at early puberty. Across pubertal groups, within sex, the most mature girls and boys had higher Ct.BMD and lower Ct.Po than their less mature peers (prepuberty) at both the radius and tibia. Girls in early, peri‐, and postpubertal groups and boys in peri‐ and postpubertal groups had higher estimates of bone strength compared with their same‐sex prepubertal peers (p < 0.001). These results provide insight into the sex‐ and maturity‐related differences in bone microstructure and estimated bone strength.