Alterations in subchondral bone plate, trabecular bone and articular cartilage properties of rabbit femoral condyles at 4 weeks after anterior cruciate ligament transection.

OBJECTIVE: To quantify early osteoarthritic-like changes in the structure and volume of subchondral bone plate and trabecular bone and properties of articular cartilage in a rabbit model of osteoarthritis (OA) induced by anterior cruciate ligament transection (ACLT). METHODS: Left knee joints from eight skeletally mature New Zealand white rabbits underwent ACLT surgery, while the contralateral (CTRL) right knee joints were left unoperated. Femoral condyles were harvested 4 weeks after ACLT. Micro-computed tomography imaging was applied to evaluate the structural properties of subchondral bone plate and trabecular bone. Additionally, biomechanical properties, structure and composition of articular cartilage were assessed. RESULTS: As a result of ACLT, significant thinning of the subchondral bone plate (P < 0.05) was accompanied by significantly reduced trabecular bone volume fraction and trabecular thickness in the medial femoral condyle compartment (P < 0.05), while no changes were observed in the lateral compartment. In both lateral and medial femoral condyles, the equilibrium modulus and superficial zone proteoglycan (PG) content were significantly lower in ACLT than CTRL joint cartilage (P < 0.05). Significant alterations in the collagen orientation angle extended substantially deeper into cartilage from the ACLT joints in the lateral femoral condyle relative to the medial condyle compartment (P < 0.05). CONCLUSIONS: In this model of early OA, significant changes in volume and microstructure of subchondral bone plate and trabecular bone were detected only in the femoral medial condyle, while alterations in articular cartilage properties were more severe in the lateral compartment. The former finding may be associated with reduced joint loading in the medial compartment due to ACLT, while the latter finding reflects early osteoarthritic changes in the lateral compartment.

Tissue viscoelasticity is related to tissue composition but may not fully predict the apparent-level viscoelasticity in human trabecular bone - An experimental and finite element study.

Trabecular bone is viscoelastic under dynamic loading. However, it is unclear how tissue viscoelasticity controls viscoelasticity at the apparent-level. In this study, viscoelasticity of cylindrical human trabecular bone samples (n=11, male, age 18-78 years) from 11 proximal femurs were characterized using dynamic and stress-relaxation testing at the apparent-level and with creep nanoindentation at the tissue-level. In addition, bone tissue elasticity was determined using scanning acoustic microscope (SAM). Tissue composition and collagen crosslinks were assessed using Raman micro-spectroscopy and high performance liquid chromatography (HPLC), respectively. Values of material parameters were obtained from finite element (FE) models by optimizing tissue-level creep and apparent-level stress-relaxation to experimental nanoindentation and unconfined compression testing values, respectively, utilizing the second order Prony series to depict viscoelasticity. FE simulations showed that tissue-level equilibrium elastic modulus (Eeq) increased with increasing crystallinity (r=0.730, p=.011) while at the apparent-level it increased with increasing hydroxylysyl pyridinoline content (r=0.718, p=.019). In addition, the normalized shear modulus g1 (r=-0.780, p=.005) decreased with increasing collagen ratio (amide III/CH2) at the tissue-level, but increased (r=0.696, p=.025) with increasing collagen ratio at the apparent-level. No significant relations were found between the measured or simulated viscoelastic parameters at the tissue- and apparent-levels nor were the parameters related to tissue elasticity determined with SAM. However, only Eeq, g2 and relaxation time τ1 from simulated viscoelastic values were statistically different between tissue- and apparent-levels (p<.01). These findings indicate that bone tissue viscoelasticity is affected by tissue composition but may not fully predict the macroscale viscoelasticity in human trabecular bone.

Effect of porosity, tissue density, and mechanical properties on radial sound speed in human cortical bone.

PURPOSE: The purpose of this study was to investigate the effect of simultaneous changes in cortical porosity, tissue mineral density, and elastic properties on radial speed of sound (SOS) in cortical bone. The authors applied quantitative pulse-echo (PE) ultrasound techniques that hold much potential especially for screening of osteoporosis at primary healthcare facilities. Currently, most PE measurements of cortical thickness, a well-known indicator of fracture risk, use a predefined estimate for SOS in bone to calculate thickness. Due to variation of cortical bone porosity, the use of a constant SOS value propagates to an unknown error in cortical thickness assessment by PE ultrasound. METHODS: The authors conducted 2.25 and 5.00 MHz focused PE ultrasound time of flight measurements on femoral diaphyses of 18 cadavers in vitro. Cortical porosities of the samples were determined using microcomputed tomography and related to SOS in the samples. Additionally, the effect of cortical bone porosity and mechanical properties of the calcified matrix on SOS was investigated using numerical finite difference time domain simulations. RESULTS: Both experimental measurements and simulations demonstrated significant negative correlation between radial SOS and cortical porosity (R(2) ≥ 0.493, p < 0.01 and R(2) ≥ 0.989, p < 0.01, respectively). When a constant SOS was assumed for cortical bone, the error due to variation of cortical bone porosity (4.9%-16.4%) was about 6% in the cortical thickness assessment in vitro. CONCLUSIONS: Use of a predefined, constant value for radial SOS in cortical bone, i.e., neglecting the effect of measured variation in cortical porosity, propagated to an error of 6% in cortical thickness. This error can be critical as characteristic cortical thinning of 1.10% ± 1.06% per yr decreases bending strength of the distal radius and results in increased fragility in postmenopausal women. Provided that the cortical porosity can be estimated in vivo, the relationship between radial SOS and cortical porosity can be utilized and a porosity based radial SOS estimate could be implemented to determine cortical thickness. This would constitute a step toward individualized quantitative ultrasound diagnostics of osteoporosis.

Relationships between tissue composition and viscoelastic properties in human trabecular bone.

Trabecular bone is a metabolically active tissue with a high surface to volume ratio. It exhibits viscoelastic properties that may change during aging. Changes in bone properties due to altered metabolism are sensitively revealed in trabecular bone. However, the relationships between material composition and viscoelastic properties of bone, and their changes during aging have not yet been elucidated. In this study, trabecular bone samples from the femoral neck of male cadavers (n=21) aged 17-82 years were collected and the tissue level composition and its associations with the tissue viscoelastic properties were evaluated by using Raman microspectroscopy and nanoindentation, respectively. For composition, collagen content, mineralization, carbonate substitution and mineral crystallinity were evaluated. The calculated mechanical properties included reduced modulus (Er), hardness (H) and the creep parameters (E1, E2, η1and η2), as obtained by fitting the experimental data to the Burgers model. The results indicated that the creep parameters, E1, E2, η1and η2, were linearly correlated with mineral crystallinity (r=0.769-0.924, p<0.001). Creep time constant (η2/E2) tended to increase with crystallinity (r=0.422, p=0.057). With age, the mineralization decreased (r=-0.587, p=0.005) while the carbonate substitution increased (r=0.728, p<0.001). Age showed no significant associations with nanoindentation parameters. The present findings suggest that, at the tissue-level, the viscoelastic properties of trabecular bone are related to the changes in characteristics of bone mineral. This association may be independent of human age.

Ultrasound backscatter measurements of intact human proximal femurs--relationships of ultrasound parameters with tissue structure and mineral density.

Ultrasound reflection and backscatter parameters are related to the mechanical and structural properties of bone in vitro. However, the potential of ultrasound reflection and backscatter measurements has not been tested with intact human proximal femurs ex vivo. We hypothesize that ultrasound backscatter can be measured from intact femurs and that the measured backscattered signal is associated with cadaver age, bone mineral density (BMD) and trabecular bone microstructure. In this study, human femoral bones of 16 male cadavers (47.0±16.1 years, range: 21-77 years) were investigated using pulse-echo ultrasound measurements at the femoral neck in the antero-posterior direction and at the trochanter major in the anteroposterior and lateromedial directions. Recently introduced ultrasound backscatter parameters, independent of cortical thickness, e.g., time slope of apparent integrated backscatter (TSAB) and mean of the backscatter difference technique (MBD) were obtained and compared with the structural properties of trabecular bone samples, extracted from the locations of ultrasound measurements. Moreover, more conventional backscatter parameters, e.g., apparent integrated backscatter (AIB) and frequency slope of apparent integrated backscatter (FSAB) were analyzed. Bone mineral density of the intact femurs was evaluated using dual energy X-ray absorptiometry (DXA). AIB and MDB measured from the femoral neck correlated significantly (p<0.01) with the neck BMD (R2=0.44 and 0.45), cadaver age (R2=0.61 and 0.41) and several structural parameters, e.g., bone volume fraction (R2=0.33 and 0.39, p<0.05 and p<0.01), respectively. To conclude, ultrasound backscatter parameters, measured from intact proximal femurs, are significantly related (p<0.05) to structural properties and mineral density of trabecular bone.

Longitudinal elastic properties and porosity of cortical bone tissue vary with age in human proximal femur.

Tissue level structural and mechanical properties are important determinants of bone strength. As an individual ages, microstructural changes occur in bone, e.g., trabeculae and cortex become thinner and porosity increases. However, it is not known how the elastic properties of bone change during aging. Bone tissue may lose its elasticity and become more brittle and prone to fractures as it ages. In the present study the age-dependent variation in the spatial distributions of microstructural and microelastic properties of the human femoral neck and shaft were evaluated by using acoustic microscopy. Although these properties may not be directly measured in vivo, there is a major interest to investigate their relationships with the linear elastic measurements obtained by diagnostic ultrasound at the most severe fracture sites, e.g., the femoral neck. However, before the validity of novel in vivo techniques can be established, it is essential to understand the age-dependent variation in tissue elastic properties and porosity at different skeletal sites. A total of 42 transverse cross-sectional bone samples were obtained from the femoral neck (Fn) and proximal femoral shaft (Ps) of 21 men (mean±SD age 47.1±17.8, range 17-82years). Samples were quantitatively imaged using a scanning acoustic microscope (SAM) equipped with a 50MHz ultrasound transducer. Distributions of the elastic coefficient (c33) of cortical (Ct) and trabecular (Tr) tissues and microstructure of cortex (cortical thickness Ct.Th and porosity Ct.Po) were determined. Variations in c33 were observed with respect to tissue type (c33Trc33(Ct.Fn)=35.3GPa>c33(Tr.Ps)=33.8GPa>c33(Tr.Fn)=31.9GPa), and cadaver age (R(2)=0.28-0.46, p<0.05). Regional variations in porosity were found in the neck (superior 13.1%; inferior 6.1%; anterior 10.1%; posterior 8.6%) and in the shaft (medial 9.5%; lateral 7.7%; anterior 8.6%; posterior 12.0%). In conclusion, significant variations in elastic coefficients were detected between femoral neck and shaft as well as between the quadrants of the cross-sections of neck and shaft. Moreover, an age-related increase in cortical porosity and a stiffening of the bone tissue were observed. These findings may explain in part the increase in susceptibility to suffer low energy fractures during aging and highlight the potential of ultrasound in clinical osteoporosis diagnostics.