Cortical Bone Specimens Research Articles

J024035 皮質骨組織の衝撃破壊強度に及ぼす微視的損傷の影響([J024-03]光学・力学のバイオメカニクス応用)

Cortical bone has anisotropic mechanical properties depending on the hierarchal structure of the tissue observed on multi-scales. Microscopic structure of bone tissue contains micro structural defects such as Haversian canal, osteocyte, boundary of osteons and microcracks. This study focused on fracture behaviors of cortical bone tissue including micro-nano scale structural defects. A compact-sized loading machine for Charpy impact tests was developed to measure the fracture toughness of bone specimens. The cortical bone specimens of 3 x 3 x 30 mm were prepared from a shaft of bovine femur. The differences relating the anisotropy of each specimen were observed in the fracture toughness and the fracture surfaces. The fracture energy values of the bone axial specimens were greater than of the circumferential specimens. The bone axial specimens were demineralized in EDTA solution for several days. The demineralized specimens showed different fracture surfaces and different fracture energy compared with intact specimens. The 48 hour demineralized specimen required the highest energy for fracture. The long term of 1 month demineralized specimen had high bendability and the specimen was not fractured in the impact tests.

Changes in the viscoelastic properties of cortical bone by selective degradation of matrix protein

We have studied stress relaxation of bovine femoral cortical bone specimens treated with KOH aqueous solution which had been known to degrade selectively protein molecules in bone. With the KOH treatment, we found an increase in specimens' volume. This increase was regarded as swelling of the bone specimen, presumably due to matrix protein network degradation including that of collagen. In an analogy of bone to gel structure, an increasing ratio of specimen volume was used as an indicating parameter for the matrix protein network degradation by the treatment. Although an empirical equation with a linearly combined form of two Kohlrausch–Williams–Watts (KWW) functions has been shown to describe the stress relaxation of bone specimens, a single KWW function was suitable for the bone specimens treated with KOH solution for as little as 3h. In KOH treated specimens, both the initial modulus and the relaxation time decreased with the volume-increasing ratio, while the relaxation time distribution did not change. A chemo-rheological consideration attributed the reduction of modulus values to the network degradation in the organic matrix phase. The relaxation time of KOH treated specimens was thought to be related to the longer relaxation time of untreated bones, although there was a discontinuity between the extrapolated relaxation time values for KOH treated specimens and untreated specimens. This discontinuity may have originated from the release of residual stress existing in the bone by the matrix protein degradation. The results of the present study suggest that the state of matrix protein is crucial for integrating the mechanical properties of bone.

Analysis of fracture processes in cortical bone tissue

Bones are the principal structural components of a skeleton; they play unique roles in the body providing its shape maintenance, protection of internal organs and transmission of forces. Ultimately, their structural integrity is vital for the quality of life. Unfortunately, bones can only sustain loads until a certain limit, beyond which they fail. Understanding a fracture behaviour of bone is necessary for prevention and diagnosis of trauma; this can be achieved by studying mechanical properties of bone, such as its fracture toughness. Generally, most of bone fractures occur in long bones consisting mostly of cortical bone tissue. Therefore, in this paper, an experimental study and numerical simulations of fracture processes in a bovine femoral cortical bone tissue were considered. A set of experiments was conducted to characterise fracture toughness of the bone tissue in order to gain basic understanding of spatial variability and anisotropy of its resistance to fracture and its link to an underlying microstructure. The data was obtained using single-edge-notch-bending specimens of cortical bone tested in a three-point bending setup; fracture surfaces of specimens were studied using scanning electron microscopy. Based on the results of those experiments, a number of finite-element models were developed in order to analyse its deformation and fracture using the extended finite-element method (X-FEM). Experimental results of this study demonstrate both variability and anisotropy of fracture toughness of the cortical bone tissue; the developed models adequately reflected the experimental data.

Accurate measurement of cortical bone elasticity tensor with resonant ultrasound spectroscopy

Resonant ultrasound spectroscopy (RUS) allows to accurately characterize the complete set of elastic constants of an anisotropic material from a set of measured mechanical resonant frequencies of a specimen. This method does not suffer from the drawbacks and limitations of the conventional sound velocity approach, but has been reported to fail to measure bone because of its strong viscoelastic damping. In this study, we take advantage of recent developments of RUS to overcome this limitation. The frequency response of a human cortical bone specimen (about 5×7×7mm3) was measured between 100 and 280kHz. Despite an important overlapping of the resonant peaks 20 resonant frequencies could be retrieved by using a dedicated signal processing method. The experimental frequencies were progressively matched to the frequencies predicted by a model of the sample whose elastic constants were adjusted. The determined diagonal elastic constants were in good agreement with concurrent sound velocity measurements performed in the principal directions of the specimen. This study demonstrates that RUS is suitable for an accurate measurement of cortical bone anisotropic elasticity. In particular, precision of measured Young and shear moduli is about 0.5%.

Design and validation of bending test method for characterization of miniature pediatric cortical bone specimens

Osteogenesis imperfecta is a genetic disorder of bone fragility; however, the effects of this disorder on bone material properties are not well understood. No study has yet measured bone material strength in humans with osteogenesis imperfecta. Small bone specimens are often extracted during routine fracture surgeries in children with osteogenesis imperfecta. These specimens could provide valuable insight into the effects of osteogenesis imperfecta on bone material strength; however, their small size poses a challenge to their mechanical characterization. In this study, a validated miniature three-point bending test is described that enables measurement of the flexural material properties of pediatric cortical osteotomy specimens as small as 5 mm in length. This method was validated extensively using bovine bone, and the effect of span/depth aspect ratio (5 vs 6) on the measured flexural properties was examined. The method provided reasonable results for both Young's modulus and flexural strength in bovine bone. With a span/depth ratio of 6, the median longitudinal modulus and flexural strength results were 16.1 (range: 14.4-19.3)GPa and 251 (range: 219-293)MPa, respectively. Finally, the pilot results from two osteotomy specimens from children with osteogenesis imperfecta are presented. These results provide the first measures of bone material strength in this patient population.

The relative influence of apatite crystal orientations and intracortical porosity on the elastic anisotropy of human cortical bone

Elastic anisotropy exhibits spatial inhomogeneity in human cortical bone, but the structural origins of anatomic variation are not well understood. In this study, the elastic anisotropy of human cortical bone was predicted using a specimen-specific multiscale model that investigated the relative influence of apatite crystal orientations and intracortical porosity. The elastic anisotropy of cortical bone specimens from the diaphysis of human femora was measured by ultrasonic wave propagation as the ratio of elastic constants in the longitudinal/radial (L/R) and longitudinal/circumferential (L/C) anatomic specimen axes. Experimental measurements of elastic constants exhibited orthotropy, with greater anisotropy in the L/R plane compared to the L/C plane. Model predictions included (1) a micromechanical model accounting for the effects of apatite crystal orientations, (2) a voxel-based finite element model accounting for the effects of intracortical porosity, and (3) a combined model accounting for both effects. The combined model provided the most accurate predictions of elastic anisotropy in both the L/R and L/C plane, with less than 10% mean error. The micromechanical model alone was able to accurately predict elastic anisotropy in the L/C plane, but predicted transverse isotropy. The finite element model alone grossly underestimated elastic anisotropy in both the L/R and L/C planes, but was able to predict orthotropy. Therefore, the results of this study suggest that the dominant and less variable transverse isotropy of human cortical bone, reflected by L/C, is governed primarily by apatite crystal orientations, while the more subtle and variable orthotropy, reflected by the difference between L/R and L/C, is governed primarily by intracortical porosity. Moreover, the combined model may be useful to investigate other structure-function relationships or in place of current numerical models, for example, in the study of bone adaptation and metabolic bone disease.

Sporicidal efficacy of genipin: a potential theoretical alternative for biomaterial and tissue graft sterilization

Terminal sterilization of musculoskeletal allografts by gamma radiation minimizes the risk of disease transmission but impairs allograft mechanical properties. Commonly employed crosslinking agents can sterilize tissues without affecting mechanical properties adversely; however, these agents are toxic. Genipin is reported to be a benign crosslinking agent that strengthens mechanical properties of tissues; however, the antimicrobial capacity of genipin is largely unknown. The present study's aims were: (1) to assess the sporicidal potential of genipin, (2) to improve antimicrobial capacity by changing chemical and physical treatment conditions. To establish genipin's sterilization potential Bacillus subtilis var. niger spore strips were treated with 0-10% genipin in PBS or in 1:1 DMSO:PBS up to 72 h at room temperature (RT). Sterilizing doses and concentrations of genipin were used to treat B. pumilus and Geobacillus stearothermophilus spores to assess broader spectrum sporicidal activity of genipin. Scanning electron microscopy (SEM) was performed to evaluate gross morphological changes after genipin treatment. Optimal sterilization conditions were determined by evaluating the effects of temperature (RT-50 °C), DMSO:PBS ratio (0:100-100:0), and treatment duration (24-72 h) on B. subtilis. Genipin penetration of full thickness bovine patellar tendon and cortical bone specimens was observed to assess the feasibility of the agent for treating grafts. Initial studies showed that after 72 h of treatment at RT with 0.63-10% genipin/DMSO:PBS B. subtilis spore strips were sterilized; 0.63% genipin/PBS did not sterilize spore strips at 72 h at RT. Genipin doses and concentrations that sterilized B. subtilis spore strips sterilized B. pumilus and G. stearothermophilus spore strips. SEM revealed no gross morphological differences between untreated and treated spores. Treatment optimization resulted in sterilization within 24 h with 100% PBS, and DMSO facilitated sporicidal activity. Genipin penetrated full thickness patellar tendon specimens and 3.72 ± 0.58 mm in cortical bone specimens. Genipin sterilizes B. subtilis, B. pumilus, and G. stearothermophilus spore strips. It penetrates soft and hard tissues at doses previously shown to be non-toxic and to improve mechanical strength in collagen-rich soft tissues. Further studies are indicated to assess genipin's effects on the mechanical properties of genipin-sterilized grafts, the ability of genipin to eradicate infectious species other than spores, and to assess whether sterilant activity persists after penetrating tissues and biomaterials.

In situ mechanical behavior of mineral crystals in human cortical bone under compressive load using synchrotron X-ray scattering techniques

It is of great interest to delineate the effect of orientation distribution of mineral crystals on the bulk mechanical behavior of bone. Using a unique synergistic approach combining a progressive loading scheme and synchrotron X-ray scattering techniques, human cortical bone specimens were tested in compression to examine the in situ mechanical behavior of mineral crystals aligned in different orientations. The orientation distribution was quantitatively estimated by measuring the X-ray diffraction intensity from the (002) plane in mineral crystals. In addition, the average longitudinal (c-axis), transverse (a-axis), and shear strains of the subset of mineral crystals aligned in each orientation were determined by measuring the lattice deformation normal to three distinct crystallographic planes (i.e. 002, 310, and 213) in the crystals. The experimental results indicated that the in situ strain and stress of mineral crystals varied with orientations. The normal strain and stress in the longitudinally aligned mineral crystals were markedly greater than those in the transversely oriented crystals, whereas the shear stress reached a maximum for the crystals aligned in ±30° with respect to the loading direction. The maximum principal strain and stress were observed in the mineral crystals oriented along the loading axis, with a similar trend observed in the maximum shear strain and stress. By examining the in situ behavior, the contribution of mineral crystals to load bearing and the bulk behavior of bone are discussed.

Cathepsin K Preferentially Solubilizes Matured Bone Matrix

Bone collagen undergoes a series of enzymatic and nonenzymatic posttranslational modifications with maturation. The aim of this study was to analyze the collagenolytic efficiency of cathepsin K in relation to the extent of bone collagen age. Bone collagen posttranslational maturation was induced in vitro by preincubating bovine fetal cortical bone specimens at 37°C for different times. The collagen enzymatic cross-links pyridinoline (PYD) and deoxypyridinoline (DPD), the advanced glycation end product pentosidine (PEN), and the native (α) and β-isomerized C-telopeptide (CTX) isomers were measured in each bone specimen. After extraction, bone collagen was incubated with human recombinant cathepsin K at different concentrations and its collagenolytic activity was measured by the release of hydroxyproline. To assess the affinity of cathepsin K for isomerized and nonisomerized CTX isomers, incubation with cathepsin K was also performed in the presence of various concentrations of a specific inhibitor. We showed that preincubation of bone collagen at 37°C induces a marked increase in the bone concentration of PYD, DPD, and PEN and of CTX isomerization as reflected by the ratio of α-/βCTX. This increase was associated with a parallel increase in the efficiency of cathepsin K to solubilize bone collagen. When cathepsin K was incubated in the presence of an inhibitor, the β-isomerized form of collagen from 3-month- and 8-year-old bovine bone was more susceptible to degradation than the native α form. These results suggest that the collagenolytic activity of cathepsin K may be increased toward more matured bone collagen.

Nonlinear resonant ultrasound spectroscopy is sensitive to the level of cortical bone damage

The objective of the study was to evaluate the sensitivity of nonlinear resonant ultrasound spectroscopy (NRUS) measurements to the accumulation of damage in cortical bone by fatigue or by controlled crack propagation. Two groups of human cortical bone specimens were prepared from the femoral mid-diaphysis. The specimens from the first group were taken through a progressive fatigue protocol consisting of four steps of cyclic four-point bending. The specimens from the second group were taken through a toughness protocol consisting of initiation and controlled propagation of a stable crack induced by 4-point bending mechanical loading. Our results evidenced a progressive increase of the normalized nonlinear elastic parameter during fatigue testing or during toughness experiments. While in specimens subjected to mechanical fatigue cycling the relative variation of nonlinear elasticity was significantly related to the relative variation of the number density of small cracks assessed with micro-computed tomogr...

Anatomical distribution of the degree of mineralization of bone tissue in human femoral neck: Impact on biomechanical properties

Osteoporotic hip fractures represent a major public health problem associated with high human and economic costs. The anatomical variation of the tissue mineral density (TMD) and of the elastic constants in femoral neck cortical bone specimens is an important determinant of bone fragility. The purpose of this study was to show that a Synchrotron radiation microcomputed tomography system coupled with a multiscale biomechanical model allows the determination of the 3-D anatomical dependence of TMD and of the elastic constants (i.e. the mechanical properties of an anisotropic material) in human femoral neck.Bone specimens from the inferior femoral neck were obtained from 18 patients undergoing standard hemiarthroplasty. The specimens were imaged using 3-D synchrotron micro-computed tomography with a voxel size of 10.13μm, leading to the determination of the anatomical distributions of porosity and TMD. The elastic properties of bone tissue were computed using a multiscale model. The model uses the experimental data obtained at the scale of several micrometers to estimate the components of the elastic tensor of bone at the scale of the organ.Statistical analysis (ANOVA) revealed a significant effect of the radial position on porosity and TMD and a significant effect of axial position on TMD only. Porosity was found to increase in the radial direction moving from the periosteum inwards (p<10−5). At any given distance from the periosteum, porosity does not vary noticeably along the bone axis. TMD was found to be significantly higher (p<10−5) in the periosteal region than in other bone locations and decreases from the periosteal to the endosteal region with an average slope of 10.05g.cm−3.m−1, the decrease being faster in the porous part of the samples (average slope equal of 30.04g.cm−3.m−1) than in dense cortical bone. TMD was found to decrease from the distal to the proximal part of the femur neck (average slope of 6.5g.cm−3.m−1). Considering TMD variations in the radial direction induces weak changes of bone properties compared to constant TMD. TMD variations in the axial direction are responsible for a significant variation of elastic constants.These results demonstrate that the anatomical variations of TMD affect the bone elastic properties, which could be explained by the complex stress field in bone affecting bone remodeling. TMD spatial variations should be taken into account to properly describe the spatial heterogeneity of elastic coefficients of bone tissue at the organ scale.

In Vitro Laser Bonding of Bovine Cortical Bone Specimen and TCP-Glass Ceramics

Orthopedic implants are widely used to fix a bone or to replace the articulating surface of a joint. Implant surgeries are performed only by highly specialized and trained surgeons. It is quite important to develop a technique that can bond bone and implant materials directly. Laser processing with high energy density is an effective tool for bonding different materials in a very short time with a high positional accuracy. In this study, to investigate the possibility of laser bonding living bones and bioceramics, an in-vitro laser processing technique was developed for bonding a bovine cortical bone specimen with a ceramics composed of synthetic tricalcium phosphate (TCP) and MgO-Al2O3-SiO2-glass (TG ceramics). By irradiating the surface of the bone specimen with a laser beam, a microporous foam-like substance was generated that bonded the bone specimen with the TG ceramics in a manner similar to spot welding. The bonding strength was defined as the shear stress between the interface of the bone specimen and the ceramics specimen. It was found that the strength of the bond increased with an increase in the duration of laser irradiation.

B107 脳卒中易発症ラットより摘出した皮質骨の生体力学的特性(B1-2 骨のバイオメカニクス)

B107 脳卒中易発症ラットより摘出した皮質骨の生体力学的特性(B1-2 骨のバイオメカニクス)

Change in porosity is the major determinant of the variation of cortical bone elasticity at the millimeter scale in aged women

At the mesoscale (i.e. over a few millimeters), cortical bone can be described as two-phase composite material consisting of pores and a dense mineralized matrix. The cortical porosity is known to influence the mesoscopic elasticity. Our objective was to determine whether the variations of porosity are sufficient to predict the variations of bone mesoscopic anisotropic elasticity or if change in bone matrix elasticity is an important factor to consider. We measured 21 cortical bone specimens prepared from the mid-diaphysis of 10 women donors (aged from 66 to 98 years). A 50-MHz scanning acoustic microscope (SAM) was used to evaluate the bone matrix elasticity (reflected in impedance values) and porosity. Porosity evaluation with SAM was validated against Synchrotron Radiation μCT measurements. A standard contact ultrasonic method was applied to determine the mesoscopic elastic coefficients. Only matrix impedance in the direction of the bone axis correlated to mesoscale elasticity (adjusted R(2)=[0.16-0.25], p<0.05). The mesoscopic elasticity was found to be highly correlated to the cortical porosity (adj-R(2)=[0.72-0.84], p<10(-5)). Multivariate analysis including both matrix impedance and porosity did not provide a better statistical model of mesoscopic elasticity variations. Our results indicate that, for the elderly population, the elastic properties of the mineralized matrix do not undergo large variations among different samples, as reflected in the low coefficients of variation of matrix impedance (less than 6%). This work suggests that change in the intracortical porosity accounts for most of the variations of mesoscopic elasticity, at least when the analyzed porosity range is large (3-27% in this study). The trend in the variation of mesoscale elasticity with porosity is consistent with the predictions of a micromechanical model consisting of an anisotropic matrix pervaded by cylindrical pores.

Detection of fatigue microdamage in whole rat femora using contrast-enhanced micro-computed tomography

Microdamage in bone tissue is typically studied using destructive, two-dimensional histological techniques. Contrast-enhanced micro-computed tomography (micro-CT) was recently demonstrated to enable non-destructive, three-dimensional (3-D) detection of microdamage in machined cortical and trabecular bone specimens in vitro. However, the accumulation of microdamage in whole bones is influenced by variations in the magnitude and mode of loading due to the complex whole bone morphology. Therefore, the objective of this study was to detect the presence, spatial location, and accumulation of fatigue microdamage in whole rat femora in vitro using micro-CT with a BaSO4 contrast agent. Microdamage was detected and observed to accumulate at specific spatial locations within the cortex of femora loaded in cyclic three-point bending to a 5% or 10% reduction in secant modulus. The ratio of the segmented BaSO4 stain volume (SV) to the total volume (TV) of cortical bone was adopted as a measure of damage. The amount of microdamage measured by micro-CT (SV/TV) was significantly greater for both loaded groups compared to the control group (p<0.05), but the difference between loaded groups was not statistically significant. At least one distinct region of microdamage, as indicated by the segmented SV, was observed in 85% of loaded specimens. A specimen-specific finite element model confirmed elevated tensile principal strains localized in regions of tissue corresponding to the accumulated microdamage. These regions were not always located where one might expect a priori based upon Euler–Bernoulli beam theory, demonstrating the utility of contrast-enhanced micro-CT for non-destructive, 3-D detection of fatigue microdamage in whole bones in vitro.

Effects of glucocorticoid on BMD, micro-architecture and biomechanics of cancellous and cortical bone mass in OVX rabbits

The incidence of osteoporosis continues to increase with progressively aging populations. The purpose of this study was to detect the effects of glucocorticoid (GC) treatment on bone mineral density (BMD), biomechanical strength and micro-architecture in cancellous and cortical bone in ovariectomized (OVX) rabbits. Twenty adult female New Zealand white rabbits were randomly divided into three groups. The OVX-GC group ( n = 8) received a bilateral ovariectomy first and then daily GC treatment (methylprednisolone sodium succinate, 1 mg/kg/day) for 4 weeks beginning 2 weeks after ovariectomy treatment. The OVX group ( n = 4) received a bilateral ovariectomy without GC treatment. The sham group ( n = 8) only received the sham operation. BMD was determined prior to and 6 weeks after the operation in the spine. Six weeks after the operation, the animals were sacrificed, and cancellous bone specimens were harvested from the femoral condyle and lumbar vertebrae. Cortical bone specimens were obtained from the femoral midshaft. The femoral specimens were scanned for apparent BMD. All specimens were tested mechanically and analyzed by microcompute tomography (micro-CT). In cancellous bone, GC treatment resulted in significant decreases in BMD, bone biomechanical strength and micro-architecture parameters in lumbar vertebrae. Similar trends in BMD and micro-architectural changes were also observed in the femoral condyle in the OVX-GC group compared with the sham group. However, there was no significant decline in any parameter in either lumbar vertebrae or femoral condyle in the OVX group. Similarly, no significant difference was found in any parameter in cortical bone among the three groups. Thus, the 4-week GC treatment in OVX rabbits could result in a significant bone loss in cancellous bone but not in cortical bone. This model is comparable to the osteoporosis-related changes in humans. OVX alone was not sufficient to induce osteoporosis.

Visualization of 3D osteon morphology by synchrotron radiation micro‐CT

Cortical bone histology has been the subject of scientific inquiry since the advent of the earliest microscopes. Histology - literally the study of tissue - is a field nearly synonymous with 2D thin sections. That said, progressive developments in high-resolution X-ray imaging are enabling 3D visualization to reach ever smaller structures. Micro-computed tomography (micro-CT), employing conventional X-ray sources, has become the gold standard for 3D analysis of trabecular bone and is capable of detecting the structure of vascular (osteonal) porosity in cortical bone. To date, however, direct 3D visualization of secondary osteons has eluded micro-CT based upon absorption-derived contrast. Synchrotron radiation micro-CT, through greater image quality, resolution and alternative contrast mechanisms (e.g. phase contrast), holds great potential for non-destructive 3D visualization of secondary osteons. Our objective was to demonstrate this potential and to discuss areas of bone research that can be advanced through the application of this approach. We imaged human mid-femoral cortical bone specimens derived from a 20-year-old male (Melbourne Femur Collection) at the Advanced Photon Source synchrotron (Chicago, IL, USA) using the 2BM beam line. A 60-mm distance between the target and the detector was employed to enhance visualization of internal structures through propagation phase contrast. Scan times were 1 h and images were acquired with 1.4-μm nominal isotropic resolution. Computer-aided manual segmentation and volumetric 3D rendering were employed to visualize secondary osteons and porous structures, respectively. Osteonal borders were evident via two contrast mechanisms. First, relatively new (hypomineralized) osteons were evident due to differences in X-ray attenuation relative to the surrounding bone. Second, osteon boundaries (cement lines) were delineated by phase contrast. Phase contrast also enabled the detection of soft tissue remnants within the vascular pores. The ability to discern osteon boundaries in conjunction with vascular and cellular porosity revealed a number of secondary osteon morphologies and provided a unique 3D perspective of the superimposition of secondary osteons on existing structures. Improvements in resolution and optimization of the propagation phase contrast promise to provide further improvements in structural detail in the future.

Near-terminal creep damage does not substantially influence fatigue life under physiological loading

Cortical bone specimens were damaged using repeated blocks of tensile creep loading until a near-terminal amount of creep damage was generated (corresponding to a reduction in elastic modulus of 15%). One group of cortical bone specimens was submitted to the near-terminal damage protocol and subsequently underwent fatigue loading in tension with a maximum strain of 2000 με (Damage Fatigue, n=5). A second group was submitted to cyclic fatigue loading but was not pre-damaged (Control Fatigue, n=5). All but one specimen (a damaged specimen) reached run-out (10 million cycles, 7.7 days). No significant differences in microscopic cracks or other tissue damage were observed between the two groups or between either group and additional, completely unloaded specimens. Our results suggest that damage in cortical bone allograft that is not obvious or associated with a stress riser may not substantially affect its fatigue life under physiologic loading.

Anatomic variation in the elastic inhomogeneity and anisotropy of human femoral cortical bone tissue is consistent across multiple donors

Numerical models commonly account for elastic inhomogeneity in cortical bone using power-law scaling relationships with various measures of tissue density, but limited experimental data exists for anatomic variation in elastic anisotropy. A recent study revealed anatomic variation in the magnitude and anisotropy of elastic constants along the entire femoral diaphysis of a single human femur (Espinoza Orías et al., 2009). The objective of this study was to confirm these trends across multiple donors while also considering possible confounding effects of the anatomic quadrant, apparent tissue density, donor age, and gender. Cortical bone specimens were sampled from the whole femora of 9 human donors at 20%, 50%, and 80% of the total femur length. Elastic constants from the main diagonal of the reduced fourth-order tensor were measured on hydrated specimens using ultrasonic wave propagation. The tissue exhibited orthotropy overall and at each location along the length of the diaphysis (p<0.0001). Elastic anisotropy increased from the mid-diaphysis toward the epiphyses (p<0.05). The increased elastic anisotropy was primarily caused by a decreased radial elastic constant (C11) from the mid-diaphysis toward the epiphyses (p<0.05), since differences in the circumferential (C22) and longitudinal (C33) elastic constants were not statistically significant (p>0.29). Anatomic variation in intracortical porosity may account for these trends, but requires further investigation. The apparent tissue density was positively correlated with the magnitude of each elastic constant (p<0.0001, R2>0.46), as expected, but was only weakly correlated with C33/C11 (p<0.05, R2=0.04) and not significantly correlated with C33/C22 and C11/C22.

In situ accumulation of advanced glycation endproducts (AGEs) in bone matrix and its correlation with osteoclastic bone resorption

Advanced glycation end products (AGEs) have been observed to accumulate in bone with increasing age and may impose effects on bone resorption activities. However, the underlying mechanism of AGEs accumulation in bone is still poorly understood. In this study, human cortical bone specimens from young (31±6years old), middle-aged (51±3years old) and elderly (76±4years old) groups were examined to determine the spatial-temporal distribution of AGEs in bone matrix and its effect on bone resorption activities by directly culturing osteoclastic cells on bone slices. The results of this study indicated that the fluorescence intensity (excitation wave length 360nm and emission wave length 470±40nm) could be used to estimate the relative distribution of AGEs in bone (pentosidine as its marker) under an epifluorescence microscope. Using the fluorescence intensity as the relative measure of AGEs concentration, it was found that the concentration of AGEs varied with biological tissue ages, showing the greatest amount in the interstitial tissue, followed by the old osteons, and the least amount in newly formed osteons. In addition, AGEs accumulation was found to be dependent on donor ages, suggesting that the younger the donor the less AGEs were accumulated in the tissue. Most interestingly, AGEs accumulation appeared to initiate from the region of cement lines, and spread diffusively to the other parts as the tissue aged. Finally, it was observed that the bone resorption activities of osteoclasts were positively correlated with the in situ concentration of AGEs and such an effect was enhanced with increasing donor age. These findings may help elucidate the mechanism of AGEs accumulation in bone and its association with bone remodeling process.