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Understanding the relationship between the microstructure and mechanical function of trabecular bone is critical for prediction and prevention of bone fragility fractures. However, a detailed understanding of the structural design of trabecular microarchitecture is still missing. This study hypothesized that there exists a commonality in the underlying probabilistic distributions of microstructural features of trabecular bones, whereas the microstructural differences among individuals are primarily describe by a set of scalar parameters. To test the hypothesis, twenty-three trabecular bone specimens were obtained from two anatomic locations (i.e., femoral neck and vertebral body) and a diverse group of seventeen donors of different age and sex. The number, size, spatial location, and orientation of individual plates and rods in the trabecular bone specimens were determined via volumetric decomposition of 3D μCT images using the Individual Trabecula Segmentation (ITS) technique. Then, m/n bootstrap Kolmogorov-Smirnov tests were performed to compare the normalized distributions of size, orientation, and spatial arrangement of trabecular plates and rods in the specimens. The results showed that 100% of the twenty-three normalized distributions of each microstructural feature were statistically equivalent irrespective of individual differences among the bone specimens, except the distributions of rod spatial arrangement (100%). On the other hand, nonparametric Mann-Whitney U tests showed that a set of scalar parameters (i.e., the number, average size, and average nearest neighbor distance of trabecular plates and rods) were statistically different among the individual specimens (p0.05). Due to the commonality of the underlying distributions, the individual differences in the trabecular microstructure among the specimens seemed to be reflected primarily by changes in the scalar parameters. The above results strongly support the hypothesis of this study and may shed more light on understanding the natural design of trabecular bone microstructures.

It is well known that loss of bone mass, quantified by areal bone mineral density (aBMD) using DXA, is associated with the increasing risk of bone fractures. However, bone mineral density alone cannot fully explain changes in fracture risks. On top of bone mass, bone architecture has been identified as another key contributor to fracture risk. In this study, we used a novel stochastic approach to assess the distribution of aBMD from 2D projection images of Micro-CT scans of trabecular bone specimens at a resolution comparable to DXA images. Sill variance, a stochastic measure of distribution of aBMD, had significant relationships with microarchitecture parameters of trabecular bone, including bone volume fraction, bone surface-to-volume ratio, trabecular thickness, trabecular number, trabecular separation and anisotropy. Accordingly, it showed significantly positive correlations with strength and elastic modulus of trabecular bone. Moreover, a combination of aBMD and sill variance derived from the 2D projection images (R2=0.85) predicted bone strength better than using aBMD alone (R2=0.63). Thus, it would be promising to extend the stochastic approach to routine DXA scans to assess the distribution of aBMD, offering a more clinically significant technique for predicting risks of bone fragility fractures.

Bone quality is significantly correlated with the inhomogeneous distribution of material and ultrastructural properties (e.g., modulus and mineralization) of the tissue. Current techniques for quantifying inhomogeneity consist of descriptive statistics such as mean, standard deviation and coefficient of variation. However, these parameters do not describe the spatial variations of bone properties. The objective of this study was to develop a novel statistical method to characterize and quantitatively describe the spatial variation of bone properties at ultrastructural levels. To do so, a random field defined by an exponential covariance function was used to represent the spatial uncertainty of elastic modulus by delineating the correlation of the modulus at different locations in bone lamellae. The correlation length, a characteristic parameter of the covariance function, was employed to estimate the fluctuation of the elastic modulus in the random field. Using this approach, two distribution maps of the elastic modulus within bone lamellae were generated using simulation and compared with those obtained experimentally by a combination of atomic force microscopy and nanoindentation techniques. The simulation-generated maps of elastic modulus were in close agreement with the experimental ones, thus validating the random field approach in defining the inhomogeneity of elastic modulus in lamellae of bone. Indeed, generation of such random fields will facilitate multi-scale modeling of bone in more pragmatic details.

Previous studies have postulated that ultrastructural changes may alter the pattern and capacity of microdamage accumulation in bone. Using an osteogenesis imperfecta (OI) mouse model, this study was performed to investigate the correlation of collagen mutation with the microdamage morphology and the associated brittleness of bone. In this study, femurs from mild OI and wild type mice were fatigued under four-point bending to create microdamage in the specimens. Then, the microdamage morphology of these specimens was examined using the bulk-staining technique with basic fuchsin. Similar with the results of previous studies, it was observed that linear microcracks were formed more easily in compression, whereas diffuse damage was induced more readily in tension for both wild-type and mild-type mice. However, less diffuse damage was found in the tensile side of mild OI mouse femurs (collagen mutation) compared with those of wild type mice, showing that the microdamage morphology is correlated to the brittleness of bone. The results of this study provide direct evidence that supports the prediction made by the previous numerical simulation studies, suggesting that microdamage morphology in bone is significantly correlated with the integrity of the collagen phase.

Since clinical measures of bone mineral density do not necessarily predict whether a person will fracture a bone without an intervention, there is a need to find supplementary tools for assessing bone quality. Presently, we hypothesized that measures of mobile and bound water by a Nuclear Magnetic Resonance (NMR) technique are correlated with bone strength and toughness, respectively. To test this, bending specimens from the mid-diaphysis of 18 human femurs were collected from 18 male donors and divided into middle aged and elderly groups. After NMR measurements of each hydrated specimen, an inversion technique was used to convert the free induction decay data into a distribution of spin-spin (T2) relaxation rates. Then, the distribution resolved into three distinct components that likely represent solid hydrogen, water bound to bone tissue, and mobile water that occupy microscopic pores within the bone specimen. The integrated signal intensities of the bound and mobile components were normalized by the wet mass of the specimen. Following NMR measurements, three point bending tests were conducted to determine the modulus of elasticity, flexure strength, and work to fracture of each specimen. Next, the porosity, mineral-to-collagen ratio, and pentosidine concentration were measured. In this sample of human cortical bone, there was no age-related difference in the amount of mobile water, but the decrease in the amount of bound water with increasing age was statistically significant. Moreover, bound water was associated with both strength and work to fracture of bone, while mobile water was correlated with modulus of elasticity and appeared to quantify the level of microscopic pores within bone. On the other hand, bound water was correlated with the concentration of non-enzymatic collagen cross-links. The results of this study indicate that quantifying mobile and bound water with magnetic resonance techniques could potentially serve as indicators of bone quality.

Collagen crosslinks are important to the quality of bone and may be contributors to the age-related increase in bone fracture. This study was performed to investigate whether age and gender effects on collagen crosslinks are similar in osteonal and interstitial bone tissues. Forty human cadaveric femurs were collected and divided into two age groups: Middle aged (42–63 years of age) and Elderly (69–90 years of age) with ten males and ten females in each group (n = 10). Micro-cores of bone tissue from both secondary osteons (newly formed) and interstitial regions (biologically old) in the medial quadrant of the diaphysis were extracted using a custom-modified, computer numerical controlled machine. The bone specimens were then analyzed using high performance liquid chromatography to determine the effects of age and gender on the concentration of mature, enzymatic crosslinks (hydroxylysyl-pyridinoline – HP and lysylpyridinoline – LP) and a non-enzymatic crosslink (pentosidine – PE) at these two bony sites. The results indicate that age has a significant effect on the concentration of LP and PE, while gender has a significant effect on HP and LP. In addition, the concentration of the crosslinks in the secondary osteons is significantly different from that in the interstitial bone regions. These results suggest that the rate of non-enzymatic crosslinking may increase while the formation of maturate enzymatic crosslinks may decrease with age. Such changes could potentially reduce the inherent quality of the bone tissue in the elderly skeleton.

The hypothesis of this study is that the mechanical integrity of the collagen network in bone deteriorates with age, and such adverse changes correlate with the decreased toughness of aged bone. To test the hypothesis, 30 human cadaveric femurs from donors ranging from 19 to 89 years of age were tested to determine the age-related changes in the mechanical properties of demineralized bone and fresh bone samples. Along with bone porosity, bone density, and weight fractions of the mineral and organic phases, collagen denaturation and concentrations of collagen cross-links (HP, hydroxylysylpyridinoline; LP, lysylpyridinoline; PE, pentosidine) were determined for these bone specimens as a function age. Analysis of variance (ANOVA) showed that age-dependent changes were reflected in the decreased strength, work to fracture, and fracture toughness of bone; in the decreased strength, elastic modulus, and work to fracture of the collagen network; as well as in the increased concentration of pentosidine (a marker of nonenzymatic glycation) and increased bone porosity. Regression analyses of the measured parameters showed that the age-related decrease in work to fracture of bone (especially its postyield portion) correlated significantly with deterioration in the mechanical integrity of the collagen network. The results of this study indicate that the adverse changes in the collagen network occur as people age and such changes may lead to the decreased toughness of bone. Also, the results suggest that nonenzymatic glycation may be an important contributing factor causing changes in collagen and, consequently, leading to the age-related deterioration of bone quality.

Age-related changes in the skeleton often lead to an increase in the susceptibility of bone to fracture. Such changes most likely occur in the constituents of bone, namely, the mineral and organic phases, and in their spatial arrangement manifested as orientation and microstructure. In the past, however, bone loss or decline in bone mineral density has been considered to be the major contributing factor for the increased risk of bone fractures, and elastic modulus and ultimate strength have been commonly used to assess bone quality and strength. However, whether these properties provide sufficient information regarding the likelihood of bone to fracture remains debatable. Using a novel fracture toughness test, which measures the energy or stress intensity required to propagate a crack within a material, the objective of this study was to investigate if the mineral density and mechanical properties of bone can accurately predict bone fragility as measured by fracture toughness. Changes in fracture toughness ( K IC ), bone mineral density (BMD), elastic modulus ( E ), yield and ultimate strength (σ y and σ s ), porosity ( P 0 ), and microhardness ( H v ) of bone were examined as a function of age in a baboon model. With increasing age, the fracture toughness of bone decreased, and its microhardness increased. However, no significant changes were found in BMD, E , P 0 , σ y , and σ s as a function of age. In addition, simple regression analyses revealed no significant correlation between bone fracture toughness and the other parameters, except for microhardness of bone. The results of this study indicate that changes in bone fracture toughness may not be necessarily reflected in its mineral density, porosity, elastic modulus, yield strength, and ultimate strength.

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±6 years old), middle-aged (51±3 years old) and elderly (76±4 years 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 360 nm and emission wave length 470±40 nm) 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.

Correction to the article “Age-Related Changes in the Collagen Network and Toughness of Bone” [Bone 31:1–7, 2002] X. Wang, X. Shen, X. Li, and C. Mauli Agrawal The elastic portion of the work to fracture of bone Wfe is misprinted as Wfc in Table 1 and in the corresponding text. In addition, the work to fracture Wf is misprinted as W in the footnote to Table 1. R Available online at www.sciencedirect.com