Three-dimensional simulation of ultrasound propagation through trabecular bone structures measured by synchrotron microtomography

Three-dimensional numerical simulations of ultrasound transmission were performed through 31 trabecular bone samples measured by synchrotron microtomography. The synchrotron microtomography provided high resolution 3D mappings of bone structures, which were used as the input geometry in the simulation software developed in our laboratory. While absorption (i.e. the absorption of ultrasound through dissipative mechanisms) was not taken into account in the algorithm, the simulations reproduced major phenomena observed in real through-transmission experiments in trabecular bone. The simulated attenuation (i.e. the decrease of the transmitted ultrasonic energy) varies linearly with frequency in the MHz frequency range. Both the speed of sound (SOS) and the slope of the normalized frequency-dependent attenuation (nBUA) increase with the bone volume fraction. Twenty-five out of the thirty-one samples exhibited negative velocity dispersion. One sample was rotated to align the main orientation of the trabecular structure with the direction of ultrasonic propagation, leading to the observation of a fast and a slow wave. Coupling numerical simulation with real bone architecture therefore provides a powerful tool to investigate the physics of ultrasound propagation in trabecular structures. As an illustration, comparison between results obtained on bone modelled either as a fluid or a solid structure suggested the major role of mode conversion of the incident acoustic wave to shear waves in bone to explain the large contribution of scattering to the overall attenuation.     

X-ray diffraction studies of tibial plateau cartilage using synchrotron radiation

X-ray diffraction studies of articular cartilage have been performed using X-rays from two sources: a conventional generator and synchrotron radiation. Blocks of tissue from the tibial plateaux of pigs and dogs were examined to find the orientations of collagen fibrils throughout their depth. Analysis of the resulting diffraction patterns revealed a zonal arrangement of fibrils in both pig and dog tissue. At the articular surface the preferred orientation of fibrils was parallel to the surface, below this was a zone with no obvious preferred orientation and the remaining tissue had fibrils oriented mainly perpendicular to the surface. Using a conventional source of X-rays the exposure time for a single diffraction pattern was a number of days but this was reduced to about 10 min using the more intense synchrotron radiation. This improvement in exposure time facilitates the examination of small areas of tissue. X-ray diffraction using the synchrotron source offers a quick method of determining collagen fibril orientations in small areas of articular cartilage.     

Lattice strains and load partitioning in bovine trabecular bone

Microdamage and failure mechanisms have been well characterized in bovine trabecular bone. However, little is known about how elastic strains develop in the apatite crystals of the trabecular struts and their relationship with different deformation mechanisms. In this study, wide-angle high-energy synchrotron X-ray diffraction has been used to determine bulk elastic strains under in situ compression. Dehydrated bone is compared to hydrated bone in terms of their response to load. During compression, load is initially borne by trabeculae aligned parallel to loading direction with non-parallel trabeculae deforming by bending. Ineffective load partitioning is noted in dehydrated bone whereas hydrated bone behaves like a plastically yielding foam.     

A synchrotron radiation microtomography system for the analysis of trabecular bone samples.

X-ray computed microtomography is particularly well suited for studying trabecular bone architecture, which requires three-dimensional (3-D) images with high spatial resolution. For this purpose, we describe a three-dimensional computed microtomography (microCT) system using synchrotron radiation, developed at ESRF. Since synchrotron radiation provides a monochromatic and high photon flux x-ray beam, it allows high resolution and a high signal-to-noise ratio imaging. The principle of the system is based on truly three-dimensional parallel tomographic acquisition. It uses a two-dimensional (2-D) CCD-based detector to record 2-D radiographs of the transmitted beam through the sample under different angles of view. The 3-D tomographic reconstruction, performed by an exact 3-D filtered backprojection algorithm, yields 3-D images with cubic voxels. The spatial resolution of the detector was experimentally measured. For the application to bone investigation, the voxel size was set to 6.65 microm, and the experimental spatial resolution was found to be 11 microm. The reconstructed linear attenuation coefficient was calibrated from hydroxyapatite phantoms. Image processing tools are being developed to extract structural parameters quantifying trabecular bone architecture from the 3-D microCT images. First results on human trabecular bone samples are presented.     

Comparison of trabecular bone anisotropies based on fractal dimensions and mean intercept length determined by principal axes of inertia

The mechanical quality of trabecular bone depends on both its stiffness and its strength characteristics, which can be predicted indirectly by the combination of bone volume fraction and architectural anisotropy. To analyze the directional anisotropy of the trabecular bone, we applied the fractal geometry technique to plain radiographs. The anisotropy of the bone was quantified from an ellipse, based on the directional fractal dimensions (FD), by the principal axes of inertia. The anisotropies based on the FD were compared with those determined using the common method of mean intercept length (MIL). The directional FD gave the fractal information obtained from a projection along the MIL orientation. For this reason, the spatial variations associated with the bone length in any direction were manifested in a related frequency band of the power spectrum determined along the direction. The directional FD and MIL plots were highly correlated, although they originated from quite different geometries. Of the angle, premolar, and incisor regions of the human mandible, the anisotropies calculated using both FD and MIL showed the highest correlation in the trabecular bone of the angle region. The method using directional FDs as determined by the principal axis of inertia measures the anisotropy directly, using two-dimensional plain radiographs. This kind of method will be a useful to provide better estimates of bone quality in vivo compared with the density measurements alone, especially for the indirect diagnosis of jawbone quality in dental clinics.     

Comparison of synchrotron radiation and conventional x-ray microcomputed tomography for assessing trabecular bone microarchitecture of human femoral heads

Microcomputed tomography (microCT) produces three-dimensional (3D) images of trabecular bone. We compared conventional microCT (CmicroCT) with a polychromatic x-ray cone beam to synchrotron radiation (SR) microCT with a monochromatic parallel beam for assessing trabecular bone microarchitecture of 14 subchondral femoral head specimens from patients with osteoarthritis (n=10) or osteoporosis (n=4). SRmicroCT images with a voxel size of 10.13 microm were reconstructed from 900 2D radiographic projections (angular step, 0.2 degrees). CmicroCT images with a voxel size of 10.77 microm were reconstructed from 205, 413, and 825 projections obtained using angular steps of 0.9 degrees, 0.45 degrees, and 0.23 degrees, respectively. A single threshold was used to binarize the images. We computed bone volume/ tissue volume (BV/TV), bone surface/bone volume (BS/BV), trabecular number (Tb.N), trabecular thickness (Tb.Th and Tb.Th*), trabecular spacing (Tb.Sp), degree of anisotropy (DA), and Euler density. With the 0.9 degrees angular step, all CmicroCT values were significantly different from SRmicroCT values. With the 0.23 degrees and 0.45 degrees rotation steps, BV/TV, Tb.Th, and BS/BV by CmicroCT differed significantly from the values by SRmicroCT. The error due to slice matching (visual site matching +/- 10 slices) was within 1% for most parameters. Compared to SRmicroCT, BV/TV, Tb.Sp, and Tb.Th by CmicroCT were underestimated, whereas Tb.N and Tb. Th* were overestimated. A Bland and Altman plot showed no bias for Tb.N or DA. Bias was -0.8 +/- 1.0%, +5.0 +/- 1.1 microm, -5.9 +/- 6.3 microm, and -5.7 +/- 29.1 microm for BV/TV, Tb.Th*, Tb.Th, and Tb.Sp, respectively, and the differences did not vary over the range of values. Although systematic differences were noted between SRmicroCT and CmicroCT values, correlations between the techniques were high and the differences would probably not change the discrimination between study groups. CmicroCT provides a reliable 3D assessment of human defatted bone when working at the 0.23 degrees or 0.45 degrees rotation step; the 0.9 degrees rotation step may be insufficiently accurate for morphological bone analysis.     

Screw insertion in trabecular bone causes peri-implant bone damage

Secure fracture fixation is still a major challenge in orthopedic surgery, especially in osteoporotic bone. While numerous studies have investigated the effect of implant loading on the peri-implant bone after screw insertion, less focus has been put on bone damage that may occur due to the screw insertion process itself. Therefore, the aim of this study was to localize and quantify peri-implant bone damage caused by screw insertion.We used non-invasive three-dimensional micro-computed tomography to scan twenty human femoral bone cores before and after screw insertion. After image registration of the pre- and post-insertion scans, changes in the bone micro-architecture were identified and quantified. This procedure was performed for screws with a small thread size of 0.3 mm (STS, N = 10) and large thread size of 0.6 mm (LTS, N = 10).Most bone damage occurred within a 0.3 mm radial distance of the screws. Further bone damage was observed up to 0.6 mm and 0.9 mm radial distance from the screw, for the STS and LTS groups, respectively. While a similar amount of bone damage was found within a 0.3 mm radial distance for the two screw groups, there was significantly more bone damage for the LTS group than the STS group in volumes of interest between 0.3–0.6 mm and 0.6–0.9 mm.In conclusion, this is the first study to localize and quantify peri-implant bone damage caused by screw insertion based on a non-invasive, three-dimensional, micro-CT imaging technique. We demonstrated that peri-implant bone damage already occurs during screw insertion. This should be taken into consideration to further improve primary implant stability, especially in low quality osteoporotic bone. We believe that this technique could be a promising method to assess more systematically the effect of peri-implant bone damage on primary implant stability. Furthermore, including peri-implant bone damage due to screw insertion into patient-specific in silico models of implant-bone systems could improve the accuracy of these models.     

Ultrasonic wave propagation in trabecular bone predicted by the stratified model

The objective of this study was to investigate ultrasound propagation in trabecular bone by considering the wave reflection and transmission in a multilayered medium. The use of ultrasound to identify those at risk of osteoporosis is a promising diagnostic method providing a measure of bone mineral density (BMD). A stratified model was proposed to study the effect of transmission and reflection of ultrasound wave within the trabecular architecture on the relationship between ultrasound and BMD. The results demonstrated that ultrasound velocity in trabecular bone was highly correlated with the bone apparent density (r=0.97). Moreover, a consistent pattern of the frequency dependence of ultrasound attenuation coefficient has been observed between simulation using this model and experimental measurement of trabecular bone. The normalized broadband ultrasound attenuation (nBUA) derived from the simulation results revealed that nBUA was nonlinear with respect to trabecular porosity and BMD. The curve of the relationship between nBUA and BMD was parabolic in shape, and the peak magnitude of nBUA was observed at 60% of bone porosity. These results agreed with the published experimental data and demonstrated that according to the stratified model, reflection and transmission were important factors in the ultrasonic propagation through the trabecular bone. © 2001 Biomedical Engineering Society. PAC01: 4380Vj, 4380Qf, 8763Df, 8710+e, 4325Ed, 8719-j     

Effect of high-energy X-ray doses on bone elastic properties and residual strains

Bone X-ray irradiation occurs during medical treatments, sterilization of allografts, space travel and in vitro studies. High doses are known to affect the post-yield properties of bone, but their effect on the bone elastic properties is unclear. The effect of such doses on the mineral-organic interface has also not been adequately addressed. Here, the evolution of elastic properties and residual strains with increasing synchrotron X-ray dose (5-3880 kGy) is examined on bovine cortical bone. It is found that these doses affect neither the degree of nanometer-level load transfer between the hydroxyapatite (HAP) platelets and the collagen up to stresses of -60 MPa nor the microscopic modulus of collagen fibrils (both measured by synchrotron X-ray scattering during repeated in situ loading and unloading). However, the residual elastic strains in the HAP phase decrease markedly with increased irradiation, indicating damage at the HAP-collagen interface. The HAP residual strain also decreases after repeated loading/unloading cycles. These observations can be explained by temporary de-bonding at the HAP/collagen interface (thus reducing the residual strain), followed by rapid re-bonding (so that load transfer capability is not affected).     

Investigation of the three-dimensional orientation of mineralized collagen fibrils in human lamellar bone using synchrotron X-ray phase nano-tomography

We investigate the three-dimensional (3-D) organization of mineralized collagen fibrils in human cortical bone based on synchrotron X-ray phase nano-tomography images. In lamellar bone the collagen fibrils are assumed to have a plywood-like arrangement, but due to experimental limitations the 3-D fibril structure has only been deduced from section surfaces so far and the findings have been controversial. Breakthroughs in synchrotron tomographic imaging have given access to direct 3-D information on the bone structure at the nanoscale level. Using an autocorrelation-based orientation measure we confirm that the fibrils are unidirectional in quasi-planes of sub-lamellae and find two specific dominant patterns, oscillating and twisted plywoods coexisting in a single osteon. Both patterns exhibit smooth orientation changes between adjacent quasi-planes. Moreover, we find that the periodic changes in collagen fibril orientation are independent of fluctuations in local mass density. These data improve our understanding of the lamellar arrangement in bone and allow more detailed investigations of structure–function relationships at this scale, providing templates for bio-inspired materials. The presented methodology can be applied to non-destructive 3-D characterization of the sub-micron scale structure of other natural and artificial mineralized biomaterials.     

An X-ray diffraction study of the effects of heat treatment on bone mineral microstructure

A series of human cortical bone specimens has been heated to temperatures up to 1200 degrees C and the mineral content examined in detail by X-ray diffraction. Line profile analysis of the diffraction data has been undertaken to characterise the microstructural (crystallite size and microstrain) features of the mineral at each temperature. Individual profile fitting of several maxima from each diffractogram has also provided precise lattice parameters of the apatite at each temperature. The apatite did not show any significant decomposition over the temperature range although CaO was increasingly formed at temperatures above 600 degrees C. Both finite crystallite size and microstrain contributed significantly to the diffraction peak broadening below 600 degrees C. When heated to > 800 degrees C, the small, rod-like mineral crystallites changed from a highly anisotropically strained state to one with significantly larger equidimensional crystals possessing little microstrain. The findings are discussed in the context of graft bone substitutes and surgical heating of bone.     

Novel assessment of hepatic iron distribution by synchrotron radiation X-ray fluorescence microscopy

Excess iron deposition in the liver is known to be hepatotoxic and may exacerbate liver injury. However, little is known about iron distribution in the lobule because of the lack of a highly sensitive detection method. The aim of this study is to determine iron distribution in the lobule of human liver by means of synchrotron radiation X-ray fluorescence (SRXRF) microscopy. Mapping of the trace elements was done with use of SRXRF microscopy and compared with the results of staining by Berlin blue and oxidative stress markers. Iron deposits were distributed predominantly in periportal hepatocytes in the normal liver in a decreasing gradient from the periportal area to the perivenous area. This distribution was consistent with the formation of oxidative stress markers, suggesting that hepatocytes in the periportal area may be predominantly primed by iron-induced free radical damage even in normal liver. On the other hand, iron deposits in the periportal area were more intense than those in the centrilobular area in both the liver with chronic hepatitis C and the cirrhotic liver. In conclusion, elemental mapping by SRXRF microscopy was a highly sensitive method for the detection and mapping of elements such as iron and copper in liver sections.     

Effects of spaceflight on trabecular bone in rats

Alterations in trabecular bone were observed in growing male Wistar rats after 18.5 days of orbital flight on the COSMOS 1129 biosatellite. Spaceflight induced a decreased mass of mineralized tissue and an increased fat content of the bone marrow in the proximal tibial and humeral metaphyses. The osteoblast population appeared to decline immediately adjacent to the growth cartilage-metaphyseal junction, but osteoclast numbers were unchanged. These results suggested that bone formation may have been inhibited during spaceflight, but resorption remained constant. With the exception of trabecular bone mass in the proximal tibia, the observed skeletal changes returned to normal during a 29-day postflight period.     

Analysis of trabecular bone texture by modified Hurst orientation transform method

There is a growing need for noninvasive and inexpensive methods that can effectively be used on a large scale, to detect an onset of early osteoarthritis in human knee joints. Of many possible options, fractal analysis of two-dimensional projection x-ray images of trabecular bone (TB) texture, appears as one of the best approaches. However, there are some problems associated with the characterization of the roughness and anisotropy of the bone texture. To resolve these problems, a modified Hurst orientation transform (HOT) method, previously developed by the authors, has been used in this study. The advantages of the HOT method over other techniques used to analyze bone texture, are that it calculates a two-dimensional fractal dimension in all possible directions and also provides a measure of anisotropy for both surfaces exhibiting strong anisotropy and surfaces exhibiting weak anisotropy. In this study, the accuracy of the HOT method in measuring the bone texture roughness and anisotropy; together with the effects of image noise, blur, exposure, magnification, and projection angle on its performance were investigated. Computer-generated images of fractal surfaces and x-ray images obtained for a human tibia head were used. Results obtained show that the HOT method can effectively be used to characterize the roughness and anisotropy (isotropy) of TB texture.     

Mechanical properties of human trabecular bone lamellae quantified by nanoindentation.

Improved preventive and therapeutic strategies for skeletal diseases such as osteoporosis rely on a better understanding of the mechanical properties of trabecular bone and their influence on cell mediated adaptation processes. The mechanical properties of trabecular bone are determined by composition as well as structural (trabecular architecture), microstructural (trabecular packets) and nanostructural (lamellae) organization. Density is the major predictor of the mechanical properties of trabecular structures and has been extended to the concept of fabric to include architectural anisotropy and improve even further the power of prediction. Recent advances in QCT and MRI technologies allow for precise assessment of 3D trabecular architecture and the mechanical consequences of structural changes can be increasingly well quantified by the means of computational methods. While single trabeculae have been tested using various techniques with contrasting results, little is known about the intrinsic mechanical properties of trabecular bone lamellae on which these computational methods rely. For instance, water and mineral content have a significant effect on the elastic, viscous, yield and postyield properties of bone tissue. In addition, collagen fiber orientation affects the mechanics of single remodeling units. Variations in composition and organization determined by age, accumulated damage or disease may therefore reduce the mechanical integrity of trabecular bone and deserve more attention. The aim of this work was to utilize a nanoindentation technique to quantify elastic modulus and hardness of human trabecular bone lamellae.