Effects of freeze–thaw and micro-computed tomography irradiation on structure–property relations of porcine trabecular bone

doi:10.1016/j.jbiomech.2014.02.022

Journal of Biomechanics

Volume 47, Issue 6, 11 April 2014, Pages 1495-1498

https://doi.org/10.1016/j.jbiomech.2014.02.022 Get rights and content

Abstract

We study the effects of freeze–thaw and irradiation on structure–property relations of trabecular bone. We measure the porosity, apparent density, mineral content, trabecular orientation, trabecular thickness, fractal dimension, surface area, and connectivity of trabecular bone using micro-computed tomography (micro-CT) and relate them to Young׳s modulus and ultimate strength measured by uniaxial compression testing. The analysis is done on six-month porcine trabecular bone from femoral heads. The effects of freeze–thaw are studied by using bones from three different groups: fresh bone and bones frozen for one and five years. We find that the porosity and apparent density have most dominant influence on the elastic modulus and strength of fresh bone. Also, five years of freezing lowers both Young׳s modulus and ultimate strength of trabecular bone. Additionally, the effects of radiation are investigated by comparing Young׳s modulus before and after micro-CT exposure. We find that the micro-CT irradiation has a negligible effect on the Young׳s modulus of trabecular bone. These findings provide insights on the effects of tissue preservation and imaging on properties of trabecular bone.

Introduction

Testing of freshly harvested bone is generally impractical (Huss et al., 1995). Previous research has shown that freeze–thaw does not affect mechanical properties of bone and thus is a popular method for storing bone (Panjabi et al., 1985). However, little is known about the effects of long term freezing on the properties of bone.

Knowledge of structure–property relations of trabecular bone is of importance for diagnosis and assessment of osteoporosis. Morphological measures of trabecular bone microarchitecture have been used, in addition to porosity and apparent density, to correlate with mechanical properties of trabecular bone (Hodgskinson and Currey, 1990a, Hodgskinson and Currey, 1990b, Goulet et al., 1994). Micro-computed tomography (micro-CT) has been employed to obtain such data (Muller, 2009, Burghardt et al., 2011). Effects of radiation on bone properties have been studied but mainly for sterilized bone (Barth et al., 2010, Barth et al., 2011, Singhal et al., 2011).

In this paper we study the effects of freeze–thaw and micro-CT irradiation on structure–property relations of porcine trabecular bone. We conduct this analysis using fresh bones, bones frozen for one and five years, and bones before and after micro-CT exposure. We measure the porosity, apparent density, mineral density, fractal dimension, surface area, orientation, thickness, and connectivity of trabeculae using micro-CT and relate them to Young׳s modulus and ultimate compressive strength obtained by compression testing. This study contributes to better understanding of the effects of tissue preservation and imaging on properties of trabecular bone.

Section snippets

Sample preparation

Femurs from six-month old pigs (Sus scrofa domestica) were obtained from the Meat Science Lab at the University of Illinois at Urbana-Champaign. Porcine bone was selected because its biology is similar to human bone (Pearce et al., 2007). All animals were healthy and raised under diets satisfying nutrient levels recommended by the Nutrient Requirements of Swine (2012). After harvesting, femurs were either tested fresh (group A) or stored in the freezer at −20 °C for one year (group B) or five

Results

Effects of freeze–thaw on Young׳s modulus and strength, studied using bone samples from all three groups (fresh bone, and bones frozen for one or five years), are summarized in Fig. 1 and Table 2. All three groups followed a normal distribution. According to Tukey׳s post hoc, the mean Young׳s modulus of fresh bone showed a significant difference compared to frozen bone. The mean of Young׳s modulus of fresh bone was significantly higher than the modulus from frozen bone. The ultimate compressive

Discussion

Results show that Young׳s modulus and ultimate strength of bone frozen for five years were lower than for fresh bone. Effects of freezing on mechanical properties of bone have been studied in literature and it was found that freezing does not alter trabecular bone properties (Panjabi et al., 1985, Linde and Sorensen, 1993, Kang et al., 1997, Borchers et al., 1995). However, most of the previous research was conducted on bones that were frozen for less than one year while this study also

Conflict of interest statement

None.

Acknowledgments

We would like to thank the staff at the Imaging Technology Group at the Beckman Institute, Leilei Yin and Mark Bee, for their help in operating the micro-CT equipment, and Travis Ross and Scott Robinson for their assistance in micro-CT imaging. We would also like to thank Professor Ryan Roeder from the University of Notre Dame for providing standards and his guidance on measuring bone mineral density. This research was supported by the National Science Foundation (CMMI 09-27909 ARRA, Dr. Ken

References (41)

H.D. Barth et al.
On the effect of X-ray irradiation on the deformation and fracture behavior of human cortical bone
Bone
(2010)
H.D. Barth et al.
Characterization of the effects of x-ray irradiation on the hierarchical structure and mechanical properties of human cortical bone
Biomaterials
(2011)
G. Bevill et al.
Side-artifact errors in yield strength and elastic modulus for human trabecular bone and their dependence on bone volume fraction and anatomic site
J. Biomech.
(2007)
R.E. Borchers et al.
Effects of selected thermal variables on the mechanical properties of trabecular bone
Biomaterials
(1995)
S.C. Cowin
The relationship between the elasticity tensor and the fabric tensor
Mech. Mater.
(1985)
S.J. Dux et al.
Alterations in damage processes in dense cancellous bone following gamma-radiation sterilization
J. Biomech.
(2010)
R.W. Goulet et al.
The relationship between the structural and orthogonal compressive properties of trabecular bone
J. Biomech.
(1994)
G. Haiat et al.
Relationship between ultrasonic parameters and apparent trabecular bone elastic modulus: a numerical approach
J. Biomech.
(2009)
T.M. Keaveny et al.
Theoretical analysis of the experimental artifact in trabecular bone compressive modulus
J. Biomech.
(1993)
T.M. Keaveny et al.
Trabecular bone modulus and strength can depend on specimen geometry
J. Biomech.
(1993)

T.M. Keaveny et al.

Trabecular bone exhibits fully linear elastic behavior and yields at low strains

J. Biomech.

(1994)

F. Linde et al.

The effect of different storage methods on the mechanical properties of trabecular bone

J. Biomech.

(1993)

A. Singhal et al.

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

J. Mech. Behav. Biomed. Mater.

(2011)

P. Zioupos et al.

Some basic relationships between density values in cancellous and cortical bone

J. Biomech.

(2008)

P.K. Zysset

A review of morphology–elasticity relationships in human trabecular bone: theories and experiments

J. Biomech.

(2003)

C.T. Badea et al.

In vivo small-animal imaging using micro-CT and digital subtraction angiography

Phys. Med. Biol.

(2008)

K.L. Brown et al.

Bone and cartilage transplantation in orthopaedic surgery. A review

J. Bone Joint Surg.

(1982)

Browner, B.D., Green N.E.. 2008. Skeletal Trauma. Saunders, MD Consult—Full Text...

A.J. Burghardt et al.

High-resolution computed tomography for clinical imaging of bone microarchitecture

Clin. Orthop. Relat. Res.

(2011)

X. Cao et al.

Irradiation induces bone injury by damaging bone marrow microenvironment for stem cells

Proc. Nat. Acad. Sci. U.S.A.

(2011)

Cited by (20)

High-Performance Computing Comparison of Implicit and Explicit Nonlinear Finite Element Simulations of Trabecular Bone
2021, Computer Methods and Programs in Biomedicine
Finite element models built from micro-computed tomography scans have become a powerful tool to investigate the mechanical properties of trabecular bone. There are two types of solving algorithms in the finite element method: implicit and explicit. Both of these methods have been utilized to study the trabecular bone. However, an investigation comparing the results obtained using the implicit and explicit solvers is lacking. Thus, in this paper, we contrast implicit and explicit procedures by analyzing trabecular bone samples as a case study.
Micro-computed tomography-based finite element analysis of trabecular bone under a direct quasi-static compression was done using implicit and explicit methods. The differences in the predictions of mechanical properties and computational time of the two methods were studied using high-performance computing.
Our findings indicate that the results using implicit and explicit solvers are well comparable, given that similar problem set up is carefully utilized. Also, the parallel scalability of the two methods was similar, while the explicit solver performed about five times faster than the implicit method. Along with faster performance, the explicit method utilized significantly less memory for the analysis, which shows another benefit of using an explicit solver for this case study.
The comparison of the implicit and explicit methods for the simulation of trabecular bone samples should be highly valuable to the bone modeling community and researchers studying complex cellular and architectured materials.
Microcomputed tomography of craniofacial mineralized tissue: A practical user's guide to study planning and generating quality data
2020, Bone
Whether in a clinical setting or a research environment using model organisms, X-ray-based computed tomography (CT) in its different forms represents the gold standard technology for the non-invasive imaging and quantification of mineralized tissues. While there are many excellent reviews on computed tomography in bone imaging, most focus on the appendicular skeleton. However, the craniofacial skeleton and mineralized dentition, which are frequently imaged for a variety of reasons, can require special considerations to ensure the best quality data are acquired and interpreted correctly. In this review, I will specifically focus on micro-computed tomography (microCT) related to the study of the craniofacial skeleton from the onset of cranioskeletal development through to adulthood using the mouse as the primary reference organism. In so doing, I will cover the important considerations when planning imaging studies, explain critical parameters of both scanning, reconstruction and 3D rendering of data that can impact quantification of different mineralized craniofacial tissues, and options for enabling accurate visualization of tomographic data.
Association between intracortical microarchitecture and the compressive fatigue life of human bone: A pilot study
2020, Bone Reports
Citation Excerpt :
The present results demonstrate similar reductions in the compressive fatigue life of bone even when the radiation dose was reduced by >75% (7.9 kGy). Although some measures of bone quality seem to be unaltered by low radiation doses associated with desktop microCT <1 kGy (Lee and Jasiuk, 2014), a systematic study to determine a tolerable dose that does not affect the fatigue behavior of bone is needed. This study was limited to morphological measures of intracortical microarchitecture and therefore we can only assess the association between these parameters and the fatigue life of bone.
Many mechanical properties of cortical bone are largely governed by the underlying microarchitecture; however, the influence of microarchitecture on the fatigue life of bone is poorly understood. Furthermore, imaging-based studies investigating intracortical microarchitecture may expose bone samples to large doses of radiation that may compromise fatigue resistance. The purpose of this pilot study was to 1) investigate the relationship between intracortical microarchitecture and the fatigue life of human bone in compression and 2) examine the effects of synchrotron irradiation on fatigue life measurements. Cortical samples were prepared from the femoral and tibial shafts of three cadaveric donors. A subset of samples was imaged using synchrotron X-ray microCT to quantify microarchitecture, including porosity, canal diameter, lacunar density, lacunar volume, and lacunar orientation. A second group of control samples was not imaged and used only for mechanical testing. Fatigue life was quantified by cyclically loading both groups in zero-compression until failure. Increased porosity and larger canal diameter were both logarithmically related to a shorter fatigue life, whereas lacunar density demonstrated a positive linear relationship with fatigue life (r² = 45–73%, depending on measure). Irradiation from microCT scanning reduced fatigue life measurements by 91%, but relationships with microarchitecture measurements remained. Additional research is needed to support the findings of this pilot study and fully establish the relationship between intracortical microarchitecture and the compressive fatigue life of bone.
MRI-based assessment of proximal femur strength compared to mechanical testing
2020, Bone
Citation Excerpt :
Another limitation is that our simulations do not model soft tissues, viscoelasticity, moduli anisotropy, and other more complicated mechanics. The cadavers were also frozen for months prior to scanning and testing, which some groups have reported can affect the elastic moduli of bone [67]. Moreover, our powerful desktop computer required around 10 h to compute nonlinear simulations on the higher resolution images.
Half of the women who sustain a hip fracture would not qualify for osteoporosis treatment based on current DXA-estimated bone mineral density criteria. Therefore, a better approach is needed to determine if an individual is at risk of hip fracture from a fall. The objective of this study was to determine the association between radiation-free MRI-derived bone strength and strain simulations compared to results from direct mechanical testing of cadaveric femora.
Imaging was conducted on a 3-Tesla MRI scanner using two sequences: one balanced steady-state free precession sequence with 300 μm isotropic voxel size and one spoiled gradient echo with anisotropic voxel size of 234 × 234 × 1500 μm. Femora were dissected free of soft-tissue and 4350-ohm strain-gauges were securely applied to surfaces at the femoral shaft, inferior neck, greater trochanter, and superior neck. Cadavers were mechanically tested with a hydraulic universal test frame to simulate loading in a sideways fall orientation.
Sideways fall forces were simulated on MRI-based finite element meshes and bone stiffness, failure force, and force for plastic deformation were computed. Simulated bone strength metrics from the 300 μm isotropic sequence showed strong agreement with experimentally obtained values of bone strength, with stiffness (r = 0.88, p = 0.0002), plastic deformation point (r = 0.89, p < 0.0001), and failure force (r = 0.92, p < 0.0001). The anisotropic sequence showed similar trends for stiffness, plastic deformation point, and failure force (r = 0.68, 0.70, 0.84; p = 0.02, 0.01, 0.0006, respectively). Surface strain-gauge measurements showed moderate to strong agreement with simulated magnitude strain values at the greater trochanter, superior neck, and inferior neck (r = −0.97, −0.86, 0.80; p ≤0.0001, 0.003, 0.03, respectively). The findings from this study support the use of MRI-based FE analysis of the hip to reliably predict the mechanical competence of the human femur in clinical settings.
Advanced interlocking systems to improve heavy-load-bearing characteristics of flexible intramedullary nailing
2016, Materials Science and Engineering C
Citation Excerpt :
A possible influence of freeze/thaw cycles on the mechanical parameters cannot be ruled out completely. However, we consider the influence of freezing as negligible due to the short time (less than 4 months) the specimens were frozen and the low number of cycles (freezing after operation and one time thawing for the potting procedure and test) [31, 32]. Load was applied via blocks of bone cement, which influences the measured absolute values of the specimen's stiffness.
Flexible intramedullary nailing (FIN) is a minimally invasive and widespread standard method for osteosynthesis of pediatric long bone fractures. In the case of unstable fractures of the lower extremity, interlocking systems need to be used to prevent axial shortening and subsequent perforation of the nail at its insertion site. In the present study, four different screw-fixed interlocking systems for FINs (Hofer TwinPlug with two 3-mm titanium interlocking screws, Hofer FixPlug with 3-mm titanium interlocking screw, Hofer Plug with 3.5-mm titanium interlocking screw, and Hofer Plug with 3-mm titanium interlocking screw) in comparison with the commonly used Ender stainless steel nails (locked with 3.5-mm screw) were experimentally investigated in cadaveric lamb tibiae, regarding their load characteristics and failure modes in the case of heavy loading. The specimens were subjected to sequential axial cyclic loading of 5000cycles with stepwise increase of the load amplitude until failure. Migration of locking screws and internal damage of bone tissue was quantified by micro-computed tomography (CT) imaging. Ender nails failed on average at a peak load of 800 N, TwinPlugs at 1367 N, FixPlugs at 1222 N, Plugs 3.5mm at 1225 N and Plugs 3.0mm at 971 N. TwinPlugs, FixPlugs, and Plugs 3.5mm failed in a slow manner over several hundred loading cycles, whereas Ender nails and Plugs 3.0mm exhibited abrupt failure without any prior indication. Our results confirm that axial stability of FIN can be further improved by screw-fixed plugs by simultaneously avoiding shortcomings of an eye-locked system, which the Ender nails are. Considering biomechanical results, plug interlocking systems with 3.5-mm screws should be favored over conventional Ender nails and plugs with 3-mm screws.
The effects of experimental freeze-thaw cycles to bone as a component of subaerial weathering
2016, Journal of Archaeological Science: Reports
Citation Excerpt :
Freezing has been theorized to affect negatively the mechanical properties of bone (Borchers et al., 1995; Boutros et al., 2000; Diefenbeck et al., 2011; Gleizes et al., 1998; Johnson, 1985; Jung et al., 2011; Karr and Outram, 2012; Kaye et al., 2012; Lee and Jasiuk, 2014; Nyman et al., 2006; Torimitsu et al., 2014; Van Haaren et al., 2008; Wilson et al., 2006). Studies have subjected human and non-human bone to temperatures between − 18 °C and − 70 °C for varying time intervals before testing specific biomechanical properties, including rigidity and elasticity (Gleizes et al., 1998); integrity of trabecular bone (Borchers et al., 1995; Lee and Jasiuk, 2014); bending, torsion, energy absorption, and hardness (Van Haaren et al., 2008); and fracturability or fracture load (Kaye et al., 2012; Torimitsu et al., 2014). For example, Gleizes et al. (1998) noted that freezing at − 18 °C for three months did not affect the rigidity or elastic behavior of sheep vertebral discs.
Subaerial weathering of bone is a taphonomic process that affects many archaeological and paleontological assemblages and is characterized by surface bleaching, loss of organic component, and progressive cracking and fragmentation of the bone surfaces. The mechanisms by which these changes occur, however, are poorly understood but are hypothesized to include ultraviolet exposure, degreasing, leaching, remineralization, wetting/drying, thermal expansion/contraction, and freezing/thawing. In order to examine the potential contribution of the latter process to weathering, a sample (n = 93) of naturally decomposed and degreased metapodials of white-tailed deer (Odocoileus virginianus) were subjected to up to 75 cycles of complete freezing and thawing under laboratory conditions, with samples withdrawn each 25 cycles for histological examination of the taphonomic alterations to the bone microstructure. Macroscopic and microscopic changes included progressive cracking that parallels part of the progressive degradation of weathered bone, but no elements unequivocally reached weathering stage 1 over this interval.

View all citing articles on Scopus

View full text

Short communicationEffects of freeze–thaw and micro-computed tomography irradiation on structure–property relations of porcine trabecular bone

Abstract

Introduction

Section snippets

Sample preparation

Results

Discussion

Conflict of interest statement

Acknowledgments

Bone

Biomaterials

J. Biomech.

Biomaterials

Mech. Mater.

J. Biomech.

J. Biomech.

J. Biomech.

J. Biomech.

J. Biomech.

J. Biomech.

J. Biomech.

J. Mech. Behav. Biomed. Mater.

J. Biomech.

J. Biomech.

In vivo small-animal imaging using micro-CT and digital subtraction angiography

Phys. Med. Biol.

Bone and cartilage transplantation in orthopaedic surgery. A review

J. Bone Joint Surg.

High-resolution computed tomography for clinical imaging of bone microarchitecture

Clin. Orthop. Relat. Res.

Irradiation induces bone injury by damaging bone marrow microenvironment for stem cells

Proc. Nat. Acad. Sci. U.S.A.

Short communication
Effects of freeze–thaw and micro-computed tomography irradiation on structure–property relations of porcine trabecular bone