Influence of nonenzymatic glycation on biomechanical properties of cortical bone

In this study, the influence of nonenzymatic glycation (NEG) on the mechanical properties of bone and bone collagen were investigated. Bovine cortical bone specimens were incubated in ribose to cause collagen cross-links in vitro, and nondestructive mechanical testing was used to determine tensile and compressive elastic modulus as a function of incubation time. Mechanical properties associated with yield, postyield, and final fracture of bone were determined at the end of the incubation period. The stiffness of the collagen network was measured using stress relaxation tests of demineralized bone cylinders extracted periodically throughout the incubation period. It was found that accumulation of nonenzymatic glycation end-products in cortical bone caused stiffening of the type I collagen network in bone (r2 = 0.92; p < 0.001) but did not significantly affect the overall stiffness of the mineralized bone (p = 0.98). The ribosylated group had significantly more NEG products and higher yield stress and strain than the control group (p < 0.05). Postyield properties including postyield strain and strain energy were lower in the ribosylated group but were not significantly different from the control group (p = 0.24). Compared with the control group, the ribosylated group was characterized by significantly higher secant modulus and lower damage fraction (p < 0.05). Taken together, the results of this study suggest that collagen in bone is susceptible to the same NEG-mediated changes as collagen in other connective tissues and that an increased stiffness of the collagen network in bone due to NEG may explain some of the age-related increase in skeletal fragility and fracture risk.     

A study of some properties of a sample of bovine cortical bone using ultrasound

Summary Both collagen fibers and long bone exhibit similar anisotropy in several respects, strongly supporting the concept that the anisotropy of bone has its source in the contained collagen fibers which are parallel to the bone axis in mature adult mammalian bone. Experimental information was obtained for a sample of adult bovine tibia from the midregion of the bone prepared as 22 specimens, 2 mm thick, in three orientations—axial, tangential, and radial—with respect to the measurement of sonic velocity. Thickness, sonic transit time, and weight were obtained for the original wet specimens in 0.15M saline, after dehydration at room temperature, after rehydration, and after demineralization in 0.5M EDTA solution. The average density and volume fractions of water, mineral, and organic components closely matched those reported by Biltz and Pellegrino for bovine bone. The distribution of orientation-dependent properties (velocity, change in velocity, and change in thickness) kept the same pattern in every state, although the intensity of the anisotropy varied. The fractional dimensional changes when samples were dehydrated were greatest in the radial direction and least in the axial, just as for collagen fibers. An almost identical decrease in thickness was observed in magnitude and distribution when the specimens were demineralized. Estimates of the sonic velocity for bone collagen were obtained with the aid of the Lees-Davidson hypothesis. These values often matched the published values for collagen fibers, summarized as follows: -1107-1     

Comparative biomechanical studies of conventional and self-tapping cortical bone screws]

In normal cortical screws the thread must be cut by a separate thread-cutter. So a high stability of the connection is achieved. On the other hand thread cutting needs time, which may be assumed to several minutes during a plate osteosynthesis. To avoid thread-cutting, self-tapping screws were used 20 years ago. These old self-tapping screws had a reduced stability and a lower torque at the final screw driving so that they tended to destroy the thread. By a modification of the cutting edges at the tip of the screw the tapping quality is increased decisively. The microscopic examination showed a better cutting of the new screws with little difference to a thread-cutter. In biomechanical tests the final torque and the maximum tension force of the self-tapping screws were at the same level as in usual pre-cut screws. In the practical use the new self-tapping screws have a good handling and save time during operation. In osteoporosis, they should be used carefully in order to meet the opposite corticalis' hole and not to destroy the ipsilateral corticalis' thread.     

Intrinsic material properties of cortical bone

The G171V mutation (high bone mass, HBM) is autosomal dominant and is responsible for high bone mass in humans. Transgenic HBM mice in which the human LRP5 G171V gene is inserted also show a similar phenotype with greater bone mass and biomechanical performance than wild-type mice, as determined by whole bone testing. Whole bone mechanics, however, depend jointly on bone mass, architecture, and intrinsic bone tissue mechanical properties. To determine whether the HBM mutation affects tissue-level biomechanical performance, we performed nano-indentation testing of unembedded cortical bone from HBM mice and their nontransgenic (NTG) littermates. Femora from 17-week-old mice (female, 8 mice/genotype) were subjected to nano-indentation using a Triboscope (Hysitron, Minneapolis, MN, USA). For each femoral specimen, approximately 10 indentations were made on the midshaft anterior surface with a target force of either 3 or 9 mN at a constant loading rate of 400 mN/s. The load–displacement data from each test were used to calculate indentation modulus and hardness for bone tissue. The intrinsic material property that reflected the bone modulus was greater (48%) in the HBM as compared to the NTG mice. Our results of intrinsic properties are consistent with the published structural and material properties of the midshaft femur in HBM and NTG mice. The greater intrinsic modulus in HBM reflects greater bone mineral content as compared to NTG (wild-type, WT) mice. This study suggests that the greater intrinsic property of cortical bone is derived from the greater bone mineral content and BMD, resulting in greater bone strength in HBM as compared to NTG (WT) mice.     

Some properties of the organic matrix of a bovine cortical bone sample in various media

Summary Dimensional stability of a demineralized bovine cortical bone sample was found in all media whether EDTA, saline or ethanol and water solutions or even 100% ethanol. A 6% volume shrinkage was observed, in strong contrast to the reported swelling for tail tendon fiber collagen. Sonic velocity was strongly dependent on the state and the medium, varying by a factor greater than 2. The medium appears to contribute strongly to the observed velocity suggesting that the Reuss formalism is applicable with the solid collagen skeleton as one component and the liquid in the pores as the second. Sonic anisotropy was noted although the intensity varied. The radial to axial velocity was greatest (0.93) in saline and least in 100% ethanol (0.80) indicating that the rigidity of the tissue influenced the character of sonic propagation. Two sets of intermolecular linkages are inferred. One set, in common with tendon collagen, controls the elastic properties. A second set in bone collagen maintains dimensional stability.     

Effect of age on mechanical properties of the collagen phase in different orientations of human cortical bone

The collagen phase plays an important role in mechanical behaviors of cortical bone. However, aging effects on the mechanical behavior of the collagen phase is still poorly understood. In this study, micro-tensile tests were performed on demineralized human cortical bone samples from young, middle-aged, and elderly donors and aging effects on the mechanical properties of the collagen phase in different orientations (i.e. longitudinal and transverse directions of bone) were examined. The results of this study indicated that the elastic modulus and ultimate strength of the demineralized bone specimens decreased with aging in both the longitudinal and transverse orientations. However, the failure strain exhibited no significant changes in both orientations regardless of aging. These results suggest that the stiffness and strength of the collagen phase in bone are deteriorated with aging in both longitudinal and transverse directions. However, the aging effect is not reflected in the failure strain of the collagen phase in both longitudinal and transverse orientations, implying that the maximum sustainable deformation of the collagen phase is independent of aging and orientation.     

Effects of three different preservation methods on the mechanical properties of human and bovine cortical bone

Background and purposeIn the development of new strategies for fracture fixation, new methods have to be tested biomechanically under in vitro conditions before clinical trials can be performed. The gold standard for laboratory evaluations is fresh-frozen specimen. As the availability of fresh-frozen specimens is limited and since their use bears infectious risks, specimens treated with various chemical embalming fluids are also used. These preservation methods may alter the mechanical properties of the specimens used. Therefore, the aims of the present study were to determine the effects of three different preservation methods (formalin fixation (FO), Thiel-fixation (TH), and alcohol–glycerine fixation (AG)) on the elastic and postyield mechanical properties of cortical bone and to compare these properties to those of fresh-frozen (FF) specimens.Materials and methodsCylindrical cortical specimens (diameter 3 mm, length 60 mm) were obtained from human femurs (n = 48) and bovine tibiae (n = 40). Before specimen immersion in different fixation fluids, bone mineral density (BMD) as well as the initial Young's modulus was determined. The Young's modulus was determined in a nondestructive bending test, and measurements were repeated after 6 months of immersion in fixative solution. Subsequent to the nondestructive test, a destructive 3-point bending test was conducted to assess the postyield and fracture properties.ResultsThe BMD as well as the initial Young's modulus showed no significant differences between the four test groups. After 6 months in fixative solution, the Young's modulus was significantly lowered in human Thiel specimens and only showed minor changes in formalin- and alcohol–glycerine-treated specimens. The plastic energy absorption of human and bovine specimens was altered significantly. Formalin as well as alcohol–glycerine fixation yielded a significant decrease in plastic energy absorption, whereas Thiel fixation significantly increased the plastic energy absorption.Discussion/conclusionBecause of the significantly altered plastic mechanical properties of cortical bone, the use fresh-frozen bone specimens is recommended in biomechanical studies investigating failure loads of orthopaedic implants. The use of embalmed specimens should be restricted to pilot tests.     

Utilizing biologic assimilation of bovine fetal collagen in staged skin grafting

Seven patients underwent 2-stage skin grafting with bovine fetal collagen (BFC) as an initial wound cover. Split-thickness skin grafts were successfully placed on the wounds after completion of interval management. BFC proved to be a resilient acellular dermal matrix that could proceed to assimilation and skin grafting under a variety of wound conditions. BFC may prove to be a valuable material, as the role of acellular dermal matrices in skin grafting becomes better defined.     

Quantitative ultrashort echo time (UTE) MRI of human cortical bone: correlation with porosity and biomechanical properties

In this study we describe the use of ultrashort echo time (UTE) magnetic resonance imaging (MRI) to evaluate short and long T2* components as well as the water content of cortical bone. Fourteen human cadaveric distal femur and proximal tibia were sectioned to produce 44 rectangular slabs of cortical bone for quantitative UTE MR imaging, microcomputed tomography (µCT), and biomechanical testing. A two‐dimensional (2D) UTE pulse sequence with a minimal nominal TE of 8 µseconds was used together with bicomponent analysis to quantify the bound and free water in cortical bone using a clinical 3T scanner. Total water concentration was measured using a 3D UTE sequence together with a reference water phantom. UTE MR measures of water content (total, free, and bound), T2* (short and long), and short and long T2* fractions were compared with porosity assessed with µCT, as well as elastic (modulus, yield stress, and strain) and failure (ultimate stress, failure strain, and energy) properties, using Pearson correlation. Porosity significantly correlated positively with total (R² = 0.23; p < 0.01) and free (R² = 0.31; p < 0.001) water content as well as long T2* fraction (R² = 0.25; p < 0.001), and negatively with short T2* fraction and short T2* (R² = 0.24; p < 0.01). Failure strain significantly correlated positively with short T2* (R² = 0.29; p < 0.001), ultimate stress significantly correlated negatively with total (R² = 0.25; p < 0.001) and bound (R² = 0.22; p < 0.01) water content, and failure energy significantly correlated positively with both short (R² = 0 30; p < 0.001) and long (R² = 0.17; p < 0.01) T2* values. These results suggest that UTE MR measures are sensitive to the structure and failure properties of human cortical bone, and may provide a novel way of evaluating cortical bone quality. © 2012 American Society for Bone and Mineral Research.     

Pyridoxine deficiency affects biomechanical properties of chick tibial bone

The mechanical integrity of bone is dependent on the bone matrix, which is believed to account for the plastic deformation of the tissue, and the mineral, which is believed to account for the elastic deformation. The validity of this model is shown in this study based on analysis of the bones of vitamin B₆-deficient and vitamin B₆-replete chick bones. In this model, when B₆-deficient and control animals are compared, vitamin B₆ deficiency has no effect on the mineral content or composition of cortical bone as measured by ash weight (63 ± 6 vs. 58 ± 3); mineral to matrix ratio of the FTIR spectra (4.2 ± 0.6 vs. 4.5 ± 0.2), line-broadening analyses of the X-ray diffraction 002 peak (β₀₀₂ = 0.50 ± 0.1 vs. 0.49 ± 0.01), or other features of the infrared spectra. In contrast, collagen was significantly more extractable from vitamin B₆ deficient chick bones (20 ± 2 % of total hydroxyproline extracted vs. 10 ± 3% p ≤ 0.001). The B₆-deficient bones also contained an increased amount of the reducible cross-links DHLNL, dehydro-dihydroxylysinonorleucine, (1.03 ± 0.07 vs. 0.84 ± 0.13 p < 0.001); and a nonsignificant increase in HLNL, dehydrohydroxylysinonorleucine, (0.51 ± 0.03 vs. 0.43 ± 0.03, p ≤ 0.10). There were no significant changes in bone length, bone diameter, or area moment of inertia. In four-point bending, no significant changes in elastic modulus, stiffness, offset yield deflection, or fracture deflection were detected. However, fracture load in the B₆-deficient animals was decreased from 203 ± 35 MPa to 151 ± 23 MPa, p ≤ 0.01, and offset yield load was decreased from 165 ± 9 MPa to 125 ± 14 MPa, p ≤ 0.05. Since earlier histomorphometric studies had demonstrated that the B₆-deficient bones were osteopenic, these data suggest that although proper cortical bone mineralization occurred, the alterations of the collagen resulted in changes to bone mechanical performance.     

Dynamic loads are determinants of peak bone mass.

This study investigated the association between non-invasive measurements of bone mass and markers of dynamic and static hip joint loads in subjects expected to be at peak bone mass. The bone mineral density (BMD) and bone mineral content (BMC) of three proximal femoral sites (neck, greater trochanter, and total) were measured by dual energy X-ray absorptiometry, and the peak external joint moments at the hip during walking and jogging were calculated from gait analyses of 31 normal human subjects ranging in age from 30 to 49 years (18 females, 13 males). Various multiple regression analyses were performed to determine how much of the variance in BMD and BMC was explained by height, body mass, and the peak hip joint moments. In total, the models explained up to 40% of the variance in BMD and 58% of the variance in BMC. Inclusion of height or body mass did not increase the explanatory power of the models for BMD and explained no more than 8% of the total variance in BMC once the joint moments from walking were allowed to enter the models. These data support the hypothesis that variance in peak bone mass is associated with variance in dynamic hip loads largely independent of the effect of static factors such as height and body mass.     

Relationship among bone mineral density, collagen composition, and biomechanical properties of callus in the healing of osteoporotic fracture

Objective： To study the change and relationship among bone mineral density （BMD）, collagen composition and biomechanical properties of the callus in the healing process of osteoporotic fracture. Methods： The osteoporotic rat model and fracture model were established through bilateral ovariectomy （OV＇X） and osteotomy of the middle shaft of the right hind tibiae, respectively. Ninety female SD rats were randomly divided into OVX group and sham group. With the samples of blood and callus, roentgenoraphic and histological observation were performed for the assessment of the healing progress of the fracture, and the serum concentration of TRAP-5b, proportion of type I collagen, BMD and biomechanical properties of the callus were measured. Results： The OVX group experienced a significant delay of fracture healing. The mean serum concentration of TRAP-5b of rats in the OVX group was much higher than that in the sham group after the operation （P 〈 0.05）, but the difference at the same time point after fracture was smaller than that before fracture （P 〈 0.05）. The BMD of the callus in both groups reached the peak value at the 6 th week after fracture while the proportion of the type I collagen and the biomechanical strength reached the peak at the 8th week. Conclusions. The deficiency of estrogen after the ovariectomy could induce the up-regulation of the osteoclasts activities, whereas the potency of further activation after fracture was depressed. Although the synthesis of collagen together with its mineralization determines the biomechanical properties of new bone, the accumulation of collagen could be assessed as an index in the prediction of biomechanical strength of bones independent of the bone mineral deposition.     

Age-related changes in mineral of rat and bovine cortical bone

Summary The mineral of cortical bones has been studied in newborn, growing, and adult rats and in the calf and cow, using X-ray diffraction and infrared spectroscopy during the thermal decomposition of bones and by microassay of carbonate. The mineral of all the bone samples, regardless of species or age, was found to be a calcium-deficient apatite containing both CO₃ ²⁻ and HPO₄ ²⁻ ions in the crystal lattice. The crystal size, Ca/P molar ratio, and CO₃ ²⁻ ion content of cortical bone all increased with increasing age in both the rat and the bovine. The Ca/P ratio varied from 1.51 in newborn rats to 1.69 in adults but remained that of Ca-deficient apatite even though its value was close to that of stoichiometric hydroxyapatite (1.67). Both the carbonate ion and the hydrogenophosphate ion contents varied from one animal species to another and with age within a given species. Maturation was correlated with an increase in carbonate ion content, which replaced the HPO₄ ²⁻ ions. In contrast, the calcium ion number per unit formula did not vary during maturation. Cortical bone mineral, in both species, regardless of age, can therefore be represented by the following formula: Ca_8.3(PO₄)_4.3(CO₃)_x(HPO₄)_y(OH)_0.3; y decreased and x increased with increasing age, (x+y) being constant, equal to 1.7.     

Improvement of success rates for abdominal component reconstructions using bovine fetal collagen

Between February 2008 and December 2010, the author performed 15 standardized component repairs in patients reinforced with bovine fetal collagen. Of 15 repairs, 14 (93%) were intact at an average follow-up of 18 months. This success rate is a significant improvement from the author's previous success rate with this procedure (67%) and may represent a specific contribution of bovine fetal collagen reinforcement.     

High bone turnover of type I collagen depends on fetal growth

The bone metabolic processes of proliferation and differentiation in preterm and term newborns have yet to be fully elucidated. Seventy-four umbilical cord blood samples were collected from preterm and term newborns delivered at 27 to 42 gestational weeks (GWs). Carboxy-terminal propeptide of type I procollagen (PICP), pyridinoline cross-linked telopeptide domain of type I collagen (ICTP), alkaline phosphatase (ALP), and bone-specific alkaline phosphatase (BAP) were measured. Calcitonin (CT), estrogen (E2), intact parathyroid hormone, and insulin-like growth factor-I (IGF-I) were also examined in 20 or 23 randomly selected samples. We conducted cross-sectional regression analyses for bone metabolic markers, fetal growth markers including GWs, birth weight (BW), height (BH) and head circumference (HC), and bone related hormones. PICP and ICTP activities were very high, but decreased significantly with fetal growth based on GWs, BW, BH, and HC changes (GWs, BW, and BH to both PICP and ICTP, P < 0.0001; HC to ICTP, P < 0.0001; HC to PICP, P < 0.05), while BAP and ALP did not change significantly. E2 and CT both showed a significant positive correlation with Ca (P < 0.05), but neither hormone had any apparent correlation with PICP, ALP, BAP, or ICTP. These results suggest very active bone formation and resorption of type I collagen to be dependent on fetal growth and that fetal osteoblasts dominate the proliferation phase of development rather than the maturation phase. However, factors contributing to high bone turnover in the fetus remain to be elucidated.     

Low-megahertz ultrasonic properties of bovine cancellous bone

Ultrasound offers a noninvasive means to detect changes that occur to the density of cancellous bone as a result of degenerative diseases such as osteoporosis. Techniques based on the velocity and frequency dependence of attenuation of ultrasonic pulses propagated through cancellous bone have proven sensitive to bone density. Most previous studies have investigated these two parameters in the frequency range of 0.1–1.0 MHz. The present study had two goals. The first was to measure three ultrasonic parameters: longitudinal mode velocity; broadband ultrasonic attenuation (BUA); and apparent integrated backscatter (AIB), at higher frequencies using a broadband 2.25 MHz measurement system. The second goal was to assess the dependence of these parameters on bone density. Twenty-one specimens of cancellous bone acquired from the proximal end of four bovine tibiae were investigated in this study. The apparent density of the specimens (determined with the bone marrow removed and the specimens thoroughly dry) ranged between 0.3 and 0.9 g/cm³. Ultrasonic measurements were performed along three mutually perpendicular directions corresponding to the anteroposterior (AP), mediolateral (ML), and superoinferior (SI) axes of the tibia. A linear regression was used to analyze the results of these measurements as a function of apparent density. Velocity demonstrated a highly significant linear increase with density for all three directions (AP: p < 0.001; ML: p < 0.001; SI: p < 0.01). AIB decreased with density in all three directions; however, only the ML and SI directions demonstrated a significant linear correlation (AP: p = n.s.; ML: p < 0.05; SI: p < 0.05). In the frequency range 0.5–1.0 MHz, BUA exhibited a significant linear increase in the AP and ML directions, but not the SI direction (AP: p < 0.05; ML: p < 0.01; SI: p = n.s.). In contrast, in the frequency range 1.0–2.0 MHz, BUA exhibited a highly significant increase with density in the SI direction, but no significant change in the AP and ML directions (AP: p = n.s., ML: p = n.s., SI: p < 0.001).     

Age-related changes in physicochemical properties of mineral crystals are related to impaired mechanical function of cortical bone

The measures of bone mass and architecture need to be supplemented with physicochemical and compositional measures for better assessment of fracture risk. In the current studies, we investigated the effects of physicochemical properties of mineral crystals on tissue and organ-level mechanical function of aging rat cortical bone. Our hypothesis was that age-related changes in physicochemical properties of mineral crystals are related to impaired elastic deformability of cortical bone tissue. Raman microspectroscopy was used to investigate the age-related changes in mineralization (relative amounts of mineral and organic matrix), the substitution of carbonate ions in phosphate positions (type-B carbonate substitution) and mineral crystallinity (the orderliness of crystal lattice) of femurs from young adult (3-month old), middle-aged (8-month old) and aged (24-month old) female Sprague-Dawley rats. Cross-sectional properties, the area and the moment of inertia at the mid-diaphysis, were histomorphometrically quantified and the elastic deformation capacity of femurs was quantified via three-point bending tests. It was observed that the elastic deformation capacity of aged rats was significantly impaired both at the tissue and the organ levels with increasing age. In parallel with this impairment in the elastic deformability and in support of our hypothesis, we found that increasing mineralization, increasing crystallinity and increasing type-B carbonate substitution were significantly correlated with decreasing elastic deformation capacity with age. We conclude that the measure of bone mass needs to be supplemented with measures reflecting the physicochemical status of mineral crystals for improved assessment of fracture susceptibility.     

Torsional stability of interference screws derived from bovine bone - a biomechanical study

Background  

In the present biomechanical study, the torsional stability of different interference screws, made of bovine bone, was tested. Interference screws derived from bovine bone are a possible biological alternative to conventional metallic or bioabsorbable polymer interference screws.