Accounting for spatial variation of trabecular anisotropy with subject-specific finite element modeling moderately improves predictions of local subchondral bone stiffness at the proximal tibia

IntroductionPreviously, a finite element (FE) model of the proximal tibia was developed and validated against experimentally measured local subchondral stiffness. This model indicated modest predictions of stiffness (R² = 0.77, normalized root mean squared error (RMSE%) = 16.6%). Trabecular bone though was modeled with isotropic material properties despite its orthotropic anisotropy. The objective of this study was to identify the anisotropic FE modeling approach which best predicted (with largest explained variance and least amount of error) local subchondral bone stiffness at the proximal tibia.MethodsLocal stiffness was measured at the subchondral surface of 13 medial/lateral tibial compartments using in situ macro indentation testing. An FE model of each specimen was generated assuming uniform anisotropy with 14 different combinations of cortical- and tibial-specific density-modulus relationships taken from the literature. Two FE models of each specimen were also generated which accounted for the spatial variation of trabecular bone anisotropy directly from clinical CT images using grey-level structure tensor and Cowin’s fabric-elasticity equations. Stiffness was calculated using FE and compared to measured stiffness in terms of R² and RMSE%.ResultsThe uniform anisotropic FE model explained 53–74% of the measured stiffness variance, with RMSE% ranging from 12.4 to 245.3%. The models which accounted for spatial variation of trabecular bone anisotropy predicted 76–79% of the variance in stiffness with RMSE% being 11.2–11.5%.ConclusionsOf the 16 evaluated finite element models in this study, the combination of Synder and Schneider (for cortical bone) and Cowin’s fabric-elasticity equations (for trabecular bone) best predicted local subchondral bone stiffness.     

Evaluation of LR white resin for histology of the undecalcified rat tibia

Histology of plastic embedded undecalcified bone represents a challenging problem to the histotechnologist. We outline here an exploration of LR White resin as a suitable medium for histologic study of undecalcified rat tibia. A procedure was developed for light microscopy of rat tibia embedded in LR White and sectioned by sawing-grinding technics. The specimens were fixed in 10% neutral buffered formalin or alcohol-acetic acid-formol, dehydrated in ethanol, defatted in chloroform followed by resin infiltration and heat-curing of embedded blocks. The procedure of dehydration, defatting, infiltration, and polymerization can be completed within 10 days. Cold curing with accelerator provided by the manufacturer did not yield superior results compared to blocks cured with heat. Thick sections were obtained using a diamond wire saw, attached to plexiform slides, then ground and polished. Surface staining with Von Kossa silver reagent or toluidine blue revealed satisfactory morphological preservation of the mineralized bone sections. Artifacts like small bubbles appeared occasionally and could not be avoided despite prolonged infiltration or cold curing of blocks. Our method is relatively simple for base-line histologic study of rat tibia. The method offers advantages such as easy adaptability, reliable stainability, contrast, and resolution of bone architecture and marrow cells. Two other embedding media, Micro-Bed resin and Unicryl, were also tested, but produced inferior results.     

Amino and carboxyl propeptides in bone collagen fibrils during embryogenesis

Summary Collagen fibrillogenesis was studied in tibiae of chick embryos, 9, 11, and 14 days old. Specimens were incubated with antibodies against the amino and the carboxyl propeptides of type-I collagen and subjected to ferritin-la-belling immuno-electron microscopy. The amino propeptide was found in thin fibrils, 20–40 nm in diameter, distributed at 60-nm periodicity. The carboxyl propeptide antibody labelled a wide spectrum of fibrils, although the majority were in the range of 40–100 nm, distinctly larger than those labelled with the amino propeptide antibody. The presence of pN (amino propeptide plus collagen) and pC (carboxyl propeptide plus collagen) collagen was also demonstrated by Western blotting in all specimens. This study suggests that the sequence of propeptide removal may regulate collagen fibril diameter.     

Longitudinal Evaluation of Mouse Hind Limb Bone Loss After Spinal Cord Injury using Novel,in vivo,Methodology

Spinal cord injury (SCI) is often accompanied by osteoporosis in the sublesional regions of the pelvis and lower extremities, leading to a higher frequency of fractures ¹. As these fractures often occur in regions that have lost normal sensory function, the patient is at a greater risk of fracture-dependent pathologies, including death. SCI-dependent loss in both bone mineral density (BMD, grams/cm²) and bone mineral content (BMC, grams) has been attributed to mechanical disuse ², aberrant neuronal signaling ³ and hormonal changes ⁴. The use of rodent models of SCI-induced osteoporosis can provide invaluable information regarding the mechanisms underlying the development of osteoporosis following SCI as well as a test environment for the generation of new therapies ^5-7 (and reviewed in ⁸). Mouse models of SCI are of great interest as they permit a reductionist approach to mechanism-based assessment through the use of null and transgenic mice. While such models have provided important data, there is still a need for minimally-invasive, reliable, reproducible, and quantifiable methods in determining the extent of bone loss following SCI, particularly over time and within the same cohort of experimental animals, to improve diagnosis, treatment methods, and/or prevention of SCI-induced osteoporosis.An ideal method for measuring bone density in rodents would allow multiple, sequential (over time) exposures to low-levels of X-ray radiation. This study describes the use of a new whole-animal scanner, the IVIS Lumina XR (Caliper Instruments) that can be used to provide low-energy (1-3 milligray (mGy)) high-resolution, high-magnification X-ray images of mouse hind limb bones over time following SCI. Significant bone density loss was seen in the tibiae of mice by 10 days post-spinal transection when compared to uninjured, age-matched control (naïve) mice (13% decrease, p<0.0005). Loss of bone density in the distal femur was also detectable by day 10 post-SCI, while a loss of density in the proximal femur was not detectable until 40 days post injury (7% decrease, p<0.05). SCI-dependent loss of mouse femur density was confirmed post-mortem through the use of Dual-energy X-ray Absorptiometry (DXA), the current gold standard for bone density measurements. We detect a 12% loss of BMC in the femurs of mice at 40 days post-SCI using the IVIS Lumina XR. This compares favorably with a previously reported BMC loss of 13.5% by Picard and colleagues who used DXA analysis on mouse femurs post-mortem 30 days post-SCI ⁹. Our results suggest that the IVIS Lumina XR provides a novel, high-resolution/high-magnification method for performing long-term, longitudinal measurements of hind limb bone density in the mouse following SCI.     

Targeting the upregulation of reactive oxygen species subsequent to hyperglycemia prevents type 1 diabetic cardiomyopathy in mice

Cardiac oxidative stress is an early event associated with diabetic cardiomyopathy, triggered by hyperglycemia. We tested the hypothesis that targeting left-ventricular (LV) reactive oxygen species (ROS) upregulation subsequent to hyperglycemia attenuates type 1 diabetes-induced LV remodeling and dysfunction, accompanied by attenuated proinflammatory markers and cardiomyocyte apoptosis. Male 6-week-old mice received either streptozotocin (55 mg/kg/day for 5 days), to induce type 1 diabetes, or citrate buffer vehicle. After 4 weeks of hyperglycemia, the mice were allocated to coenzyme Q₁₀ supplementation (10 mg/kg/day), treatment with the angiotensin-converting-enzyme inhibitor (ACE-I) ramipril (3 mg/kg/day), treatment with olive oil vehicle, or no treatment for 8 weeks. Type 1 diabetes upregulated LV NADPH oxidase (Nox2, p22^phox, p47^phox and superoxide production), LV uncoupling protein UCP3 expression, and both LV and systemic oxidative stress (LV 3-nitrotyrosine and plasma lipid peroxidation). All of these were significantly attenuated by coenzyme Q₁₀. Coenzyme Q₁₀ substantially limited type 1 diabetes-induced impairments in LV diastolic function (E:A ratio and deceleration time by echocardiography, LV end-diastolic pressure, and LV −dP/dt by micromanometry), LV remodeling (cardiomyocyte hypertrophy, cardiac fibrosis, apoptosis), and LV expression of proinflammatory mediators (tumor necrosis factor-α, with a similar trend for interleukin IL-1β). Coenzyme Q_10's actions were independent of glycemic control, body mass, and blood pressure. Coenzyme Q₁₀ compared favorably to improvements observed with ramipril. In summary, these data suggest that coenzyme Q₁₀ effectively targets LV ROS upregulation to limit type 1 diabetic cardiomyopathy. Coenzyme Q₁₀ supplementation may thus represent an effective alternative to ACE-Is for the treatment of cardiac complications in type 1 diabetic patients.     

Modeling and remodeling responses to normal loading in the human lower limb

Limb bones are designed to be strong enough to support the body and yet be energetically conservative during locomotion. Bones of the distal segment, which are relatively costly to move, are often more slender than bones of the proximal segments, even though they must sustain proportionally greater loads. As a result, they are expected to experience a higher incidence of microdamage. With this constraint in mind, Lieberman and Crompton (1998 Principles of Animal Design, Cambridge: Cambridge University Press, p. 78-86) proposed that bones response to strain varies along the proximo-distal axis of the limb. In order to avoid fatigue fractures due to the accumulation of microdamage, the distal segment, in comparison to the proximal segment, will have an increase in remodeling events to replace damaged bone. In this paper, we test the hypothesis of Lieberman and Crompton (1998) with respect to the human lower limb. With a sample of adult individuals, we compare tibiae and femora for mid-diaphyseal cross-sectional geometry and Haversian remodeling differences. Our results indicate that the human limb is not designed like that of quadrupedal cursorial animals. The tibia is not less resistant in bending and torsion, and does not remodel more than the femur. Our findings fail to support the hypothesis of Lieberman and Crompton (1998) and suggest, instead, that the human lower limb is not designed like a cursorial animal limb. In addition, our results support previous observations that remodeling is not uniform within the cross section of a bone, probably a reflection of different loading histories within the different regions of the cross section.     

The impact of incorporation of n-3 fatty acids into eggs on ovarian follicular development, immune response, antioxidative status and tibial bone characteristics in aged laying hens

The objectives of this study were to investigate the effects of different dietary sources of unsaturated fatty acids (fish oil (FO) and/or linseed oil (LO)) on laying performance, egg yolk fatty acid composition, ovarian follicular development, antioxidative properties, immune response and tibial bone characteristics in aged laying hens. A total of 100 Hisex Brown hens at 56 weeks of age were housed individually in laying cages in an open-sided building under a 16 h light:8 h dark lighting schedule. Hens were randomly divided into four experimental treatments (n=25 each). Birds were fed ad libitum diets containing 2.5% vegetable oil (C, control), 2.5% FO, 2.5% LO and a mixture of 1.25% LO+1.25% FO (LO+FO) from 56 to 68 weeks of age. Egg production, egg quality characteristics and yolk lipid profile were analyzed. At 68 weeks of age, ovarian follicles were classified and tibial bone characteristics were determined. Serum thiobarbituric acid reactive substance (TBARS), glutathione peroxidase (GSH-Px) activity and total antioxidant capacity were measured. Incorporation of n-3 polyunsaturated fatty acids (n-3PUFA) into the egg yolks has been successful by using dietary FO and/or LO. There were no significant effects of treatments on hen-day egg production, feed intake, egg weight, egg shape index, albumen height, Haugh units and yolk height. However, dietary FO and/or LO supplementation had a significantly positive effect on eggshell percentage, eggshell thickness and yolk color. At 68 weeks of age, there was no significant difference among dietary treatments for tibial bone measurements. Also, no negative effects were detected in ovarian follicular development and weights of the ovary and oviduct, expressed in both absolute terms and relative to body weight. Dietary 2.5% LO, 2.5% FO and a mixture of 1.25% FO+1.25% LO enhanced GSH-Px activity, total antioxidant capacity and antibody titers significantly in comparison with control. It could be concluded that inclusion of mixed sources of n-3PUFA in diets at moderate levels (2.5%) increased the n-3PUFA content and decreased the n-6/n-3 ratio content in the yolk, improved the antioxidative status, reduced lipid peroxidation, enhanced the antibody response and did not have any negative influence on ovarian follicular development and tibial bone characteristics in aged laying hens.     

Periosteal versus true cross‐sectional geometry: A comparison along humeral,femoral, and tibial diaphyses

Cross‐sectional geometric (CSG) properties of human long bone diaphyses are typically calculated from both periosteal and endosteal contours. Though quantification of both is desirable, periosteal contours alone have provided accurate predictions of CSG properties at the midshaft in previous studies. The relationship between CSG properties calculated from external contours and “true” (endosteal and periosteal) CSG properties, however, has yet to be examined along the whole diaphysis. Cross‐sectional computed tomography scans were taken from 21 locations along humeral, femoral, and tibial diaphyses in 20 adults from a late prehistoric central Illinois Valley cemetery. Mechanical properties calculated from images with (a) artificially filled medullary cavities (“solid”) and (b) true unaltered cross‐sections were compared at each section location using least squares regression. Results indicate that, in this sample, polar second moments of area (J), polar section moduli (Z_p), and cross‐sectional shape (I_max/I_min) calculated from periosteal contours correspond strongly with those calculated from cross‐sections that include the medullary cavity. Correlations are high throughout most of the humeral diaphysis and throughout large portions of femoral and tibial diaphyses (R² = 0.855–0.998, all P < 0.001, %SEE ≤ 8.0, %PE ≤ 5.0), the major exception being the proximal quarter of the tibial diaphysis for J and Z_p. The main source of error was identified as variation in %CA. Results reveal that CSG properties quantified from periosteal contours provide comparable results to (and are likely to detect the same differences among individuals as) true CSG properties along large portions of long bone diaphyses. Am J Phys Anthropol, 2013. © 2013 Wiley Periodicals, Inc.