A novel methodological approach to simultaneously extract high-quality total RNA and proteins from cortical and trabecular bone.

Molecular differences between cortical and trabecular bone, of relevance to understanding the pathophysiological basis of bone diseases, can be determined only through effective isolation methods for RNA and proteins. Here we present a TRIzol-based method, which combines bone pulverization and homogenization to extract simultaneously total RNA and proteins from human cortical and trabecular bone from the same carrot. RNA integrity and purity were determined as the 260/280 nm and 260/230 nm absorbance ratios and the 28S/18S rRNA ratio. Protein integrity and quality were evaluated by Coomassie blue staining. Reverse transcription quantitative polymerase chain reaction and immunoblotting for bone-specific genes and proteins were performed to verify the suitability of the isolated material in downstream applications. The 260/280 nm and 260/230 nm absorbance ratios were, on average, less than or equal to 1.8. Bands on agarose gel were consistent with intact RNA, with mean 28S/18S ratios of 1.68 ± 0.35 and 1.88 ± 0.10 for cortical and trabecular bone, respectively. Band patterns after Coomassie blue staining confirmed protein integrity. Successful gene and protein expression analysis, with relevant differences between the two compartments, highlighted the suitability of the material in downstream applications. The method presented here is appropriate and effective for the study of human bone.

Effects of bovine cancellous bone powder/poly amino acid composites on cellular behaviors and osteogenic performances.

Xenogeneic bone has good biological activity, but eliminating immunogenicity, while retaining osteogenic abilities, is a challenge. By combining xenogeneic bone with poly amino acid (PAA) that has an amide bond structure, a new type of composite conforming to bionics and low immunogenicity may be obtained. In this study, according to the principles of component bionics, three composites of delipidized cancellous bone powder (DCBP) and PAA were designed and obtained by anin situpolycondensation method, an extrusion molding (EM) method, and a solution-blend method. The three composites were all macroscopically uniform, non-cytotoxic, and demonstrated low immunogenicity by effective removal of residual antigens during preparation. Compared with PAA, mouse bone marrow mesenchymal stem cells (BMSCs) on the surfaces of three composites showed different cellular morphologies. The effects of different preparation methods and cellular morphology on cellular differentiation were confirmed by alkaline phosphatase activity, calcium nodule formation and the expression levels of osteogenic differentiation-related genes (bone morphogenetic protein 2, runt-related transcription factor 2, osteopontin and osteocalcin). Among these composites, DCBP/PAA EM showed best cell proliferation and osteogenic differentiationin vitro, and possessed greater bone formation than PAA in a rabbit femoral condyle study. This study may provide a new method for preparing bioactive bone repair materials with low immunogenicity and superior ability to stimulate differentiation of BMSCsin vitroand osteogenesisin vivo. DCBP/PAA EM might be a promising bone repair material for bone defect treatment.

A new cancellous bone material of silk fibroin/cellulose dual network composite aerogel reinforced by nano-hydroxyapatite filler.

Composites materials comprised of biopolymeric aerogel matrices and inorganic nano-hydroxyapatite (n-HA) fillers have received considerable attention in bone engineering. Although with significant progress in aerogel-based biomaterials, the brittleness and low strengths limit the application. The improvements in toughness and mechanical strength of aerogel-based biomaterials are in great need. In this work, an alkali urea system was used to dissolve, regenerate and gelate cellulose and silk fibroin (SF) to prepare composite aerosol. A dual network structure was shaped in the composite aerosol materials interlaced by sheet-like SF and reticular cellulose wrapping n-HA on the surface. Through uniaxial compression, the density of the composite aerogel material was close to the one of natural bone, and mechanical strength and toughness were high. Our work indicates that the composite aerogel has the same mechanical strength range as cancellous bone when the ratio of cellulose, n-HA and SF being 8:1:1. In vitro cell culture showed HEK-293T cells cultured on composite aerogels had high ability of adhesion, proliferation and differentiation. Totally, the presented biodegradable composite aerogel has application potential in bone tissue engineering.

Study on mechanical properties and permeability of elliptical porous scaffold based on the SLM manufactured medical Ti6Al4V.

In this paper, we take the elliptical pore structure which is similar to the microstructure of cancellous bone as the research object, four groups of bone scaffolds were designed from the perspective of pore size, porosity and pore distribution. The size of the all scaffolds were uniformly designed as 10 × 10 × 12 mm. Four groups of model samples were prepared by selective laser melting (SLM) and Ti6Al4V materials. The statics performance of the scaffolds was comprehensively evaluated by mechanical compression simulation and mechanical compression test, the manufacturing error of the scaffold samples were evaluated by scanning electron microscope (SEM), and the permeability of the scaffolds were predicted and evaluated by simulation analysis of computational fluid dynamics (CFD). The results show that the different distribution of porosity, pore size and pores of the elliptical scaffold have a certain influence on the mechanical properties and permeability of the scaffold, and the reasonable size and angle distribution of the elliptical pore can match the mechanical properties and permeability of the elliptical pore scaffold with human cancellous bone, which has great potential for research and application in the field of artificial bone scaffold.

Preparation of a novel bovine cancellous bone/poly-amino acid composite with low immunogenicity, proper strength, and cytocompatibility in vitro.

In this work, the delipidized and deproteinized bovine cancellous bone powder/poly-amino acid (DDBP/PAA) composite was fabricated by extrusion-injection molding method for the first time. After about 70% clearance rate by the delipidization and deproteinization procedures, the residual antigens of galactosyl α-(1, 3)-galactosyl ß-1,4-N-aeetylglueosaminyl (α-Gal) and major histocompatibility complex (MHC) II were basically eliminated by the extrusion-injection molding process, which may cause high titer of antibody and lead to hyperacute rejection or chronic immune toxicity. Meanwhile, the natural BMP II and apatite in bovine bone were kept in DDBP/PAA composite. After 26 weeks of immersion in simulated body fluid, the DDBP/PAA composite remained the intact appearance, 96.4% of weight, and 69.2% of compressive strength, and these showed sufficient degradation stability. The composite also exhibited excellent attachment and proliferation abilities of mouse bone marrow mesenchymal stem cells (mMSCs). The results herein suggested that the DDBP/PAA composite was expected to be a load-bearing transplant with some natural ingredients for hard tissue repair.

Quantitative Magnetic Resonance Imaging of Cortical and Trabecular Bone.

Bone is a composite material consisting of mineral, organic matrix, and water. Water in bone can be categorized as bound water (BW), which is bound to bone mineral and organic matrix, or as pore water (PW), which resides in Haversian canals as well as in lacunae and canaliculi. Bone is generally classified into two types: cortical bone and trabecular bone. Cortical bone is much denser than trabecular bone that is surrounded by marrow and fat. Magnetic resonance (MR) imaging has been increasingly used for noninvasive assessment of both cortical bone and trabecular bone. Bone typically appears as a signal void with conventional MR sequences because of its short T2*. Ultrashort echo time (UTE) sequences with echo times 100 to 1,000 times shorter than those of conventional sequences allow direct imaging of BW and PW in bone. This article summarizes several quantitative MR techniques recently developed for bone evaluation. Specifically, we discuss the use of UTE and adiabatic inversion recovery prepared UTE sequences to quantify BW and PW, UTE magnetization transfer sequences to quantify collagen backbone protons, UTE quantitative susceptibility mapping sequences to assess bone mineral, and conventional sequences for high-resolution imaging of PW as well as the evaluation of trabecular bone architecture.

Timing of Antimicrobial Prophylaxis and Tourniquet Inflation: A Randomized Controlled Microdialysis Study.

BACKGROUND: Tourniquets are widely used during extremity surgery. In order to prevent surgical site infection, correct timing of antimicrobial prophylaxis and tourniquet inflation is important. We aimed to evaluate the time for which the free drug concentration of cefuroxime is maintained above the minimum inhibitory concentration (t > MIC) in porcine subcutaneous adipose tissue and calcaneal cancellous bone during 3 clinically relevant tourniquet application scenarios. METHODS: Twenty-four female Danish Landrace pigs were included. Microdialysis catheters were placed bilaterally for sampling of cefuroxime concentrations in calcaneal cancellous bone and subcutaneous adipose tissue, and a tourniquet was applied to a randomly picked leg of each pig. Subsequently, the pigs were randomized into 3 groups to receive 1.5 g of cefuroxime by intravenous injection 15 minutes prior to tourniquet inflation (Group A), 45 minutes prior to tourniquet inflation (Group B), and at the time of tourniquet release (Group C). The tourniquet duration was 90 minutes in all groups. Dialysates and venous blood samples were collected for 8 hours after cefuroxime administration. Cefuroxime and various ischemic marker concentrations were quantified. RESULTS: Cefuroxime concentrations were maintained above the clinical breakpoint MIC for Staphylococcus aureus (4 µg/mL) in calcaneal cancellous bone and subcutaneous adipose tissue throughout the 90-minute tourniquet duration in Groups A and B. Cefuroxime administration at the time of tourniquet release (Group C) resulted in concentrations of >4 µg/mL for approximately of 3.5 hours in the tissues on the tourniquet side. Furthermore, tourniquet application induced ischemia (increased lactate:pyruvate ratio) and cell damage (increased glycerol) in subcutaneous adipose tissue and calcaneal cancellous bone. Tissue ischemia was sustained for 2.5 hours after tourniquet release in calcaneal cancellous bone. CONCLUSIONS: Administration of cefuroxime (1.5 g) in the 15 to 45-minute window prior to tourniquet inflation resulted in sufficient concentrations in calcaneal cancellous bone and subcutaneous adipose tissue throughout the 90-minute tourniquet application. Furthermore, tourniquet-induced tissue ischemia fully resolved 2.5 hours after tourniquet release. CLINICAL RELEVANCE: Cefuroxime administration 15 to 45 minutes prior to tourniquet inflation seems to be a safe window. If the goal is to maintain postoperative cefuroxime concentrations above relevant MIC values, our results suggest that a second dose of cefuroxime should be administered at the time of tourniquet release.

Effect of chitosan infiltration on hydroxyapatite scaffolds derived from New Zealand bovine cancellous bones for bone regeneration.

Hydroxyapatite (HA) derived from bovine bones garnered wider interest as a bone substitute due to their abundant availability as meat wastes and similarities in morphology and mineral composition to human bone. In our previous work, we developed an easy and reproducible method to prepare xenograft HA scaffolds from NZ bovine cancellous bones (BHA). However, the processing methodology rendered the material mechanically weak. The present study investigated the infiltration of chitosan (CS) into the bovine HA scaffolds (CSHA) to improve the mechanical properties of BHA. The presence of characteristic functional groups of HA and CS as detected by infrared spectroscopy confirmed the infiltration of CS into the BHA scaffolds. X-ray Diffraction study confirmed the presence of the hydroxyapatite phase in both BHA and CSHA scaffolds. SEM and µCT analyses showed the CSHA scaffolds presented adequate porosity and an interconnected porous architecture required for cell migration and attachment. CSHA scaffolds presented good thermal, chemical and structural stability while demonstrating sustained biodegradability in simulated body fluid. CSHA scaffolds presented mechanical properties significantly higher than the BHA scaffolds. CSHA scaffolds were biocompatible with Saos-2 osteoblast cells and supported cell proliferation significantly better than the BHA scaffolds indicating their potential in bone tissue engineering.

Influence of processing parameters on mechanical properties of a 3D-printed trabecular bone microstructure.

Natural bone microstructure has shown to be the most efficient choice for the bone scaffold design. However, there are several process parameters involved in the generation of a microCT-based 3D-printed (3DP) bone. In this study, the effect of selected parameters on the reproducibility of mechanical properties of a 3DP trabecular bone structure is investigated. MicroCT images of a distal radial sample were used to reconstruct a 3D ROI of trabecular bone. Nine tensile tests on bulk material and 54 compression tests on 8.2 mm cubic samples were performed (9 cases × 6 specimens/case). The effect of input-image resolution, STL mesh decimation, boundary condition, support material, and repetition parameters on the weight, elastic modulus, and strength were studied. The elastic modulus and the strength of bulk material showed consistent results (CV% = 9 and 6%, respectively). The weight, elastic modulus, and strength of the cubic samples showed small intragroup variation (average CV% = 1.2, 9, and 5.5%, respectively). All studied parameters had a significant effect on the outcome variables with less effect on the weight. Utmost care to every step of the 3DP process and involved parameters is required to be able to reach the desired mechanical properties in the final printed specimen. © 2019 The Authors. Journal of Biomedical Materials Research Part B: Applied Biomaterials published by Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 108B:38-47, 2020.

Characterization of a prevascularized biomimetic tissue engineered scaffold for bone regeneration.

Significant bone loss due to disease or severe injury can result in the need for a bone graft, with over 500,000 procedures occurring each year in the United States. However, the current standards for grafting, autografts and allografts, can result in increased patient morbidity or a high rate of failure respectively. An ideal alternative would be a biodegradable tissue engineered graft that fulfills the function of bone while promoting the growth of new bone tissue. We developed a prevascularized tissue engineered scaffold of electrospun biodegradable polymers PLLA and PDLA reinforced with hydroxyapatite, a mineral similar to that found in bone. A composite design was utilized to mimic the structure and function of human trabecular and cortical bone. These scaffolds were characterized mechanically and in vitro to determine osteoinductive and angioinductive properties. It was observed that further reinforcement is necessary for the scaffolds to mechanically match bone, but the scaffolds are successful at inducing the differentiation of mesenchymal stem cells into mature bone cells and vascular endothelial cells. Prevascularization was seen to have a positive effect on angiogenesis and cellular metabolic activity, critical factors for the integration of a graft.

Which type of bone releases the most vancomycin? Comparison of spongious bone, cortical powder and cortico-spongious bone.

Bone infections can be challenging to treat and can lead to several surgeries and relapses. When a graft is needed, cavitary bone loss can be grafted with cancellous or cortical bone. Both can be used for grafting. However, the antibiotic releasing capacity of these grafts has not been compared. Which type of bone is best at releasing the most antibiotic has not been well established. The aim of this study was to determine which type of bone is best for antibiotic release when the bone is suffused with antibiotics by the surgeon. The hypothesis is that there would be a difference between the type of bone tested due to different release capacities of cortical and cancellous bone. This was an experimental study. Cortical spongy bone in chips, Spongy bone in chips and demineralized cortical bone powder were compared. For each type of bone, 5 samples were tested. Processed and decontaminated grafts were freeze-dried to be kept at room temperature. The primary endpoint was the amount of vancomycin released by the graft as it affects the concentration of antibiotic around the graft in clinical practice. The procedure for the study consisted of full graft immersion in a vancomycin solution. Then, the liquid was removed with aspiration. In order to measure the quantity of antibiotic released, the bone was put into distilled water in agitation in a heated rocker at 37 °C. After 30 min of soaking, 1 mL of the liquid was removed. The same extraction process was also carried out after 60 min soaking, 2 h, 3 h, 24 h, and 48 h. No differences were found between each type of bone relative to the concentration of vancomycin released at each time of the assessment. There was a significant difference in the weight of the bone with a higher weight for the cortical powder (1.793 g) versus cortical spongy bone and spongy bone (1.154 g and 1.013 g) with a p value < 0.0001. A significant difference was seen in the weight of the bone with vancomycin after the aspiration of the liquid with 3.026 g for cortical powder, 2.140 g and 2.049 g for the cortical spongy bone and the spongy bone with a p value < 0.0001. In daily clinical practice, one can use cancellous bone, cortico-cancellous bone or cortical powder in order to add vancomycin to a bone graft. Our results show the release kinetics of the soaked allografts. With a maximum of 14 mg/mL in the first minutes and a rapid decrease it shows a pattern comparable to antibiotic loaded bone cement. The method used appears favourable for prophylactic use, protecting the graft against contamination at implantation, but is not sufficient for treating chronic bone infection. LEVEL OF EVIDENCE: V.

Inter and intra-individual variation in skeletal DNA preservation in buried remains.

Our ability to identify skeletal remains often relies on the quality and quantity of DNA extracted from bone and teeth. Current research on buried remains has been retrospective, and no study to our knowledge has comprehensively assessed both intra-individual and inter-individual variation in human skeletal DNA from all representative skeletal element types recovered from a burial. Three individuals were interred together in a single grave for four years. Following disinterment, skeletal DNA was extracted, quantified, and GlobalFiler™ results were produced from 49 bones per skeleton, representing all bone types. Multiple sites per bone were also tested to determine intra-bone variability. Co-extracted bacterial and fungal DNA were quantified to determine microbial loads in the bones. Results show that the small, cancellous bones of the feet outperformed other bones in terms of DNA yield, measured as nanograms per gram of bone powder, and short tandem repeat (STR) profile completeness. The cuneiforms, in particular, had consistently high human DNA yields for all three individuals. DNA yield varied by individual and depth within the grave, with the shallowest individual demonstrating the highest DNA yields While the feet exhibited the greatest variation in DNA yield across bone type and sampling site, they also demonstrated some of the highest DNA yields and the most complete STR profiles, evoking a re-evaluation of their use for skeletal DNA sampling and analysis.

A Review of Bomb Pulse Dating and its Use in the Investigation of Unidentified Human Remains.

In cases where there is limited antemortem information, the examination of unidentified human remains as part of the investigation of long-term missing person's cases is a complex endeavor and consequently requires a multidisciplinary approach. Bomb pulse dating, which involves the analysis and interpretation of 14C concentration, is one technique that may assist in these investigations by providing an estimate of year of birth and year of death. This review examines the technique of bomb pulse dating and its use in the identification of differentially preserved unknown human remains. Research and case studies implementing bomb pulse dating have predominantly been undertaken in the Northern Hemisphere and have demonstrated reliable and accurate results. Limitations were, however, identified throughout the literature. These included the small sample sizes used in previous research/case studies which impacted on the statistical significance of the findings, as well as technique-specific issues. Such limitations highlight the need for future research.

Is bone loss a physiological cost of reproduction in the Great fruit-eating bat Artibeus lituratus?

During mammalian pregnancy and lactation, the maternal demand for calcium is increased to satisfy fetus and newborn skeletal growth. In addition to the dietary intake, females use the calcium contained in their bones to supply this increased demand, leading to a decrease in maternal bone mineral content. In reproductive insectivorous female bats, bone loss has been described as a physiological cost of reproduction, due to the reported increased risk of bone fracture. This physiological cost may be the mechanism underlying the conflict between increasing litter size and maintaining wing skeletal integrity, which would help to explain the small litter size of most bat species. If bone loss is a linking cost between reproduction and survival in bats, and most bat species have small litter sizes, one would expect to find a loss of bone and an increasing probability of bone fracture during pregnancy and lactation in other non-insectivorous bats. In this study, we tested for the existence of this cost in the Great-fruit eating bat, Artibeus lituratus. We analyzed trabecular structure, bone strength and bone mineral content for the humerus bone, hypothesizing that bone loss during reproduction in females would increase the risk of fracture. Our results showed a decrease of 22-31% in bone trabecular area in lactating females, rapidly compensated following weaning. Bone strength did not differ among reproductive and non-reproductive groups and seems to be more influenced by bone organic components rather than mineral contents. Since we observed bone loss during reproduction yet the humerus strength seems to be unaffected, we suggest that bone loss may not represent a physiological cost during reproduction for this frugivorous bat.

CoCr porous scaffolds manufactured via selective laser melting in orthopedics: Topographical, mechanical, and biological characterization.

Over the last decade, advances in additive manufacturing have allowed to obtain complex 3D porous lattice in materials suitable for orthopedic applications. Whereas 3D-melted titanium alloys have been extensively investigated, little is the current knowledge on the feasibility of bone-replicating CoCr porous scaffolds manufactured via selective laser melting (SLM). Moreover, the effect of topography on bone cells viability and proliferation has not been fully explored yet. Small cylindrical porous lattices were modeled from micro-CT images of human trabecular bone, and from the repetition of spherical-hollow and body-centered cubic unit cells, and manufactured via SLM from CoCr powder. Macro- and microcharacterization of the porous samples were assessed using optical microscope, micro-CT, and SEM. The scaffolds mechanical properties, measured via ISO testing, compared well with those of the human bone. Osteoblast-like cells proliferation and viability were assessed in vitro, and compared to those cultured on a standard nonporous implant-to-bone interface, showing steady increase on all geometries over time. SEM analysis confirmed the quality of cells morphology, spread, and organization on all lattices. The SLM process appeared not to alter the biocompatibility of CoCr; however, 15-100 µm irregularities and macroalterations were observed in the porous scaffolds with respect to the 3D nominal models. © 2019 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B: 2343-2353, 2019.

Effects of dietary omega-3 fatty acids on bones of healthy mice.

BACKGROUND & AIMS: Altering the lipid component in diets may affect the incidence of metabolic bone disease in patients dependent on parenteral nutrition. Consumption of polyunsaturated fatty acids (PUFA) can impact bone health by modulating calcium metabolism, prostaglandin synthesis, lipid oxidation, osteoblast formation, and osteoclastogenesis. The aim of this study was to evaluate the dietary effects of PUFA on murine bone health. METHODS: Three-weeks-old male (n = 30) and female (n = 30) C57BL/6J mice were randomized into one of three dietary groups. The diets differed only in fat composition: soybean oil (SOY), rich in ω-6 PUFA; docosahexaenoic acid alone (DHA), an ω-3 PUFA; and DHA with arachidonic acid, an ω-6 PUFA, at a 20:1 ratio (DHA/ARA). After 9 weeks of dietary treatment, femurs were harvested for micro-computed tomographic analysis and mechanical testing via 3-point bending. Separate mice from each group were used solely for serial blood draws for measurement of biomarkers of bone formation and resorption. RESULTS: At the microstructural level, although some parameters in cortical bone reached differences that were statistically significant in female mice, these were too small to be considered biologically relevant. Similarly, trabecular bone parameters in male mice were statistically different in some dietary groups, although the biological interpretation of such subtle changes translate into a lack of effect in favor of any of the experimental diets. No differences were noted at the mechanical level and in blood-based biomarkers of bone metabolism across dietary groups within gender. CONCLUSIONS: Subtle differences were noted at the bones' microstructural level, however these are likely the result of random effects that do not translate into changes that are biologically relevant. Similarly, differences were not seen at the mechanical level, nor were they reflected in blood-based biomarkers of bone metabolism. Altogether, dietary consumption of PUFA do not seem to affect bone structure or metabolism in a healthy model of growing mice.

The Effect of Material Heterogeneity, Element Type, and Down-Sampling on Trabecular Stiffness in Micro Finite Element Models.

Preclinical and clinical bone strength predictions can be elucidated by understanding bone mechanics at a variety of hierarchical levels. As such, down-sampled micro-CT images are often used to make comparisons across image resolutions or used to reduce computational resources in micro finite element models (µFEMs). Therefore, the objectives of this study were to compare trabecular apparent modulus among (i) hexahedral and tetrahedral µFEMs, (ii) µFEMs generated from 32, 64, and 64 µm down-sampled from 32 µm µCT scans, and (iii) µFEMs with homogeneous and heterogeneous tissue moduli. Trabecular µFEMs were generated from scans at the three spatial resolutions taken from the glenoid vault of 14 cadaveric specimens. Simulated unconstrained compression was performed and used to calculate and compare the apparent modulus of each µFEM. It was found that models derived from high-resolution images that account for material heterogeneity are nearly equivalent whether hexahedral or tetrahedral elements are used. However, translation of stiffness from down-sampled scans are not equivalent to scans performed at the down-sampled resolution, or that account for trabecular material heterogeneity. Material heterogeneity is most representative of in vivo trabecular bone and to accurately model trabecular mechanical properties, material heterogeneity should be considered in future µFEM development.

Consumption of sheep milk compared to cow milk can affect trabecular bone ultrastructure in a rat model.

Cow milk has a positive effect on bone health due to the effects of its protein, fat, lipid, vitamin, and mineral components. Sheep milk contains a unique composition of all of these components. However, to the best of our knowledge the benefits of sheep milk in relation to bone development have not been studied. The aim of the present study was to determine the effects of the consumption of sheep milk in comparison to cow milk on the structural and physical characteristics of growing bone in a rat model. Newly weaned male rats were fed either cow milk, sheep milk or sheep milk diluted to the same total solids content as cow milk for 28 days in addition to a basal chow. At the end of the feeding period animals were euthanized, the femora harvested and stored. The femora were analysed by µ-CT, mechanical bending tests, and ICP-MS. Rats consuming sheep milk in the trial were found to have significantly higher trabecular bone surface density and trabecular bone surface to volume ratio (p < 0.05) when compared to the rats consuming cow milk. No significant differences were observed in the mechanical properties and the mineral composition of the bones (p > 0.05). With the exception of Rb, which was found to be present in higher concentrations in rats consuming sheep milk (p < 0.05). Our results indicate that consumption of sheep milk may positively influence the structural integrity of bone, which may result in an enhancement of bone health.

Effect of the trabecular bone microstructure on measuring its thermal conductivity: A computer modeling-based study.

The objective of this work is to quantify the relation between the value of the effective thermal conductivity of trabecular bone and its microstructure and marrow content. The thermal conductivity of twenty bovine trabecular bone samples was measured prior to and after defatting at 37, 47, and 57 °C. Computer models were built including the microstructure geometry and the gap between the tissue and measurement probe. The thermal conductivity (k) measured was 0.39 ±â€¯0.06 W m-1 K-1 at 37 °C, with a temperature dependence of + 0.2%°C-1. Replacing marrow by phosphate-buffered saline (defatting) increased both the computer simulations and measurement results by 0.04 W m-1 K-1. The computer simulations showed that k increases by 0.02-0.04 W m-1 K-1 when the model includes a gap filled by phosphate-buffered saline between the tissue and measurement probe. In the presence of microstructure and fatty red marrow, k varies by ±â€¯0.01 W m-1 K-1 compared with the case considering matrix only, which suggests that there are no significant differences between cortical and trabecular bone in terms of k. The computer results showed that the presence of a gap filled by phosphate-buffered saline around the energy applicator changes maximum temperature by < 0.7 °C, while including the bone microstructure involved a variation of < 0.2 mm in the isotherm location. Future experimental studies on measuring the value of k involving the insertion of a probe into the bone through a drill hole should consider the bias found in the simulations. Thermal models based on a homogeneous geometry (i.e. ignoring the microstructure) could provide sufficient accuracy.

Dynamic structure and composition of bone investigated by nanoscale infrared spectroscopy.

Bone is a highly organized tissue in which each structural level influences the macroscopic and microscopic mechanical behavior. In particular, the quantity, quality, and distribution of the different bone components, i.e. collagen matrix and hydroxyapatite crystals, are associated with bone strength or fragility. Common spectroscopic techniques used to assess bone composition have resolutions limited to the micrometer range. In this study, our aims were two-fold: i) to develop and validate the AFM-IR methodology for skeletal tissues and ii) to apply the methodology to sheep cancellous bone with the objective to obtain novel findings on the composition and structure of trabecular packets.To develop the methodology, we assessed spatial and temporal reproducibility using a known homogeneous material (polymethylmethacrylate, PMMA). We verified that the major peak positions were similar and not shifted when compared to traditional Fourier Transform Infrared imaging (FTIRI). When AFM-IR was applied to sheep cancellous bone, the mineral-to-matrix ratio increased and the acid phosphate substitution ratio decreased as a function of tissue maturity. The resolution of the technique enabled visualization of different stages of the bone maturation process, particularly newly-formed osteoid prior to mineralization. We also observed alternating patterns of IR parameters in line and imaging measurements, suggesting the apposition of layers of alternating structure and / or composition that were not visible with traditional spectroscopic methods. In conclusion, nanoscale IR spectroscopy demonstrates novel compositional and structural changes within trabecular packets in cancellous bone. Based on these results, AFM-IR is a valuable tool to investigate cancellous bone at the nanoscale and, more generally, to analyze small dynamic areas that are invisible to traditional spectroscopic methods.