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Massive bone allografts enable the reconstruction of critical bone defects in numerous conditions (e.g. tumoral, infection or trauma). Unfortunately, their biological integration remains insufficient and the reconstruction may suffer from several postoperative complications. Perfusion-decellularization emerges as a tissue engineering potential solution to enhance osseointegration. Therefore, an intrinsic vascular study of this novel tissue engineering tool becomes essential to understand its efficacy and applicability. 32 porcine long bones (humeri and femurs) were used to assess the quality of their vascular network prior and after undergoing a perfusion-decellularization protocol. 12 paired bones were used to assess the vascular matrix prior (N = 6) and after our protocol (N = 6) by immunohistochemistry. Collagen IV, Von Willebrand factor and CD31 were targeted then quantified. The medullary macroscopic vascular network was evaluated with 12 bones: 6 were decellularized and the other 6 were, as control, not treated. All 12 underwent a contrast-agent injection through the nutrient artery prior an angio CT-scan acquisition. The images were processed and the length of medullary vessels filled with contrast agent were measured on angiographic cT images obtained in control and decellularized bones by 4 independent observers to evaluate the vascular network preservation. The microscopic cortical vascular network was evaluated on 8 bones: 4 control and 4 decellularized. After injection of gelatinous fluorochrome mixture (calcein green), non-decalcified fluoroscopic microscopy was performed in order to assess the perfusion quality of cortical vascular lacunae. The continuity of the microscopic vascular network was assessed with Collagen IV immunohistochemistry (p -value = 0.805) while the decellularization quality was observed through CD31 and Von Willebrand factor immunohistochemistry (p -values <0.001). The macroscopic vascular network was severely impaired after perfusion-decellularization; nutrient arteries were still patent but the amount of medullary vascular channels measured was significantly higher in the control group compared to the decellularized group (p -value <0.001). On average, the observers show good agreement on these results, except in the decellularized group where more inter-observer discrepancies were observed. The microscopic vascular network was observed with green fluoroscopic signal in almost every canals and lacunae of the bone cortices, in three different bone locations (proximal metaphysis, diaphysis and distal metaphysis). Despite the aggressiveness of the decellularization protocol on medullary vessels, total porcine long bones decellularized by perfusion retain an acellular cortical microvascular network. By injection through the intact nutrient arteries, this latter vascular network can still be used as a total bone infusion access for bone tissue engineering in order to enhance massive bone allografts prior implantation. [Display omitted] • This study shows how a total bone can be used as a pediculed whole organ for tissue engineering purpose. • This study brings insights in the understanding of total bone decellularization by perfusion. • This study is a step forward a new generation of massive bone allografts. • This study reinforces the importance and complexity of bone vascularization. [ABSTRACT FROM AUTHOR]

High-resolution peripheral quantitative computed tomography (HR-pQCT) is a non-invasive method of measuring volumetric bone mineral density (vBMD) and microarchitecture at the distal radius and tibia. With increasing use of this technology, it is crucial to understand the potential impact of overlying soft tissue on the accuracy of HR-pQCT measures. Thus, we examined the effects of a simulated increase in adiposity (via 6- and 12-mm thick layers of overlying circumferential fat) on HR-pQCT measures of a hydroxyapatite (HA) phantom and in women (n = 20, aged 18–75 years). In the phantom, increasing the amount of overlying fat tissue led to a corresponding decrease in the mean measured density for each HA rod. In women, fat-layering led to a decrease in total vBMD (−2.9 to −3.7%, p < 0.001), cortical vBMD (−1.4% to −5.5%, p < 0.001), and estimated failure load (−1.4 to −5.7%, p = 0.002) at the radius, with similar changes in the tibia. Trabecular microarchitectural measurements were also impacted by simulated adiposity, with fat-layering leading to decreased trabecular thickness and separation and increased trabecular number at the radius (Δ's = 5 to 12%) with more pronounced differences at the tibia (Δ's = 14 to 40%). At the tibia, fat-layering also led to decreased cortical thickness and increased cortical porosity. Altogether, these results demonstrate that overlying adipose tissue can lead to artifacts in bone measurements by HR-pQCT, resulting in an underestimation of vBMD and generally, an overestimation of bone microarchitecture impairment. Therefore, soft tissue artifact should be considered when interpreting HR-pQCT results, particularly in those with high BMI and/or marked changes in adiposity. • High-resolution peripheral QCT (HR-pQCT) is used increasingly. • Overlying adipose tissue can lead to artifacts in bone measurements by HR-pQCT. • Volumetric bone mineral density is underestimated with increased adipose tissue. • Bone microarchitecture impairment is overestimated with increased adipose tissue. • Soft tissue artifact should be considered when interpreting HR-pQCT results. [ABSTRACT FROM AUTHOR]

Deposition of calcium oxalate crystals in the kidney and bone is a hallmark of systemic oxalosis. Since the bone compartment can store massive amounts of oxalate, patients present with recurrent low-trauma fractures, bone deformations, severe bone pains and specific oxalate osteopathy on plain X-ray. Bone biopsy from the iliac crest displays specific features such as oxalate crystals surrounded by a granulomatous reaction due to an invasion of bone surface by macrophages. We present data obtained in 10 samples from 8 patients with oxalosis (16–68 years) who underwent iliac crest bone biopsy and bone quality analysis using modern methods (microradiography, microindentation, Fourier Transform InfraRed Microspectroscopy, transmission electron microscopy) in addition to histomorphometry. Disseminated calcium oxalate deposits (whewellite) were found in the bone marrow space (with a granulomatous reaction) but not in the bone matrix. Calcium oxalate deposits were totally surrounded by macrophages and multinucleated giant cells, and a phagocytosis activity was sometimes observed. Very few calcium oxalate crystals were directly in close contact with the mineral substance of the bone. Bone mineralization was not modified by the presence of calcium oxalate even in close vicinity. Bone quality analysis also revealed a harder bone than normal, perhaps in relationship with decreased carbonate content in the mineral. This increase in bone hardness could explain a more “brittle” bone. In patients with oxalosis, the formation and growth of calcium oxalate crystals in the bone appeared independent of apatite. The mechanisms leading to nucleation and growth of oxalate deposits are still unclear and deserve further studies. [ABSTRACT FROM AUTHOR]

The vertebral endplate and cortical shell play an important structural role and contribute to the overall strength of the vertebral body, are at highest risk of initial failure, and are involved in degenerative disease of the spine. The ability to accurately measure the thickness of these structures is therefore important, even if difficult due to relatively low resolution clinical imaging. We posit that digital tomosynthesis (DTS) may be a suitable imaging modality for measurement of endplate and cortical shell thickness owing to the ability to reconstruct multiplanar images with good spatial resolution at low radiation dose. In this study, for 25 cadaveric L1 vertebrae, average and standard deviation of endplate and cortical shell thickness were measured using images from DTS and microcomputed tomography (μCT). For endplate thickness measurements, significant correlations between DTS and μCT were found for all variables when comparing thicknesses measured in both the overall endplate volume (R2 = 0.25–0.54) and when measurements were limited to a central range of coronal or sagittal slices (R2 = 0.24–0.62). When compared to reference values from the overall shell volume, DTS thickness measurements were generally nonsignificant. However, when measurement of cortical shell thickness was limited to a range of central slices, DTS outcomes were significantly correlated with reference values for both sagittal and coronal central regions (R2 = 0.21–0.49). DTS may therefore offer a means for measurement of endplate thickness and, within a limited sagittal or coronal measurement volume, for measurement of cortical shell thickness. • Vertebral endplate and shell thickness were measured from tomosynthesis (DTS) and μCT. • DTS derived thickness was 9.4% to 21.7% less than reference values from μCT volume. • The differences were 0.71% to 16% when DTS and μCT volumes were matched. • Moderate correlations were found (R2 to 0.62 for endplate, 0.49 for shell outcomes). • DTS offers a means for measuring endplate, and shell within central analysis regions. [ABSTRACT FROM AUTHOR]

Abstract: Advanced bone imaging techniques provide structural information, beyond bone mineral density (BMD), and growing evidence indicates that BMD only partially explains bone strength and fracture resistance. Assessing glucocorticoid-induced osteoporosis (GIO) is important, especially the documentation of glucocorticoid (GC) impact on trabecular and cortical bone and on macro and microstructural features. Advanced methods for assessing macrostructure of bone include volumetric quantitative computed tomography (vQCT), high-resolution computed tomography (hrCT), and high-resolution magnetic resonance imaging (hrMRI). The methods for assessing bone microstructure include micro computed tomography (μCT) and micro magnetic resonance imaging (μMRI). Many advanced imaging techniques have been used in vitro and in vivo to examine structural effects of GIO in animals and in humans, and these applications are explored in this review. In human in vitro studies, investigators have used standard bone histomorphometry and μCT to compare trabecular microarchitecture and bone remodeling in postmenopausal women and in males with GIO, and have found that high-dose GC produces dramatic bone loss, accompanied by major reduction in trabecular connectivity and increases in trabecular perforations. In animal studies, investigators have used standard histomorphometry along with pQCT, vQCT, hrMRI or μCT to examine GIO in a variety of animal models including rats, minipigs and sheep. They generally have found excellent relationships between treatment-induced structural changes assessed by these advanced imaging techniques and changes in BMD and biomechanical properties. They also have examined various therapeutic interventions in animals and monitored their efficacy using quantitative imaging methods. In human in vivo studies, investigators have serially examined postmenopausal women and males with GIO in order to assess the extent of skeletal deterioration and to determine the best advanced measures of BMD and structure, with which to monitor disease activity and therapeutic response, and to predict fracture risk. They generally have found that bone density and structural measures obtained by pQCT, vQCT and hrMRI contributed substantially to understanding the skeletal effects of glucocorticoids and to predicting the risk of fracture in human GIO. These animal and human applications, illustrating advanced imaging in GIO, are still in early stages of development. However, as discussed in this review, the novelty and power of the imaging approaches are compelling, and their utility is promising. [Copyright &y& Elsevier]

Abstract: An accurate analysis and quantification of microdamage is critical to understand how microdamage affects the mechanics and biology of bone fragility. In this study we demonstrate the development and validation of a novel in vitro micro-computed tomography (microCT) method that employs lead–uranyl acetate as a radio-opaque contrast agent for automated quantification of microdamage in trabecular bone. Human trabecular bone cores were extracted from the femoral neck, scanned via microCT, loaded in unconfined compression to a range of apparent strains (0.5% to 2.25%), stained in lead–uranyl acetate, and subsequently re-scanned via microCT. An investigation of the regions containing microdamage using the backscatter mode of a scanning electron microscope (BSEM) showed that the lead–uranyl sulfide complex was an effective contrast agent for microdamage in bone. Damaged volume fraction (DV/BV), as determined by microCT, increased exponentially with respect to applied strains and proportionately to mechanically determined modulus reduction (p <0.001). Furthermore, the formation of microdamage was observed to occur before any apparent stiffness loss, suggesting that the localized tissue yielding occurs prior to the structural yielding of trabecular bone. This non-invasive in vitro technique for the detection of microdamage using microCT may serve as a valuable complement to existing morphometric analyses of bone. [Copyright &y& Elsevier]