Bone matrix mineralization and osteocyte lacunae characteristics in patients with chronic kidney disease - mineral bone disorder (CKD-MBD).

OBJECTIVES: Little is known about bone mineralization and osteocyte lacunae properties in chronic kidney disease mineral bone disorder (CKD-MBD). METHODS: In this retrospective study, we measured the bone mineralization density distribution (BMDD) and osteocyte lacunar section (OLS) 2D-characteristics by quantitative backscatter electron imaging in Straumann drill biopsy samples from n=58 patients with CKD-MBD. Outcomes were studied in relation to serum parathyroid hormone (PTH), alkaline phosphatase (APH), histomorphometric bone turnover and treatment with cinacalcet or phosphate binders. RESULTS: Lower calcium concentrations in bone from high turnover (average degree of bone mineralization -6.2%, p<0.001) versus low turnover patients were observed. OLS-characteristics were distinctly different (p<0.01 to p<0.05) in patients with highest compared to those with lowest turnover. Patients with cinacalcet had different OLS-characteristics (p<0.05) compared to those without cinacalcet. Furthermore, patients with phosphate binders had differences in BMDD and OLS-characteristics (p<0.05) compared to patients without phosphate binders. CONCLUSIONS: Our findings suggest that in patients with CKD-MBD secondary hyperparathyroidism and increased bone turnover decrease the average degree of bone matrix mineralization. Conversely, density and lacunar size of the osteocytes are increased compared to adynamic bone disease pointing at distinct patterns of bone mineralization and osteocyte lacunar properties in these two disease entities.

Consideration of bone regeneration effect of stem cells: comparison between adipose-derived stem cells and demineralized bone matrix.

BACKGROUND: Currently, many studies have sought to address the regeneration of extensive bone defects using stem cells. Here, the authors injected adipose-derived stem cells and demineralized bone matrix (DBM) into areas of bone defect in rabbits and compared their effect on bone regeneration to study the clinical usefulness of stem cells. METHODS: This study used 20 male New Zealand white rabbits. Four craniectomies were made in 20 male New Zealand white rabbits' calvaria, and 4 different groups of experimental conditions were applied to each of the 4 cranial defects. To the first group, 0.2 mL of DBX, a commercially available clinical preparation ofDBM, was applied with fibrin glue. To the second group, 0.2 mL of adipose-derived stem cells, with confirmed bone differentiation ability, was applied with fibrin glue. To the third group, 0.1 mL of DBX, 0.1 mL of adipose-derived stem cells, and fibrin glue were applied. The fourth group of defects acted as the control and was left unaltered. After 6 weeks, regenerated bone from each defect site in each rabbit was collected and measured for volume change. Bone regeneration was assessed with three-dimensional skull bone computed tomography and histological analysis. RESULTS: Osteoblasts were confirmed in all defect groups after 6 weeks. Overall, bone regeneration was weakest in the control group, whereas other groups of defects showed distinct bone regeneration. In particular, group 3, to which adipose-derived stem cells and DBM were applied, demonstrated the most active regeneration. CONCLUSIONS: Both adipose-derived stem cells and DBM demonstrated regeneration effect on cranial defects in rabbits, but it is difficult to conclude which was better, because in each case the amount of regenerated bone was within the margin of error. However, as the most active bone regeneration was observed when both adipose-derived stem cells and DBM were applied together, this combination could be helpful in the correction of extensive bone defects.

Effects of 3 years treatment with once-yearly zoledronic acid on the kinetics of bone matrix maturation in osteoporotic patients.

UNLABELLED: Once-yearly administration of intravenous zoledronic acid for 3 years in humans affects the kinetics of matrix filling in by mineral, independent of bone turnover. INTRODUCTION: Yearly 5-mg infusions of zoledronic acid (ZOL) for 3 years have shown pronounced antifracture efficacy. The purpose of the present study was to test whether ZOL affects the kinetics of forming bone material properties maturation. METHODS: Iliac crest biopsies from the HORIZON-PFT clinical trial were analyzed by Raman microspectroscopy in actively bone-forming surfaces as a function of tissue age in trabecular and osteonal bone, to determine ZOL's effect on bone material quality indices maturation kinetics. RESULTS: Mineral/matrix ratio increased in both groups as a function of tissue age, at both osteonal- and trabecular-forming surfaces; ZOL exhibiting the greatest increase in the trabecular surfaces only. The proteoglycan content showed a dependency on tissue age in both trabecular and osteonal surfaces, with ZOL exhibiting lower values in the tissue age 8-22 days in the trabecular surfaces. Mineral crystallinity (crystallite length and thickness) showed a dependence on tissue age, with ZOL exhibiting lower crystallite length compared with placebo only in the 8- to 22-day-old tissue at trabecular surfaces, while crystal thickness was lower in the 1- to 5-day-old tissue at both osteonal and trabecular surfaces. CONCLUSIONS: The results of the present study suggest that once-yearly administration of intravenous ZOL for 3 years in humans does not exert any adverse effects on the evolution of bone material properties at actively forming osteonal and trabecular surfaces, while it may have a beneficial effect on the progression of the mineral-to-matrix ratio and mineral maturity bone quality indices.

No effect of short-term hypertension on bone matrix mineralization in a surgical animal model in immature rabbits.

Epidemiological studies show that arterial hypertension is associated with bone loss and an increased risk of fractures. Bone material properties are essential for bone strength. However, little is known about the effects of hypertension on bone matrix mineralization. Genetic animal models of hypertension are not ideal for studying bone matrix properties since these mutations may affect mineralization per se. The purpose of this study was to evaluate the effects of short-term hypertension on bone mineral density distribution (BMDD) using quantitative backscattered electron imaging in the proximal humerus in an established surgical model of pressure-overload cardiac hypertrophy in immature rabbits. Banding of the descending aorta was performed in 10-day-old rabbits (n = 4). Systolic blood pressure was elevated at all timepoints in the upper extremity but reached statistical significance at 5 and 6 weeks of age (+30.1% and +25.1 mm Hg; P < 0.05 each, respectively). Diastolic blood pressure was not affected. The left proximal humerus was harvested at 6 weeks of age, which is the maximum time in this animal model. Four non-operated, matched animals served as controls. Bone mineral density distribution parameters were determined in the epiphyseal and metaphyseal regions of the proximal humerus. The bone mineral density distribution parameters which describe the degree and heterogeneity of mineralization as well as the amount of low mineralized matrix showed no significant differences. Moreover no difference in bone length was found. Our study indicates that short-term elevation of blood pressure has no effects on bone matrix mineralization in this surgical model of pressure-overload cardiac hypertrophy in immature rabbits.

Severe developmental bone phenotype in ClC-7 deficient mice.

Bone development is dependent on the functionality of three essential cell types: chondrocytes, osteoclasts and osteoblasts. If any of these cell types is dysfunctional, a developmental bone phenotype can result. The bone disease osteopetrosis is caused by osteoclast dysfunction or impaired osteoclastogenesis, leading to increased bone mass. In ClC-7 deficient mice, which display severe osteopetrosis, the osteoclast malfunction is due to abrogated acidification of the resorption lacuna. This study sought to investigate the consequences of osteoclast malfunction on bone development, bone structure and bone modeling/remodeling in ClC-7 deficient mice. Bones from wildtype, heterozygous and ClC-7 deficient mice were examined by bone histomorphometry and immunohistochemistry. ClC-7 deficient mice were found to have a severe developmental bone phenotype, characterized by dramatically increased bone mass, a high content of cartilage remnants, impaired longitudinal and radial growth, as well as lack of compact cortical bone development. Indices of bone formation were reduced in ClC-7 deficient mice; however, calcein labeling indicated that mineralization occurred on most trabecular bone surfaces. Osteoid deposition had great regional variance, but an osteopetrorickets phenotype, as observed in oc/oc mice, was not apparent in the ClC-7 deficient mice. A striking finding was the presence of very large abnormal osteoclasts, which filled the bone marrow space within the ClC-7 deficient bones. The development of these giant osteoclasts could be due to altered cell fate of the ClC-7 deficient osteoclasts, caused by increased cellular fusion and/or prolonged osteoclast survival. In summary, malfunctional ClC-7 deficient osteoclasts led to a severe developmental bone phenotype including abnormally large and non-functional osteoclasts. Bone formation paremeters were reduced; however, bone formation and mineralization were found to be heterogenous and continuing.

Fragility fractures in men with idiopathic osteoporosis are associated with undermineralization of the bone matrix without evidence of increased bone turnover.

The pathogenesis of primary osteoporosis in younger individuals is still elusive. An important determinant of the biomechanical competence of bone is its material quality. In this retrospective study we evaluated bone material quality based on quantitative backscattered electron imaging to assess bone mineralization density distribution (BMDD) in bone biopsies of 25 male patients (aged 18-61 years) who sustained fragility fractures but were otherwise healthy. BMDD of cancellous bone was compared with previously established adult reference data. Complementary information was obtained by bone histomorphometry. The histomorphometric results showed a paucity of osteoblasts and osteoclasts on the bone surface in the majority of patients. BMDD revealed a significant shift to lower mineralization densities for cancellous bone values: CaMean (weighted mean Ca content, -5.9%), CaPeak (mode of the BMDD, -5.6%), and CaHigh (portion of fully mineralized bone, -76.8%) were decreased compared to normative reference; CaWidth (heterogeneity in mineralization, +18.5%) and CaLow (portion of low mineralized bone, +68.8; all P < 0.001) were significantly increased. The shift toward lower mineral content in the bone matrix in combination with reduced indices of bone formation and bone resorption suggests an inherent mineralization defect leading to undermineralized bone matrix, which might contribute to the susceptibility to fragility fractures of the patients. The alteration in bone material might be related to osteoblastic dysfunction and seems fundamentally different from that in high bone turnover osteoporosis with a negative bone balance.

Micro-CT analysis of alveolar bone healing using a rat experimental model of critical-size defects.

OBJECTIVE: This study was designed to establish a rat model of a critical size alveolar bone defect. MATERIALS AND METHODS: Standardized buccal or mesiobuccal alveolar bone defects were made around the right first mandibular molar of 12-week-old rats, and the left was used as a control. Alveolar bone healing was examined quantitatively by three-dimensional micro-computed tomographic imaging. Bone matrix production of osteoblasts and osteocytes during repair of alveolar bone defects was examined with in situ hybridization for type I collagen. RESULTS: Buccal defects were repaired significantly and the volume decreased by 88.3% in week 24, whereas mesiobuccal defects were repaired little. Osteoblasts and osteocytes expressed type I collagen in both defects in week 3 but showed little expression by week 6 and thereafter, leaving the mesiobuccal defects largely unrepaired. CONCLUSION: The mesiobuccal defect is a critical-size defect that is not ultimately repaired with bone. It may be an appropriate experimental model for investigating the effectiveness of bone regenerative agents in human alveolar bone loss.

Bone matrix hypermineralization associated with low bone turnover in a case of Nasu-Hakola disease.

Analysis of tissue from a 34-years-old male patient from Austrian origin with a history of multiple fractures associated with painful episodes over the carpal, tarsal and at the end of the long bones respectively is presented. Radiographic images and axial 3DCT scans showed widespread defects in trabecular bone architecture and ill-defined cortices over these skeletal sites in the form of discrete cystic-like lesions. Family history indicated two sisters (one half and one full biological sisters) also with a history of fractures. Whole exome sequencing revealed two heterozygous missense mutations in TYROBP (MIM 604142; NM_003332.3) gene encoding for a cell-surface adaptor protein, which is part of a signaling complex triggering activation of immune responses. It is expressed in cells of the ectoderm cell linage such as NK and dendritic cells, macrophages, monocytes, myeloid cells, microglia cells and osteoclasts. The phenotype and genotype of the patient were consistent with the diagnosis of Nasu-Hakola disease (NHD) (OMIM 221770). Investigations at the bone material level of a transiliac bone biopsy sample from the patient using polarized light microscopy and backscatter electron imaging revealed disordered lamellar collagen fibril arrangement and extensively increased matrix mineralization. These findings are the first bone material data in a patient with NHD and point toward an osteoclast defect involvement in this genetic condition.

Deletion of OPN in BSP knockout mice does not correct bone hypomineralization but results in high bone turnover.

The two SIBLING (Small Integrin Binding Ligand N-linked Glycoproteins), bone sialoprotein (BSP) and osteopontin (OPN) are expressed in osteoblasts and osteoclasts. In mature BSP knockout (KO, -/-) mice, both bone formation and resorption as well as mineralization are impaired. OPN-/- mice display impaired resorption, and OPN is described as an inhibitor of mineralization. However, OPN is overexpressed in BSP-/- mice, complicating the understanding of their phenotype. We have generated and characterized mice with a double KO (DKO) of OPN and BSP, to try and unravel their respective contributions. Despite the absence of OPN, DKO bones are still hypomineralized. The SIBLING, matrix extracellular phosphoglycoprotein with ASARM motif (MEPE) is highly overexpressed in both BSP-/- and DKO and may impair mineralization through liberation of its ASARM (Acidic Serine-Aspartate Rich MEPE associated) peptides. DKO mice also display evidence of active formation of trabecular, secondary bone as well as primary bone in the marrow-ablation repair model. A higher number of osteoclasts form in DKO marrow cultures, with higher resorption activity, and DKO long bones display a localized and conspicuous cortical macroporosity. High bone formation and resorption parameters, and high cortical porosity in DKO mice suggest an active bone modeling/remodeling, in the absence of two key regulators of bone cell performance. This first double KO of SIBLING proteins thus results in a singular, non-trivial phenotype leading to reconsider the interpretation of each single KO, concerning in particular matrix mineralization and the regulation of bone cell activity.

DMP1 Ablation in the Rabbit Results in Mineralization Defects and Abnormalities in Haversian Canal/Osteon Microarchitecture.

DMP1 (dentin matrix protein 1) is an extracellular matrix protein highly expressed in bones. Studies of Dmp1 knockout (KO) mice led to the discovery of a rare autosomal recessive form of hypophosphatemic rickets (ARHR) caused by DMP1 mutations. However, there are limitations for using this mouse model to study ARHR, including a lack of Haversian canals and osteons (that occurs only in large mammalian bones), high levels of fibroblast growth factor 23 (FGF23), and PTH, in comparison with a moderate elevation of FGF23 and unchanged PTH in human ARHR patients. To better understand this rare disease, we deleted the DMP1 gene in rabbit using CRISPR/Cas9. This rabbit model recapitulated many features of human ARHR, such as the rachitic rosary (expansion of the anterior rib ends at the costochondral junctions), moderately increased FGF23, and normal PTH levels, as well as severe defects in bone mineralization. Unexpectedly, all DMP1 KO rabbits died by postnatal week 8. They developed a severe bone microarchitecture defect: a major increase in the central canal areas of osteons, concurrent with massive accumulation of osteoid throughout all bone matrix (a defect in mineralization), suggesting a new paradigm, where rickets is caused by a combination of a defect in bone microarchitecture and a failure in mineralization. Furthermore, a study of DMP1 KO bones found accelerated chondrogenesis, whereas ARHR has commonly been thought to be involved in reduced chondrogenesis. Our findings with newly developed DMP1 KO rabbits suggest a revised understanding of the mechanism underlying ARHR. © 2019 American Society for Bone and Mineral Research.

Bone quality issues an matrix properties in OP cancellous bone.

There is increasing evidence nowadays that diseases or conditions, like osteoporosis (OP), which are conventionally defined in terms of bone quantity/mass, are also associated with concomitant changes at the bone matrix level. The present study examined the composition, density and mineral content of OP cancellous bone at the tissue level and the hardness values at the trabecular level to establish correlations between these variables. The results showed that changes in porosity (Bone volume/Tissue volume) are accompanied by changes in mineral content and in the hardness of individual trabeculae. In other words in OP there are both quantitative and qualitative effects that take place with the progress of this condition.

Mechanical behavior of metastatic vertebrae are influenced by tissue architecture, mineral content, and organic feature alterations.

Diminished vertebral mechanical behavior with metastatic involvement is typically attributed to modified architecture and trabecular bone content. Previous work has identified organic and mineral phase bone quality changes in the presence of metastases, yet limited work exists on the potential influence of such tissue level modifications on vertebral mechanical characteristics. This work seeks to determine correlations between features of bone (structural and tissue level) and mechanical behavior in metastatically involved vertebral bone. It is hypothesized that tissue level properties (mineral and organic) will improve these correlations beyond architectural properties and BMD alone. Twenty-four female athymic rats were inoculated with HeLa or Ace-1 cancer cells lines producing osteolytic (N = 8) or mixed (osteolytic/osteoblastic, N = 7) metastases, respectively. Twenty-one days post-inoculation L1-L3 pathologic vertebral motion segments were excised and µCT imaged. 3D morphometric parameters and axial rigidity of the L2 vertebrae were quantified. Sequential loading and µCT imaging measured progression of failure, stiffness and peak force. Relationships between mechanical testing (whole bone and tissue-level) and tissue-level material property modifications with metastatic involvement were evaluated utilizing linear regression models. Osteolytic involvement reduced vertebral trabecular bone volume, structure, CT-derived axial rigidity, stiffness and failure force compared to healthy controls (N = 9). Mixed metastases demonstrated similar trends. Previously assessed collagen cross-linking and proline-based residues were correlated to mechanical behavior and improved the predictive ability of the regression models. Similarly, collagen organization improved predictive regression models for metastatic bone hardness. This work highlights the importance of both bone content/architecture and organic tissue-level features in characterizing metastatic vertebral mechanics. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:3013-3022, 2018.

Cortical Matrix Mineral Density Measured Noninvasively in Pre- and Postmenopausal Women and a Woman With Vitamin D-Dependent Rickets.

Reduced bone mineral density (BMD) may be due to reduced mineralized bone matrix volume, incomplete secondary mineralization, or reduced primary mineralization. Because bone biopsy is invasive, we hypothesized that noninvasive image acquisition at high resolution can accurately quantify matrix mineral density (MMD). Quantification of MMD was confined to voxels attenuation photons above 80% of that produced by fully mineralized bone matrix because attenuation at this level is due to variation in mineralization, not porosity. To assess accuracy, 9 cadaveric distal radii were imaged at a voxel size of 82 microns using high-resolution peripheral quantitative computed tomography (HR-pQCT; XtremeCT, Scanco Medical AG, Bruttisellen, Switzerland) and compared with VivaCT 40 (µCT) at 19-micron voxel size. Associations between MMD and porosity were studied in 94 healthy vitamin D-replete premenopausal women, 77 postmenopausal women, and in a 27-year-old woman with vitamin D-dependent rickets (VDDR). Microstructure and MMD were quantified using StrAx (StraxCorp, Melbourne, Australia). MMD measured by HR-pQCT and µCT correlated (R = 0.87; p < 0.0001). The precision error for MMD was 2.43%. Cortical porosity and MMD were associated with age (r2 = 0.5 and -0.4, respectively) and correlated inversely in pre- and postmenopausal women (both r2 = 0.9, all p < 0.001). Porosity was higher, and MMD was lower, in post- than in premenopausal women (porosity 40.3% ± 7.0 versus 34.7% ± 3.5, respectively; MMD 65.4% ± 1.8 versus 66.6% ± 1.4, respectively, both p < 0.001). In the woman with VDDR, MMD was 5.6 SD lower and porosity was 5.6 SD higher than the respective trait means in premenopausal women. BMD was reduced (Z-scores femoral neck -4.3 SD, lumbar spine -3.8 SD). Low-radiation HR-pQCT may facilitate noninvasive quantification of bone's MMD and microstructure in health, disease, and during treatment. © 2018 American Society for Bone and Mineral Research.

Bone deficit in ovariectomized rats. Functional contribution of the marrow stromal cell population and the effect of oral dihydrotachysterol treatment.

This study investigates the proliferative and osteogenic role of marrow stromal/osteoprogenitor cells in the development of the cortical bone deficit in ovariectomized (OVX) female rats. In vitro, clonal growth of marrow stromal cells from OVX rats was significantly impaired (vs. sham-operated controls). Yet in vivo, cells from sham-operated and OVX rats had equal osteogenic potential in several in vivo experimental situations, such as in intraperitoneally implanted millipore diffusion chambers and in intramuscular implants of marrow plus osteoinductive bone matrix (composite grafts). Long-term (6 mo) dihydrotachysterol (DHT) treatment of OVX rats enhanced their in vitro proliferative potential and clonal growth, as well as their osteogenic expression in composite grafts. The observation that the in vivo osteogenic performance of OVX rat marrow stromal cells was normal at extraosseous sites suggests that the mechanisms leading to osteopenia may involve an abnormality in cell-matrix interactions.

Short-term aluminum administration in the rat. Effects on bone formation and relationship to renal osteomalacia.

Aluminum may be pathogenic in the osteomalacia observed in some patients receiving hemodialysis. To study the early effects of Al on bone growth, bone formation, mineralization, and resorption were measured during short-term Al exposure in the tibial cortex of pair-fed control (C, n = 10), aluminum-treated (AL, n = 9), subtotally nephrectomized control (NX-C, n = 7), and subtotally nephrectomized aluminum-treated (NX-AL, n = 8) rats using double tetracycline labeling of bone. Animals received 2 mg/d of elemental Al intraperitoneally for 5 d/wk over 4 wk. Total bone and matrix (osteoid) formation, periosteal bone and matrix formation, and periosteal bone and matrix apposition fell by 20% in AL from C, P less than 0.05 for all values, and by 40% in NX-AL from NX-C, P less than 0.01 for all values. Moreover, each measurement was significantly less in NX-AL than in AL, P less than 0.05 for all values. Osteoid width did not increase following aluminum administration in either AL or NX-AL. Resorption surface increased from control values in both AL and NX-AL; also, resorptive activity at the endosteum was greater in NX-AL than in NX-C, P less than 0.05. Thus, aluminum impairs new bone and matrix formation but does not cause classic osteomalacia in the cortical bone of rats whether renal function is normal or reduced. These findings may represent either a different response to aluminum administration in cortical bone as contrasted to trabecular bone or an early phase in the development of osteomalacia. Aluminum may increase bone resorption and contribute to osteopenia in clinical states associated with aluminum accumulation in bone.

Effect of fatigue loading and associated matrix microdamage on bone blood flow and interstitial fluid flow.

Functional adaptation of bone to cyclic fatigue involves a complex physiological response that is targeted to sites of microdamage. The mechanisms that regulate this process are not understood, although lacunocanalicular interstitial fluid flow is likely important. We investigated the effect of a single period of cyclic fatigue on bone blood flow and interstitial fluid flow. The ulnae of 69 rats were subjected to cyclic fatigue unilaterally using an initial peak strain of -6000 muepsilon until 40% loss of stiffness developed. Groups of rats (n=23 per group) were euthanized immediately after loading, at 5 days, and at 14 days. The contralateral ulna served as a treatment control, and a baseline control group (n=23) that was not loaded was also included. After euthanasia, localization of intravascular gold microspheres within the ulna (n=7 rats/group) and tissue distribution of procion red tracer were quantified (n=8 rats/group). Microcracking, modeling, and remodeling (Cr.S.Dn, microm/mm(2), Ne.Wo.B.T.Ar, mm(2), and Rs.N/T.Ar, #/mm(2) respectively) were also quantified histologically (n=8 rats/group). Cyclic fatigue loading induced hyperemia of the loaded ulna, which peaked at 5 days after loading. There was an associated overall decrease in procion tracer uptake in both the loaded and contralateral control ulnae. Tracer uptake was also decreased in the periosteal region, when compared with the endosteal region of the cortex. Pooling of tracer was seen in microdamaged bone typically adjacent to an intracortical stress fracture at all time points after fatigue loading; in adjacent bone tracer uptake was decreased. New bone formation was similar at 5 days and at 14 days, whereas formation of resorption spaces was increased at 14 days. These data suggest that a short period of cyclic fatigue induces bone hyperemia and associated decreased lacunocanalicular interstitial fluid flow, which persists over the time period in which osteoclasts are recruited to sites of microdamage for targeted remodeling. Matrix damage and development of stress fracture also interfere with normal centrifugal fluid flow through the cortex. Changes in interstitial fluid flow in the contralateral ulna suggest that functional adaptation to unilateral fatigue loading may include a more generalized neurovascular response.

Role of matrix behavior in compressive fracture of bovine cortical bone.

In compressive fracture of dry plexiform bone, we examine the individual roles of overall mean porosity, the connectivity of the porosity network, and the elastic as well as the failure properties of the nonporous matrix, using a random spring network model (RSNM). Porosity network structure is shown to reduce the compressive strength by up to 30%. However, the load-bearing capacity increases with an increase in either of the matrix properties-the elastic modulus or the failure strain threshold. To validate the porosity-based RSNM model with available experimental data, bone-specific failure strain thresholds for the ideal matrix of similar elastic properties were estimated to be within 60% of each other. Further, we observe the avalanche size exponents to be independent of the bone-dependent parameters as well as the structure of the porosity network.

Low intensity pulsed ultrasound increases the matrix hardness of the healing tissues at bone-tendon insertion-a partial patellectomy model in rabbits.

BACKGROUND: This study evaluated the low intensity pulsed ultrasound enhancement on matrix hardness of the healing tissues at the bone-tendon junction. METHODS: Sixteen 18 week-old mature female rabbits were used. An established transverse partial patellectomy was performed at the distal one-third of the patella. Animals were then divided according to their body weight into ultrasound group (n = 8) with daily treatment of low intensity pulsed ultrasound and control group (n = 8) without ultrasound treatment. Animals were euthanized at week 8 and 16 postoperatively to evaluate the radiographic new bone formation and the Vickers hardness of the matrix of the healing tissues at the bone-tendon junction. FINDINGS: (1) Comparing with the control group, the anterior-posterior area of the new bone in the ultrasound treated group was found on average to be 3.0 and 3.1 times greater at week 8 and 16, respectively (P < 0.01). (2) The Vickers hardness of the new bone in ultrasound group was 11.3% (P < 0.05) significantly lower at week 8 but 20.0% (P < 0.05) significantly higher at week 16 as compared with that of the control group. (3) The Vickers hardness of the newly regenerated fibrocartilage zone, healing tendon, and cartilaginous metaplasia in ultrasound group was found higher than the control group at both week 8 and 16, but the difference was significant at week 16 only, being 44.1% (P < 0.05), 20.1% (P < 0.01), and 46.4% (P < 0.01) higher, respectively. INTERPRETATION: The preliminary findings suggested for the first time that low intensity pulsed ultrasound treatment resulted in the enhancement of the matrix hardness in new bone, fibrocartilage, cartilaginous metaplasia, and healing tendon at the healing bone-tendon junction. These findings can be extrapolated into clinical practice, i.e. the more rapid healing induced by low intensity pulsed ultrasound, the earlier mobilization of the affected joint. The beneficial effects on prevention of the musculoskeletal deterioration resulting from the prolonged immobilization would be therefore expected.

Multiscale alterations in bone matrix quality increased fragility in steroid induced osteoporosis.

A serious adverse clinical effect of glucocorticoid steroid treatment is secondary osteoporosis, enhancing fracture risk in bone. This rapid increase in bone fracture risk is largely independent of bone loss (quantity), and must therefore arise from degradation of the quality of the bone matrix at the micro- and nanoscale. However, we lack an understanding of both the specific alterations in bone quality n steroid-induced osteoporosis as well as the mechanistic effects of these changes. Here we demonstrate alterations in the nanostructural parameters of the mineralized fibrillar collagen matrix, which affect bone quality, and develop a model linking these to increased fracture risk in glucocorticoid induced osteoporosis. Using a mouse model with an N-ethyl-N-nitrosourea (ENU)-induced corticotrophin releasing hormone promoter mutation (Crh(-120/+)) that developed hypercorticosteronaemia and osteoporosis, we utilized in situ mechanical testing with small angle X-ray diffraction, synchrotron micro-computed tomography and quantitative backscattered electron imaging to link altered nano- and microscale deformation mechanisms in the bone matrix to abnormal macroscopic mechanics. We measure the deformation of the mineralized collagen fibrils, and the nano-mechanical parameters including effective fibril modulus and fibril to tissue strain ratio. A significant reduction (51%) of fibril modulus was found in Crh(-120/+) mice. We also find a much larger fibril strain/tissue strain ratio in Crh(-120/+) mice (~1.5) compared to the wild-type mice (~0.5), indicative of a lowered mechanical competence at the nanoscale. Synchrotron microCT show a disruption of intracortical architecture, possibly linked to osteocytic osteolysis. These findings provide a clear quantitative demonstration of how bone quality changes increase macroscopic fragility in secondary osteoporosis.