How osteons form: A quantitative hypothesis-testing analysis of cortical pore filling and wall asymmetry.

Osteon morphology provides valuable information about the interplay between different processes involved in bone remodelling. The correct quantitative interpretation of these morphological features is challenging due to the complexity of interactions between osteoblast behaviour, and the evolving geometry of cortical pores during pore closing. We present a combined experimental and mathematical modelling study to provide insights into bone formation mechanisms during cortical bone remodelling based on histological cross-sections of quiescent human osteons and hypothesis-testing analyses. We introduce wall thickness asymmetry as a measure of the local asymmetry of bone formation within an osteon and examine the frequency distribution of wall thickness asymmetry in cortical osteons from human iliac crest bone samples from women 16-78 years old. Our measurements show that most osteons possess some degree of asymmetry, and that the average degree of osteon asymmetry in cortical bone evolves with age. We then propose a comprehensive mathematical model of cortical pore filling that includes osteoblast secretory activity, osteoblast elimination, osteoblast embedment as osteocytes, and osteoblast crowding and redistribution along the bone surface. The mathematical model is first calibrated to symmetric osteon data, and then used to test three mechanisms of asymmetric wall formation against osteon data: (i) delays in the onset of infilling around the cement line; (ii) heterogeneous osteoblastogenesis around the bone perimeter; and (iii) heterogeneous osteoblast secretory rate around the bone perimeter. Our results suggest that wall thickness asymmetry due to off-centred Haversian pores within osteons, and that nonuniform lamellar thicknesses within osteons are important morphological features that can indicate the prevalence of specific asymmetry-generating mechanisms. This has significant implications for the study of disruptions of bone formation as it could indicate what biological bone formation processes may become disrupted with age or disease.

Disturbed bone marrow adiposity in patients with Cushing's syndrome and glucocorticoid- and postmenopausal- induced osteoporosis.

Background: Skeletal stem/progenitor cells (SSPCs) in the bone marrow can differentiate into osteoblasts or adipocytes in response to microenvironmental signalling input, including hormonal signalling. Glucocorticoids (GC) are corticosteroid hormones that promote adipogenic differentiation and are endogenously increased in patients with Cushing´s syndrome (CS). Here, we investigate bone marrow adiposity changes in response to endogenous or exogenous GC increases. For that, we characterize bone biopsies from patients with CS and post-menopausal women with glucocorticoid-induced osteoporosis (GC-O), compared to age-matched controls, including postmenopausal osteoporotic patients (PM-O). Methods: Transiliac crest bone biopsies from CS patients and healthy controls, and from postmenopausal women with GC-O and matched controls were analysed; an additional cohort included biopsies from women with PM-O. Plastic-embedded biopsies were sectioned for histomorphometric characterization and quantification of adipocytes. The fraction of adipocyte area per tissue (Ad.Ar/T.Ar) and marrow area (Ad.Ar/Ma.Ar), mean adipocyte profile area (Ad.Pf.Ar) and adipocyte profile density (N.Ad.Pf/Ma.Ar) were determined and correlated to steroid levels. Furthermore, the spatial distribution of adipocytes in relation to trabecular bone was characterized and correlations between bone marrow adiposity and bone remodeling parameters investigated. Results: Biopsies from patients with CS and GC-O presented increased Ad.Ar/Ma.Ar, along with adipocyte hypertrophy and hyperplasia. In patients with CS, both Ad.Ar/Ma.Ar and Ad.Pf.Ar significantly correlated with serum cortisol levels. Spatial distribution analyses revealed that, in CS, the increase in Ad.Ar/Ma.Ar near to trabecular bone (<100 µm) was mediated by both adipocyte hypertrophy and hyperplasia, while N.Ad.Pf/Ma.Ar further into the marrow (>100 µm) remained unchanged. In contrast, patients with GC-O only presented increased Ad.Ar/Ma.Ar and mean Ad.Pf.Ar>100 µm from trabecular bone surface, highlighting the differential effect of increased endogenous steroid accumulation. Finally, the Ad.Ar/Ma.Ar and Ad.Ar/T.Ar correlated with the canopy coverage above remodeling events. Conclusion: Increased cortisol production in patients with CS induces increased bone marrow adiposity, primarily mediated by adipocyte hypertrophy. This adiposity is particularly evident near trabecular bone surfaces, where hyperplasia also occurs. The differential pattern of adiposity in patients with CS and GC-O highlights that bone marrow adipocytes and their progenitors may respond differently in these two GC-mediated bone diseases.

Spatial Organization of Osteoclastic Coupling Factors and Their Receptors at Human Bone Remodeling Sites.

The strictly regulated bone remodeling process ensures that osteoblastic bone formation is coupled to osteoclastic bone resorption. This coupling is regulated by a panel of coupling factors, including clastokines promoting the recruitment, expansion, and differentiation of osteoprogenitor cells within the eroded cavity. The osteoprogenitor cells on eroded surfaces are called reversal cells. They are intermixed with osteoclasts and become bone-forming osteoblast when reaching a critical density and maturity. Several coupling factors have been proposed in the literature, but their effects and expression pattern vary between studies depending on species and experimental setup. In this study, we investigated the mRNA levels of proposed secreted and membrane-bound coupling factors and their receptors in cortical bone remodeling events within the femur of healthy adolescent human controls using high-sensitivity RNA in situ hybridization. Of the proposed coupling factors, human osteoclasts showed mRNA-presence of LIF, PDGFB, SEMA4D, but no presence of EFNB2, and OSM. On the other hand, the osteoblastic reversal cells proximate to osteoclasts presented with LIFR, PDGFRA and PLXNB1, but not PDGFRB, which are all known receptors of the proposed coupling factors. Although EFNB2 was not present in mature osteoclasts, the mRNA of the ligand-receptor pair EFNB2:EPHB4 were abundant near the central blood vessels within intracortical pores with active remodeling. EPHB4 and SEMA4D were also abundant in mature bone-forming osteoblasts. This study highlights that especially LIF:LIFR, PDGFB:PDGFRA, SEMA4D:PLXNB1 may play a critical role in the osteoclast-osteoblast coupling in human remodeling events, as they are expressed within the critical cells.

Neuronal Sprouting and Reorganization in Bone Tissue Infiltrated by Human Breast Cancer Cells.

Background: Pain is a common complication for patients with metastatic bone disease. Animal models suggest that the pain, in part, is driven by pathological sprouting and reorganization of the nerve fibers innervating the bone. Here, we investigate how these findings translate to humans. Methods: Bone biopsies were collected from healthy volunteers (n = 7) and patients with breast cancer and metastatic bone disease (permissions H-15000679, S-20180057 and S-20110112). Cancer-infiltrated biopsies were from patients without recent anticancer treatment (n = 10), patients with recent anticancer treatment (n = 10), and patients with joint replacement surgery (n = 9). Adjacent bone sections were stained for (1) protein gene product 9.5 and CD34, and (2) cytokeratin 7 and 19. Histomorphometry was used to estimate the area of bone marrow and tumor burden. Nerve profiles were counted, and the nerve profile density calculated. The location of each nerve profile within 25 µm of a vascular structure and/or cancer cells was determined. Results: Cancer-infiltrated bone tissue demonstrated a significantly higher nerve profile density compared to healthy bone tissue. The percentage of nerve profiles found close to vascular structures was significantly lower in cancer-infiltrated bone tissue. No difference was found in the percentage of nerve profiles located close to cancer between the subgroups of cancer-infiltrated bone tissue. Interestingly, no correlation was found between nerve profile density and tumor burden. Conclusions: Together, the increased nerve profile density and the decreased association of nerve profiles to vasculature strongly suggests that neuronal sprouting and reorganization occurs in human cancer-infiltrated bone tissue.

Alendronate prolongs the reversal-resorption phase in human cortical bone remodeling.

Despite their ability to reduce fracture-risk and increase Bone Mineral Density (BMD) in osteoporotic women, bisphosphonates are reported to reduce formation of new bone. Reduced bone formation has been suggested to lead to accumulation of microfractures and contribute to rare side effects in cortical bone such as atypical femur fractures. However, most studies are limited to trabecular bone. In this study, the cortical bone remodeling in human iliac bone specimens of 65 non-treated and 24 alendronate-treated osteoporotic women was investigated using a new histomorphometric classification of intracortical pores. The study showed that only 12.4 ± 11% of the cortical pore area reflected quiescent pores/osteons in alendronate-treated patients versus 8.5 ± 5% in placebo, highlighting that new cortical remodeling events remain to be activated. The percent and size of eroded pores (events in resorption-reversal phase) remained unchanged, but their contribution to total pore area was 1.4-fold higher in alendronate versus placebo treated patients (66 ± 22% vs 48 ± 22%, p < 0.001). On the other hand, the mixed eroded-formative pores (events with mixed resorption-reversal-formation phases) was 2-fold lower in alendronate versus placebo treated patients (19 ± 14% vs 39 ± 23% of total pore area, p < 0.001), and formative pores (event in formation phase) was 2.2-fold lower in alendronate versus placebo treated patients (2.1 ± 2.4% vs 4.6 ± 3.6%, p < 0.01), and their contribution to total pore area was 2.4-fold lower (1.3 ± 2.1% vs 3.1 ± 4.4%, p < 0.05). Importantly, these differences between alendronate and placebo treated patients were significant in patients after 3 years of treatment, not after 2 years of treatment. Collectively, the results support that cortical remodeling events activated during alendronate treatment has a prolonged reversal-resorption phase with a delayed transition to formation, becoming increasingly evident after 3-years of treatment. A potential contributor to atypical femur fractures associated with long-term bisphosphonate treatment.

Vitamin E-doped total hip arthroplasty liners show similar head penetration to highly cross-linked polyethylene at five years: a multi-arm randomized controlled trial.

AIMS: The most frequent indication for revision surgery in total hip arthroplasty (THA) is aseptic loosening. Aseptic loosening is associated with polyethylene liner wear, and wear may be reduced by using vitamin E-doped liners. The primary objective of this study was to compare proximal femoral head penetration into the liner between a) two cross-linked polyethylene (XLPE) liners (vitamin E-doped (vE-PE)) versus standard XLPE liners, and b) two modular femoral head diameters (32 mm and 36 mm). METHODS: Patients scheduled for a THA were randomized to receive a vE-PE or XLPE liner with a 32 mm or 36 mm metal head (four intervention groups in a 2 × 2 factorial design). Head penetration and acetabular component migration were measured using radiostereometric analysis at baseline, three, 12, 24, and 60 months postoperatively. The Harris Hip Score, University of California, Los Angeles (UCLA) Activity Score, EuroQol five-dimension questionnaire (EQ-5D), and 36-Item Short-Form Health Survey questionnaire (SF-36) were assessed at baseline, three, 12, 36, and 60 months. RESULTS: Of 220 screened patients, 127 were included in this study. In all, 116 received the allocated intervention, and 94 had their results analyzed at five years. Head penetration was similar between liner materials and head sizes at five years, vE-PE versus XLPE was -0.084 mm (95% confidence interval (CI) -0.173 to 0.005; p = 0.064), and 32 mm versus 36 mm was -0.020 mm (95% CI -0.110 to 0.071; p = 0.671), respectively. No differences were found in acetabular component migration or in the patient-reported outcome measures. CONCLUSION: No significant difference in head penetration was found at five years between vE-PE and XLPE liners, nor between 32 mm and 36 mm heads. Cite this article: Bone Joint J 2020;102-B(10):1303-1310.

Osteoporosis Treatments Affect Bone Matrix Maturation in a Rat Model of Induced Cortical Remodeling.

To test how osteoporosis drugs affect bone matrix maturation during cortical bone remodeling, 72 pregnant rats were switched from a 0.4% to a 0.01% calcium diet at parturition for a 23-day lactation period. At weaning, eight dams were sacrificed to establish baseline values, while the remaining dams were returned to 0.4% calcium and treated with vehicle (saline), sodium fluoride (NaF), zoledronic acid (ZA), or sclerostin antibody (Scl-Ab) for either 7 or 28 days (eight animals per group per time point). Femora were examined by µCT, dynamic histomorphometry, Fourier transform infrared imaging, and three-point bending of notched specimens. Cortical porosity decreased in all groups from baseline to day 28. Intracortical mineralizing surface (MS/BS) and mineral apposition rate (MAR), as well as the mineral-to-matrix ratio were unaffected by treatment, but intracortical crystallinity was increased in the ZA group at day 10 compared with vehicle. Cortical area increased in all groups over 28 days mainly because of an addition of bone at the endocortical surface. Endocortical MS/BS did not vary among the groups, but endocortical MAR was suppressed in the NaF group at day 2 and elevated in the Scl-Ab group at day 4 compared with vehicle. Endocortical mineral-to-matrix ratio was increased at days 5 and 10 following NaF treatment and endocortical crystallinity was increased at day 5 following ZA treatment compared with vehicle. Fracture toughness did not differ among the groups. Thus, the treatments affected matrix maturation more strongly at the endocortical then intracortical envelope. In this model of induced remodeling, the bone formation phase is synchronized at multiple sites, facilitating study of the effects of drugs or other bone-targeting agents on matrix maturation independent of their effects on the initiation of remodeling. © 2020 The Authors. JBMR Plus published by Wiley Periodicals, Inc. on behalf of American Society for Bone and Mineral Research.

Absence of an osteopetrosis phenotype in IKBKG (NEMO) mutation-positive women: A case-control study.

BACKGROUND: NF-κB essential modulator (NEMO), encoded by IKBKG, is necessary for activation of the ubiquitous transcription factor nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB). Animal studies suggest NEMO is required for NF-κB mediated bone homeostasis, but this has not been thoroughly studied in humans. IKBKG loss-of-function mutation causes incontinentia pigmenti (IP), a rare X-linked disease featuring linear hypopigmentation, alopecia, hypodontia, and immunodeficiency. Single case reports describe osteopetrosis (OPT) in boys carrying hypomorphic IKBKG mutations. METHOD: We studied the bone phenotype in women with IP with evaluation of radiographs of the spine and non-dominant arm and leg; lumbar spine and femoral neck aBMD using DXA; µ-CT and histomorphometry of trans-iliac crest biopsy specimens; bone turnover markers; and cellular phenotype in bone marrow skeletal (stromal) stem cells (BM-MSCs) in a cross-sectional, age-, sex-, and BMI-matched case-control study. X-chromosome inactivation was measured in blood leucocytes and BM-MSCs using a PCR method with methylation of HpaII sites. NF-κB activity was quantitated in BM-MSCs using a luciferase NF-κB reporter assay. RESULTS: Seven Caucasian women with IP (age: 24-67 years and BMI: 20.0-35.2 kg/m2) and IKBKG mutation (del exon 4-10 (n = 4); c.460C>T (n = 3)) were compared to matched controls. The IKBKG mutation carriers had extremely skewed X-inactivation (>90:10%) in blood, but not in BM-MSCs. NF-κB activity was lower in BM-MSCs from IKBKG mutation carriers (n = 5) compared to controls (3094 ±â€¯679 vs. 5422 ±â€¯1038/µg protein, p < 0.01). However, no differences were identified on skeletal radiographics, aBMD, µ-architecture of the iliac crest, or bone turnover markers. The IKBKG mutation carriers had a 1.7-fold greater extent of eroded surfaces relative to osteoid surfaces (p < 0.01), and a 2.0-fold greater proportion of arrested reversal surface relative to active reversal surface (p < 0.01). CONCLUSION: Unlike mutation-positive males, the IKBKG mutation-positive women did not manifest OPT.

Understanding age-induced cortical porosity in women: Is a negative BMU balance in quiescent osteons a major contributor?

Cortical bone is remodeled by intracortical basic multicellular units (BMUs), whose end result can be observed as quiescent osteons in histological sections. These osteons offer a unique opportunity to investigate the BMU balance between the magnitude of bone resorption and subsequent bone formation at the BMU level. Our main objective was to investigate whether the latter parameters change between defined categories of osteons and with age, and to which extend these changes contribute to age-induced cortical porosity. Cortices of iliac bone specimens from 35 women (aged 16-78 years) with a higher porosity with age were investigated. A total of 3084 quiescent osteons reflecting 75% of the intracortical pores were histological examined. The osteons diameter, pore diameter, wall thickness, prevalence and contribution to the porosity were highly variable, but unchanged with age. Next, the osteons were categorized according to whether they reflected the remodeling of existing canals (type 2Q osteons) or the generation of new canals (type 1Q osteons). Type 2Q osteons versus type 1Q osteons: (i) had more frequently a pore diameter > 75 µm (7.4 vs. 1.3%; p < 0.001); (ii) had a larger mean pore diameter (40 ±â€¯10 vs. 25 ±â€¯4 µm; p < 0.001), osteon diameter (120 ±â€¯21 vs. 94 ±â€¯21 µm; p < 0.001) and wall thickness (40 ±â€¯10 vs. 35 ±â€¯9; p < 0.05); (iii) had a larger contribution to the cortical porosity (29 ±â€¯18 vs. 8 ±â€¯8%; p < 0.001); (iv) were more prevalent (44 ±â€¯10 vs. 31 ±â€¯11%; p < 0.001); and (v) were more prevalent with age. Collectively, this study demonstrates that quiescent osteons with age more frequently result from remodeling of existing canals, which in some cases had a more negative BMU balance. Still, the osteons showed no overall age-related change in their pore diameter i.e. BMU balance. In contrast to conventional wisdom, these data show that non-quiescent pores, not pores of quiescent osteons, were the main contributor to a higher cortical porosity.

Intracortical Bone Mechanics Are Related to Pore Morphology and Remodeling in Human Bone.

During aging and in osteoporosis, cortical bone becomes more porous, making it more fragile and susceptible to fractures. The aim of this study was to investigate the intracortical compression- induced strain energy distribution, and determine whether intracortical pores associated with high strain energy density (SED) in the surrounding bone matrix have a different morphology and distribution, as well as different remodeling characteristics than matrix with normal SED. Fibular diaphyseal specimens from 20 patients undergoing a jaw reconstruction (age range 41 to 75 years; 14 men and 6 women) were studied. Bone specimens were µCT-scanned, plastic embedded, and sectioned for histology. Three-dimensional microfinite element models of each specimen were tested in compression, and the SED of the bone immediately surrounding the intracortical pores was calculated within a plane of interest corresponding to the histological sections. The SED of a pore, relative to the distribution of the SED of all pores in each specimen, was used to classify pores as either a high or normal SED pore. Pores with high SED were larger, less circular, and were located closer to the endosteal surface of the cortex than normal SED pores (p < 0.001). Histological analysis of the remodeling events generating the pores revealed that the high SED pores compared with normal SED pores had 13.3-fold higher odds of being an erosive (70%) or formative (7%) pore versus a quiescent pore (p < 0.001), 5.9-fold higher odds of resulting from remodeling upon existing pores (type 2 pore) versus remodeling generating new pores (type 1 pore) (p < 0.001), and 3.2-fold higher odds of being a coalescing type 2 pore versus a noncoalescing type 2 pore (p < 0.001). Overall, the study demonstrates a strong relationship between cortical bone mechanics and pore morphology, distribution, and remodeling characteristics in human fibular bone. © 2018 American Society for Bone and Mineral Research.

Effects of substitute coated with hyaluronic acid or poly-lactic acid on implant fixation: Experimental study in ovariectomized and glucocorticoid-treated sheep.

Investigated in healthy animal models, hyaluronic acid (HyA) and poly-D,L -lactic acid (PDLLA) demonstrate osteoconductive properties when coated onto hydroxyapatite (HA) and ß-tricalcium phosphate (ßTCP) scaffolds. In this study, we examined the efficacy of HA/ßTCP granules coated with HyA or PDLLA on implant fixation when applied as graft materials in 2-mm size defects created in the femur condyles of ovariectomized (OVX) glucocorticoid-treated (GC) sheep. Titanium alloys were inserted into the femur condyles of OVX and GC-treated sheep, and the concentric gaps were filled with either allograft obtained from a healthy donor sheep (control), pure HA/ßTCP, HA/ßTCP-HyA or HA/ßTCP-PDLLA. After 12 weeks, the bone formation adjacent to the implant surface was evaluated by histology and histomorphometry, while the implant fixation was measured by a push-out test. The investigation showed a bone formation in the HA/ßTCP-HyA and HA/ßTCP-PDLLA groups not significantly different from allograft (p  > â€…0.05), whereas the HA/ßTCP group revealed a significantly reduced formation of bone compared with allograft (p  < â€…0.05). Bone-implant contact (BIC) and mechanical properties were similar comparing HA/ßTCP-HyA and HA/ßTCP-PDLLA with allograft (p  > â€…0.05). This study demonstrated that bone substitutes infiltrated with PDLLA and HyA possess osteoconductive properties comparable to allograft when tested in sheep with an OVX and GC-induced bone loss. With no significant difference in implant fixation and bone formation, HyA and PDDLA are indeed considered valuable as new coating materials for composite ceramics when tested in a sheep model - even in bones of a compromised quality.

The efficacy of poly-d,l-lactic acid- and hyaluronic acid-coated bone substitutes on implant fixation in sheep.

BACKGROUND/OBJECTIVE: The present study investigated the efficacy of poly-d,l-lactic acid (PDLLA) and hyaluronic acid (HyA) on implant fixation when coated onto hydroxyapatite/beta-tri-calcium phosphate (HA/ßTCP) granules. METHODS: The effect was assessed in a clinically relevant in vivo gap model in sheep. Thus, four titanium implants combined with either allograft (control), pure HA/ßTCP, HyA infiltrated HA/ßTCP, or PDLLA reinforced HA/ßTCP granules were bilaterally inserted into the trabecular bone of the distal femurs in eight sheep. The insertion created a 2-mm peri-implant gap. After 12 weeks, histomorphometry and push-out test was used for quantification of newly formed bone in the gap, bone-implant contact, and implant fixation. RESULTS: The histomorphometric analysis revealed the presence of newly formed bone in all groups, though substitute groups showed fragments of nonabsorbed substitute material. A significant larger bone volume was found in the allograft group versus the HA/ßTCP-PDLLA group (Zone 1), and in Zone 2 a statistically significantly larger bone volume was found in the allograft compared with the HA/ßTCP group. The mechanical properties and the bone-implant contact revealed no statistically significant differences between the groups. CONCLUSION: This study demonstrates that HA/ßTCP granules coated with PDLLA and HyA have similar bone ingrowth and implant fixation as those with allograft, and with mechanical properties resembling those of allograft in advance, they may be considered as alternative substitute materials for bone formation in sheep.

Bone Formation by Sheep Stem Cells in an Ectopic Mouse Model: Comparison of Adipose and Bone Marrow Derived Cells and Identification of Donor-Derived Bone by Antibody Staining.

Background. Scaffolds for bone tissue engineering (BTE) can be loaded with stem and progenitor cells (SPC) from different sources to improve osteogenesis. SPC can be found in bone marrow, adipose tissue, and other tissues. Little is known about osteogenic potential of adipose-derived culture expanded, adherent cells (A-CEAC). This study compares in vivo osteogenic capacity between A-CEAC and bone marrow derived culture expanded, adherent cells (BM-CEAC). Method. A-CEAC and BM-CEAC were isolated from five female sheep and seeded on hydroxyapatite granules prior to subcutaneous implantation in immunodeficient mice. The doses of cells in the implants were 0.5 × 106, 1.0 × 106, or 1.5 × 106 A-CEAC and 0.5 × 106 BM-CEAC, respectively. After eight weeks, bone volume versus total tissue volume (BV/TV) was quantified using histomorphometry. Origin of new bone was assessed using human vimentin (HVIM) antibody staining. Results. BM-CEAC yielded significantly higher BV/TV than any A-CEAC group, and differences between A-CEAC groups were not statistically significant. HVIM antibody stain was successfully used to identify sheep cells in this model. Conclusion. A-CEAC and BM-CEAC were capable of forming bone, and BM-CEAC yielded significantly higher BV/TV than any A-CEAC group. In vitro treatment to enhance osteogenic capacity of A-CEAC is suggested for further research in ovine bone tissue engineering.

Erratum: Pit- and trench-forming osteoclasts: a distinction that matters.

[This corrects the article DOI: 10.1038/boneres.2015.32.].

Pit- and trench-forming osteoclasts: a distinction that matters.

Osteoclasts (OCs) seeded on bone slices either drill round pits or dig long trenches. Whereas pits correspond to intermittent resorption, trenches correspond to continuous and faster resorption and require a distinct assembly of the resorption apparatus. It is unknown whether the distinction between pits and trenches has any biological relevance. Using OCs prepared from different blood donors, we found that female OCs achieved increased resorption mainly through pit formation, whereas male OCs did so through trench formation. Trench formation went along with high collagenolytic activity and high cathepsin K (CatK) expression, thereby allowing deeper demineralization. A specific CatK inhibitor abrogated the generation of trenches, while still allowing the generation of pits. OCs obtained from bone marrow were more prone to generate trenches than those obtained from blood. Scanning electron microscopy of bone surfaces eroded in vivo showed trenches and pits of similar size as those made by OCs in culture. We conclude that the distinction between trench- and pit-forming OCs is relevant to the differences among OCs from different skeletal sites, different individuals, including gender, and results from differences in collagenolytic power. This indicates a biological relevance and highlights the importance of discriminating between pits and trenches when assessing resorption.

A reversal phase arrest uncoupling the bone formation and resorption contributes to the bone loss in glucocorticoid treated ovariectomised aged sheep.

Large animals as sheep are often used as models for human osteoporosis. Our aim was therefore to determine how glucocorticoid treatment of ovariectomised sheep affects the cancellous bone, determining the cellular events within the bone remodelling process that contributes to their bone loss. Twenty female sheep were assigned for two groups; an untreated control group and an ovariectomised group treated with glucocorticoids (0.6 mg/kg/day, 5 times weekly) for 7 months. At 7 months the glucocorticoid-treated ovariectomised sheep showed a significant change in the bone microstructure revealed by a decreased trabecular bone volume and thickness compared to the control sheep. The treatment led to a temporary elevation of the bone resorption marker CTX (c-terminal collagen telopeptide), while the bone formation marker osteocalcin remained suppressed all 7 months. Histomorphometrically, the treated sheep had a complete absence of osteoid surfaces, and a 5-fold increase in the extent of eroded/reversal surfaces after 7 months. Most of these reversal surfaces were actually arrested reversal surfaces, defined as reversal surfaces without the presence of neighbouring osteoid surfaces or osteoclasts, which is classically observed next to active reversal surfaces. As in humans, these arrested reversal surfaces had compared to active reversal surfaces a reduced canopy coverage, a significantly decreased cell density, and a decreased immunoreactivity for the osteoblastic markers osterix, runx2 and smooth muscle actin in the mononuclear reversal cells colonising the surfaces. In conclusion, glucocorticoid treatment of ovariectomised sheep induced a significant bone loss, caused by an arrest of the reversal phase, resulting in an uncoupling of the bone formation and resorption during the reversal phase, as recently demonstrated in postmenopausal women with glucocorticoid-induced osteoporosis. This supports the relevance of the sheep model to the pathophysiology of glucocorticoid-induced osteoporosis in postmenopausal women, making it a relevant preclinical model for orthopaedic implant and biomaterial research.

Efficacy of a small cell-binding peptide coated hydroxyapatite substitute on bone formation and implant fixation in sheep.

Cylindrical critical size defects were created at the distal femoral condyles bilaterally of eight female adult sheep. Titanium implants with 2-mm concentric gaps were inserted and the gaps were filled with one of the four materials: allograft; a synthetic 15-amino acid cell-binding peptide coated hydroxyapatite (ABM/P-15); hydroxyapatite + ßtricalciumphosphate+ Poly-Lactic-Acid (HA/ßTCP-PDLLA); or ABM/P-15+HA/ßTCP-PDLLA. After nine weeks, bone-implant blocks were harvested and sectioned for micro-CT scanning, push-out test, and histomorphometry. Significant bone formation and implant fixation could be observed in all four groups. Interestingly, the microarchitecture of the ABM/P-15 group was significantly different from the control group. Tissue volume fraction and thickness were significantly greater in the ABM/P-15 group than in the allograft group. Bone formation and bone ingrowth to porous titanium implant were not significantly different among the four groups. The ABM/P-15 group had similar shear mechanical properties on implant fixation as the allograft group. Adding HA/ßTCP-PDLLA to ABM/P-15 did not significantly change these parameters. This study revealed that ABM/P-15 had significantly bone formation in concentric gap, and its enhancements on bone formation and implant fixation were at least as good as allograft. It is suggested that ABM/P-15 might be a good alternative biomaterial for bone implant fixation in this well-validated critical-size defect gap model in sheep. Nevertheless, future clinical researches should focus on prospective, randomized, controlled trials in order to fully elucidate whether ABM/P-15 could be a feasible candidate for bone substitute material in orthopedic practices.