Non-osseous Sites Research Articles

Bone without borders - Monetite-based calcium phosphate guides bone formation beyond the skeletal envelope.

Calcium phosphates (CaP) represent an important class of osteoconductive and osteoinductive biomaterials. As proof-of-concept, we show how a multi-component CaP formulation (monetite, beta-tricalcium phosphate, and calcium pyrophosphate) guides osteogenesis beyond the physiological envelope. In a sheep model, hollow dome-shaped constructs were placed directly over the occipital bone. At 12 months, large amounts of bone (∼75%) occupy the hollow space with strong evidence of ongoing remodelling. Features of both compact bone (osteonal/osteon-like arrangements) and spongy bone (trabeculae separated by marrow cavities) reveal insights into function/need-driven microstructural adaptation. Pores within the CaP also contain both woven bone and vascularised lamellar bone. Osteoclasts actively contribute to CaP degradation/removal. Of the constituent phases, only calcium pyrophosphate persists within osseous (cutting cones) and non-osseous (macrophages) sites. From a translational perspective, this multi-component CaP opens up exciting new avenues for osteotomy-free and minimally-invasive repair of large bone defects and augmentation of the dental alveolar ridge.

Coupling between macrophage phenotype, angiogenesis and bone formation by calcium phosphates

The ability of calcium phosphate (CaP) materials to induce bone formation varies with their physicochemical properties, with surface topography as one of the most crucial triggers. In view of the natural wound healing processes (e.g., inflammation, angiogenesis, tissue formation and remodeling) initiated after surgical implantation, we here comparatively investigated the biological cascades occurring upon ectopic implantation of a tricalcium phosphate with submicron surface topography (TCP-S, osteoinductive) and a tricalcium phosphate with micron-scale topography (TCP-B, non-osteoinductive). In vitro, TCP-S facilitated M2 polarization of macrophages derived from a human leukemic cell line (THP-1) as shown by the enhanced secretion of TGF-β and CCL18. Interestingly, the conditioned media of polarized M2 macrophages on TCP-S enhanced tube formation by human umbilical vein endothelial cells (HUVECs), while had no influence on the osteogenic differentiation of human bone marrow stromal cells (HBMSCs). Following an intramuscular implantation in canines, TCP-S locally increased typical M2 macrophage markers (e.g., IL-10) at week 1 to 3 and enhanced blood vessel formation after week 3 as compared to TCP-B. Bone formation was observed histologically in TCP-S 6 weeks after implantation, and bone formation was inhibited when an angiogenesis inhibitor (KRN633) was loaded onto TCP-S. No bone formation was observed for TCP-B. The data presented herein suggest strong links between macrophage polarization, angiogenesis and CaP-induced bone formation. Statement of significanceThe ability of calcium phosphate (CaP) materials to induce bone formation varies with their physicochemical properties, and the key physicochemical properties relevant to CaP-induced bone formation have been outlined in the last two decades. However, the biological mechanism underlying this material-driven osteoinduction remains largely unknown. This manuscript presented demonstrates strong links between surface topography, macrophage polarization, angiogenesis and bone formation in CaP materials implanted in non-osseous sites. The finding may provide new clues for further exploring the possible mechanism underlying osteoinduction by CaP materials.

Exercise Promotes the Osteoinduction of HA/β-TCP Biomaterials via the Wnt Signaling Pathway

To investigate the osteoinductive mechanism triggered by hydroxyapatite/β-tricalcium phosphate (HA/β-TCP) biomaterials in mice which keep exercising. Methods: The HA/β-TCP biomaterials were implanted in the muscle of bilateral thighs (non-osseous sites) of eighty Balb/C mice. All animals were then randomly divided into 4 groups (n = 20). In group 1 (negative control group), the mice were fed routinely. In group 2 (running group), all mice were put on a treadmill which was set to a 60-degree incline. The mice ran 20 min thrice each day. A 5-minute break was included in the routine from day three onwards. In group 3 (weight-bearing group), all mice underwent weight-bearing running. The mice in this group performed the same routine as group 2 while carrying 5 g rubber weights. In group 4 (positive control group), dexamethasone was injected in the implanted sites of the biomaterials from the day of the operation. All mice were injected once per week and received a total of 8 injections. One and eight weeks after surgery, the blood serum was collected to detect inflammatory and immunological factors by ELISA. In addition to this, biomaterial specimens were obtained to observe inflammatory and osteogenic levels via histological staining and to facilitate analysis of the osteogenic mechanism by Western Blot. Results: The inflammation indexes caused by surgery were alleviated through running or weight-bearing running: The tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) levels were significantly reduced in groups 2 and 3 at week 8. Exercise also enhanced the secretion of interferon-γ (IFN-γ) in mice; this can strengthen their immunity. The new bone tissues were observed in all groups; however, the area percentage of new bone tissues and the number of osteoblasts were highest in the weight-bearing group. Furthermore, the key proteins of wingless/integrated (Wnt) signaling pathway, Wnt1, Wnt3a, and β-catenin, were up-regulated during osteoinduction. This up-regulation activated runt-related transcription factor-2 (Runx2), increased the expression of osteopontin (OPN) and osteocalcin (OCN). Conclusion: Weight-bearing exercise can promote the bone and bone marrow formation through the Wnt signaling pathway: Observations documented here suggest that the proper exercise is beneficial to the recovery of bone damage.

Alpha-Particle Therapy for Multifocal Osteosarcoma: A Hypothesis.

Osteosarcoma (OST) is the most common bone tumor in children and adolescents with a second peak of incidence in elderly adults usually diagnosed as secondary tumors in Paget's disease or irradiated bone. Subjects with metastatic disease or whose disease relapses after the initial therapy have a poor prognosis. Moreover, multifocal OST contains tumor-initiating cells that are resistant to chemotherapy. The use of aggressive therapies in an attempt to eradicate these cells can have long-term negative consequences in these vulnerable patient populations. 227Th-labeled molecular probes based on ligands to OST-associated receptors such as IGF-1R (insulin-like growth factor receptor 1), HER2 (human epidermal growth factor receptor 2), and PSMA (prostate-specific membrane antigen) are expected to detect and treat osseous and nonosseous sites of multifocal OST. Published reports indicate that 227Th has limited myelotoxicity, can be stably chelated to its carriers and, as it decays at targeted sites, 227Th produces 223Ra that is subsequently incorporated into the areas of increased osteoblastic activity, that is, osseous metastatic lesions. Linear energy transfer of α particles emitted by 227Th and its daughter 223Ra is within the range of the optimum relative biological effectiveness. The radiotoxicity of α particles is virtually independent of the phase in the cell cycle, oxygenation, and the dose rate. For these reasons, even resistant OST cells remain susceptible to killing by high-energy α particles, which can also kill adjacent quiescent OST cells or cells with low expression of targeted receptors. Systemic side effects are minimized by the limited range of these intense radiations. Quantitative single-photon emission computed tomography of 227Th and 223Ra is feasible. Additionally, the availability of radionuclide pairs, for example, 89Zr for positron emission tomography and 227Th for therapy, establish a strong basis for the theranostic use of 227Th in the individualized treatment of multifocal OST.

Coupling between Macrophage Phenotype, Angiogenesis and Bone Formation by Calcium Phosphates

The ability of calcium phosphate (CaP) materials to induce bone formation varies with their physicochemical properties, with surface topography as one of the most crucial triggers. In view of the natural wound healing processes initiated after surgical implantation, involving inflammation, angiogenesis, tissue formation and remodeling, we here comparatively investigated the biological cascades occurring upon ectopic implantation of a tricalcium phosphate with submicron surface topography (TCP-S, osteoinductive) and a tricalcium phosphate with micron-scale topography (TCP-B, non-osteoinductive). In vitro, TCP-S facilitated M2 polarization of macrophages derived from a human leukemic cell line (THP-1) as shown by the enhanced secretion of TGF-β and CCL18. Interestingly, the conditioned media of polarized M2 macrophages on TCP-S enhanced tube formation by human umbilical vein endothelial cells (HUVECs), while had no influence on the osteogenic differentiation of human bone marrow stromal cells (HBMSCs). Following implantation in a canine intramuscular defect, TCP-S locally increased typical M2 macrophage markers (e.g. IL-10) at week 1 and enhanced blood vessel formation after week 3 compared to TCP-B. Bone formation was observed histologically for TCP-S 6 weeks after implantation, and bone formation was reduced when an angiogenesis inhibitor (KRN633) was added to TCP-S. No bone formation was observed for TCP-B. The data here presented demonstrates strong links between surface topography, macrophage polarization, angiogenesis and bone formation in CaP materials implanted in non-osseous sites.

Correlations between macrophage polarization and osteoinduction of porous calcium phosphate ceramics

The host immune response is critical for in situ osteogenesis, but correlations between local inflammatory reactions and biomaterial osteoinduction are still poorly understood. This study used a murine intramuscular implantation model to demonstrate that calcium phosphate ceramics with different phase compositions exhibited divergent osteoinductivities. The osteoinductive potential of each ceramic was closely associated with the immunomodulatory capacity of the material, and especially with the regulation of macrophage polarization and functional status. Biphasic calcium phosphate (BCP) ceramics with superior osteoinductive potential enhanced the fraction of CD206+ M2 macrophages, up-regulated expression of M2 phenotypic markers in vitro, and increased the ARG+ M2 population in vivo. This suggested that BCP ceramics could ameliorate long-term inflammation and build a pro-osteogenic microenvironment. However, β-tricalcium phosphate (β-TCP) ceramics with no obvious osteoinductivity increased the fraction of CCR7+ M1 macrophages, promoted the secretion of M1 phenotypic markers in vitro, and maintained a high proportion of iNOS+ M1 macrophages in vivo. It indicated that β-TCP ceramics could exacerbate inflammation and inhibit ectopic bone formation. Hydroxyapatite ceramics with an intermediate osteoinductivity exhibited a moderate amount of both M1 and M2 macrophages. These findings highlight the critical role of macrophage polarization in biomaterial-dependent osteoinduction, which not only deepens our understanding of osteoinductive mechanisms but also provides a strategy to design bone substitutes by endowing materials with the proper immunomodulatory abilities to achieve the desired clinic performance. Statement of SignificanceCalcium phosphate (CaP) ceramics with osteoinductive capacities are able to induce ectopic bone formation in non-osseous sites. However, its underlying mechanism is largely unknown. Previous studies have demonstrated an indispensable role of macrophages in osteogenesis, inspiring us that local inflammatory reaction may affect material-dependent osteoinduction. This study indicated that CaP ceramics with different phase composition could present divergent osteoinductive capacities through modulating polarization and functional status of macrophages, as biphasic calcium phosphate with potent osteoinductivity ameliorated long-term inflammation and induced a healing-associated M2 phenotype to initiate bone formation. These findings not only get an insight into the mechanism of CaP-involved osteoinduction, but also help the design of tissue-inducing implants by endowing biomaterials with proper immunomodulatory ability.

Effects of hydroxyapatite surface nano/micro-structure on osteoclast formation and activity.

The surface structure of calcium phosphate (CaP) ceramic plays an important role in its osteoinductivity; however, little is known about its effects on osteoclastogenesis. In this study, an intramuscular implantation model suggested a potential relationship between hydroxyapatite (HA)-induced bone formation and osteoclast appearance in the non-osseous site, which might be modulated by scaffold surface structure. Then, three dense HA discs with different grain sizes from biomimetic nanoscale (∼100 nm) to submicron scale (∼500 nm) were fabricated via distinct sintering procedures, and their impacts on osteoclastic differentiation of RAW 264.7 macrophages under RANKL stimulation were further investigated. Our results showed that compared with the ones in the submicron-scale dimension, nano-structured HA discs markedly impaired osteoclastic formation and function, as evidenced by inhibited cell fusion, reduced osteoclast size, less-defined actin ring, increased osteoclast apoptosis, suppressed expression of osteoclast specific genes and proteins, decreased TRAP-positive cells, and hampered resorption activity. This demonstrated that the surface structure of CaP ceramics has a great influence on osteoclastogenesis, which might be further related to its osteoinductive capacity. These findings might not only help us gain insight into biomolecular events during CaP-involved osteoinduction, but also offer a principle for designing orthopaedic implants with an ability of regulating both osteogenesis and osteoclastogenesis to achieve the desired performance.

Evaluation of osteoinductive calcium phosphate ceramics repairing alveolar cleft defects in dog model.

Alveolar cleft repair is an important step in the sequence of treatments for cleft lip and palate. Intrinsically osteoinductive materials have been the subject of research interest. The aim of this study was to explore the use of osteoinductive biphasic calcium phosphate (BCP) ceramics to repair alveolar cleft defects in dogs. We prepared two kinds of BCP ceramic with different physical characteristics: osteoinductive BCP (OBCP) and non-osteoinductive BCP (NBCP). Bilateral alveolar cleft models were surgically established in dogs. On one side, OBCP was implanted in the defect; on the opposite side NBCP was implanted as a control. The materials were also implanted in the femoral muscles to test their properties at non-osseous sites. The osteogenic ability of materials was evaluated with imaging, spiral CT, histology and fluorescent dye tests. At the muscular implantation sites, new bone formed in all of the OBCP samples, but none in the NBCP samples. Imaging and spiral CT revealed good appearance and continuity of the alveolar cleft postoeration, with normal eruption of the bilateral permanent teeth in the groups. Histological and fluorescent dye testing revealed new bone formation in both groups in situ. However, earlier osteogenesis initiation and bone remodeling were superior with OBCP. Osteogenic process in the intramuscular samples with OBCP was similar to that seen in situ. Our findings indicated that osteoinductive biphasic calcium phosphate ceramics (OBCP) have superior characteristics in alveolar cleft repair compared with non-osteoinductive calcium phosphate ceramics (NBCP).

Pattern and Distribution of Distant Metastases in Anaplastic Prostate Carcinoma: A Single-Institute Experience With 101 Patients.

The aim of this study was to evaluate the sites and frequencies of distant metastases in patients with anaplastic prostate carcinoma and to correlate those findings with prostate-specific antigen (PSA) levels. Patients with anaplastic prostate carcinoma (n = 101) underwent CT and bone scans before platinum-based chemotherapy. CT findings were retrospectively reviewed to identify the sites of metastases. CT findings were correlated with baseline PSA levels. The Wilcoxon rank sum test was used to correlate PSA levels between patients with metastases at osseous and nonosseous sites. The Wilcoxon rank sum test was also used to correlate the type of bone metastases (blastic vs lytic) and the PSA levels. Eighty-three of 101 patients (82%) had osseous metastases. PSA levels were significantly higher in patients with bone metastases than in patients without osseous metastases. However, 23 of the 83 patients (28%) with bone metastases had PSA levels in the normal range (i.e., < 4 ng/mL). The type of bone metastases (blastic vs lytic) did not show any statistically significant correlation to the PSA levels. Overall, 63 of 101 patients (62%) had nonosseous distant metastases at one or more sites, including the liver (n = 34), lung (n = 24), mediastinum (n = 31), pleura (n = 7), brain (n = 9), adrenal glands (n = 6), peritoneum (n = 4), and spleen (n = 1). PSA levels were not significantly elevated in patients with nonosseous distant metastases. Twenty-six of the 63 patients (41%) with nonosseous metastases had PSA levels in the normal range (< 4 ng/mL). Patients with the anaplastic clinical variant of prostate cancer have a high frequency of typical and atypical sites of metastases. Common sites of nonosseous distant metastases include the liver, lung, mediastinum, pleura, brain, and adrenal glands. PSA levels are unreliable and may be disproportionately low, despite the presence of multifocal large-volume metastases. CT of the chest, abdomen, and pelvis should be considered in routine staging and follow-up of patients with anaplastic prostate carcinoma regardless of their PSA levels.

Calcium phosphate scaffolds with different macro-pores constructed vascularized natural bone-like grafts at non-osseous sites in vivo for bone defect repairing

Calcium phosphate scaffolds with different macro-pores constructed vascularized natural bone-like grafts at non-osseous sites in vivo for bone defect repairing

Feasibility analysis of pathology and genetic yield from a prospective trial of tissue biopsies in metastatic castrate-resistant prostate cancer (mCRPC).

249 Background: Evaluating tumor specific pathologic and genetic profiles in mCRPC stage is difficult due to the limited availability of mCRPC tissue. We describe findings from a prospective cohort study performed to obtain concomitant histopathology and genetic information in mCRPC. Methods: Patients with mCRPC initiating abiraterone acetate therapy underwent 2 serial (3 months apart) metastatic (met) site needle core biopsies (NCB1/2). Bone lesions were biopsied using 11-13G core needles and 18G core biopsy devices were used for non-osseous masses. Up to 4 cores were obtained at each biopsy with the 1st core (S1) sent for DNA sequencing, the 2nd (S2) for RNA sequencing, and the 3rd/4th (X3, X4) submitted for xenograft implantation. From each, 1-2 mm segments were separated and formalin-fixed for histopathologic examination (HPE). Results: A total of 54 patients, enrolled between June 2013 and July 2014, underwent 94 NCB (54 NCB1, 40 NCB2), rendering a total of 259 samples (94 S1; 75 S2; 59 X3 and 31 X4) of which 85 (32%) were positive (pos) for tumor by HPE. Positivity for tumor in S1, S2, X3, and X4 cores was 42%, 33%, 22%, and 19%, respectively. At least one core pos for met tumor was observed in 62% NCB1 and 52% NCB2 (overall 52/94; 56%). Met sites biopsied include bone (71), lymph nodes (18), liver (3), penile (1) and pelvic (1) soft tissues. HPE revealed met adenocarcinoma in 44/52, and poorly differentiated carcinoma in 8/52. Gleason grade applied to the met ranged from 3+4 to 5+5. 45/85 pos samples (53%) had &gt; 50% tumor cellularity. Pos NCB in bone lesions was observed in 23/71 (32%), compared to 17/23 (73%) of non-bone sites (p=.0004). 85 S1 samples yielded DNA material, of which 66 (77%) had ≥10% tumoral DNA. 24 cases with negative HPE had ≥10% tumoral DNA; 7 cases with pos HPE had &lt;10 tumoral DNA. In all, 76/94 (81%) NCB yielded tumor material either by HPE or DNA analysis. Conclusions: NCB of mCRPC is feasible and provides adequate tissue for pursuing HPE and sequencing studies. HPE of extracted material correlates with DNA sequencing data and provides complementary information on tumor features. Bone lesions yield significantly less tumoral material than non-osseous sites.

Influence of fluoride in poly(d,l-lactide)/apatite composites on bone formation.

The influence of fluoride in poly(d,l-lactide)/apatite composites on ectopic bone formation was evaluated in sheep. Nano-apatite powders with different replacement levels of OH groups by fluoride (F) (0% (F0), 50% (F50), 100% (F100), and excessive (F200)) were co-extruded with poly (d,l-lactide) at a weight ratio of 1:1. Fluoride release from the composites (CF0, CF50, CF100, and CF200) was evaluated in vitro and bone formation was assessed after intramuscular implantation in sheep. After 24 weeks in simulated physiological solution, CF0 and CF50 showed negligible fluoride release, whereas it was considerable from the CF100 and CF200 composites. Histology showed that the incidence of de novo bone formation decreased in implants with increasing fluoride content indicating a negative influence of fluoride on ectopic bone formation. Furthermore, a significant decrease in resorption of the high fluoride-content composites and a reduction in the number of multinucleated giant cells were seen. These results show that instead of promoting, the presence of fluoride in poly(d,l-lactide)/apatite composites seemed to suppresses their resorption and osteoinductive potential in non-osseous sites.

Is prostate cancer changing?: evolving patterns of metastatic castration-resistant prostate cancer.

Metastatic castration-resistant prostate cancer (mCRPC) most commonly metastasizes to the bone, and less commonly to nonosseous sites (eg, lymph nodes, liver, lung). With new therapies extending survival in mCRPC, it was hypothesized that the pattern of metastases is changing over time. The pattern of metastatic disease was evaluated in men with mCRPC, as reported in baseline characteristics of prospective clinical trials over 2 decades. This study identified all phase 2 and 3 therapeutic studies in men with mCRPC in PubMed and American Society of Clinical Oncology abstracts from 1990 to 2012. Studies were excluded if they did not report demographic data and sites of metastasis, or excluded patients with a specific site of metastatic disease (except brain). For each type of metastasis, weighted least squares linear regression models were used to evaluate temporal trends. A total of 290 eligible studies (270 phase 2 studies and 20 phase 3 studies) involving 19,110 patients were identified. Between 1990 and 2012, the rate of nonosseous metastasis increased significantly at 1.6% per year (P < .0001), whereas the rate of osseous metastasis decreased at 0.5% per year (P < .0001). The rate of lymph node metastasis increased at 1.4% per year (P < .0001), but the rate of liver and lung metastasis remained relatively stable. A notable change was found in the pattern of metastasis in patients with mCRPC. Because these evolving patterns may have important implications in treatment selection and prognosis, it is crucial that future clinical trials of patients with mCRPC define patients with a uniform reporting of nonosseous metastasis.

English

Plasmacytoma is a neoplastic proliferation of plasma cells that may manifest as multiple myeloma, primary amyloidosis, or monoclonal gammapathy of unknown significance. Plasmacytoma may be primary or secondary to disseminated multiple myeloma and may arise from osseous (medullary) or non-osseous (extramedullary) sites. Primary extramedullary plasmacytoma can be solitary or multiple1. The International Myeloma Working Group in 2003 recognized a separate classification of plasmacytomas that occur as multiple sites of disease in soft tissue, bone, or both soft tissue and bone as multiple solitary plasmacytoma. Primary extramedullary plasmacytoma is rare, accounting for only 4% of all plasma cell tumors. Here we report a rare case of extra medullary ethmoidal sinus plasmacytoma with intracranial extension with its clinical, radiological and histological features.

A novel porous bioceramics scaffold by accumulating hydroxyapatite spherulites for large bone tissue engineering in vivo. II. Construct large volume of bone grafts

In vivo engineering of bone autografts using bioceramic scaffolds with appropriate porous structures is a potential approach to prepare autologous bone grafts for the repair of critical-sized bone defects. This study investigated the evolutionary process of osteogenesis, angiogenesis, and compressive strength of bioceramic scaffolds implanted in two non-osseous sites of dogs: the abdominal cavity and the dorsal muscle. Hydroxyapatite (HA) sphere-accumulated scaffolds with controlled porous structures were prepared and placed in the two sites for up to 6 months. Analyses of retrieved scaffolds found that osteogenesis and angiogenesis were faster in scaffolds implanted in dorsal muscles compared with those placed in abdominal cavities. The abdominal cavity, however, can accommodate larger bone grafts with designed shape. Analyses of scaffolds implanted in abdominal cavities [an environment of a low mesenchymal stem cell (MSC) density] further demonstrated that angiogenesis play critical roles during osteogenesis in the scaffolds, presumably by supplying progenitor cells and/or MSCs as seed cells. This study also examined the relationship between the volume of bone grafts and the physiological environment of in vivo bioreactor. These results provide basic information for the selection of appropriate implanting sites and culture time required to engineer autologous bone grafts for the clinical bone defect repair. Based on these positive results, a pilot study has applied the grafts constructed in canine abdominal cavity to repair segmental bone defect in load-bearing sites (limbs).

Poor osteoinductive potential of subcutaneous bone cement-induced membranes for tissue engineered bone

Background: Large segmental bone defects remain a challenge for reconstructive surgeons. A two-stage repair strategy may offer a potential solution. Here, we sought to evaluate the osteoinductive potential of bone cement-induced membranes in an ectopic site. Methods: First, bone cements were inserted into the subcutaneous tissues of 16 rabbits to induce membrane formation. After 2, 4, 6 and 8 weeks, the induced membranes were harvested to assess their vascularization and osteoinductive potential. Next, bone cements were subcutaneously inserted into 12 rabbits for 4 weeks. These bone cements were then harvested from the newly formed membranes and replaced with granular porous β-TCP, with or without bone mesenchymal stem cells. New bone formation was then evaluated after 3, 6 and 9 weeks. Results: The highest level of blood vessel formation and bone morphogenetic protein-2 expression in the membranes were found at 4 weeks (p < 0.05). In addition, vascular endothelial growth factor concentration was highest after 2 weeks (p < 0.001), persisting until 8 weeks. However, the results showed little ectopic bone formation at these time points. Conclusion: While bone cement-induced membranes appear to provide a suitable environment for bone formation, they fail to drive osteoinduction in non-osseous sites for the purposes of bone tissue engineering.

The homing of bone marrow MSCs to non-osseous sites for ectopic bone formation induced by osteoinductive calcium phosphate

Osteoinductive biomaterials are promising for bone repair. There is no direct proof that bone marrow mesenchymal stem cells (BMSCs) home to non-osseous sites and participate in ectopic bone formation induced by osteoinductive bioceramics. The objective of this study was to use a sex-mismatched beagle dog model to investigate BMSC homing via blood circulation to participate in ectopic bone formation via osteoinductive biomaterial. BMSCs of male dogs were injected into female femoral marrow cavity. The survival and stable chimerism of donor BMSCs in recipients were confirmed with polymerase chain reaction (PCR) and fluorescence in situ hybridization (FISH). Biphasic calcium phosphate (BCP) granules were implanted in dorsal muscles of female dogs. Y chromosomes were detected in samples harvested from female dogs which had received male BMSCs. At 4 weeks, cells with Y-chromosomes were distributed in the new bone matrix throughout the BCP granule implant. At 6 weeks, cells with Y chromosomes were present in newly mineralized woven bone. TRAP positive osteoclast-like cells were observed in 4-week implants, and the number of such cells decreased from 4 to 6 weeks. These results show that osteoprogenitors were recruited from bone marrow and homed to ectopic site to serve as a cell source for calcium phosphate-induced bone formation. In conclusion, BMSCs were demonstrated to migrate from bone marrow through blood circulation to non-osseous bioceramic implant site to contribute to ectopic bone formation in a canine model. BCP induced new bone in muscles without growth factor delivery, showing excellent osteoinductivity that could be useful for bone tissue engineering.

Bone conduction activation through soft tissues following complete immobilization of the ossicular chain, stapes footplate and round window

Classically it has been thought that bone conduction activation at the mastoid leads to relative motion between the stapes footplate and the oval window due to inertial and to compression (distortion) mechanisms. However, several recent clinical findings and experimental manipulations may point to additional mechanisms. These manipulations were extended in the present study. In ten fat sand rats, following obliteration of one ear, auditory nerve brainstem evoked response (ABR) thresholds were recorded in response to broad band click stimuli, either air conducted (AC) through insert earphones or bone conducted (BC) delivered directly to the exposed skull bone. Following this, the entire ossicular chain, stapes footplate and round window were completely immobilized with super glue, leading to a mean AC threshold elevation of 44 dB, but to a mean BC threshold change (elevation) of only 3.5 dB. In this state of complete immobilization, the bone vibrator was applied to a pool of saline in the surgical area and ABR was elicited with a mean threshold which was not significantly different from that of the BC threshold. When the bone vibrator was then applied to the eye without touching the bone at the orbit, the resulting ABR threshold was about 20 dB greater than the BC threshold. In conclusion, BC stimulation can activate the cochlea without two mobile windows. Furthermore, the cochlea can be activated by a fluid pathway and by application of a bone vibrator to non-osseous sites (soft tissue conduction).

Osteoinduction of porous Ti implants with a channel structure fabricated by selective laser melting

Many studies have shown that certain biomaterials with specific porous structures can induce bone formation in non-osseous sites without the need for osteoinductive biomolecules, however, the mechanisms responsible for this phenomenon (intrinsic osteoinduction of biomaterials) remain unclear. In particular, to our knowledge the type of pore structure suitable for osteoinduction has not been reported in detail. In the present study we investigated the effects of interconnective pore size on osteoinductivity and the bone formation processes during osteoinduction. Selective laser melting was employed to fabricate porous Ti implants (diameter 3.3 mm, length 15 mm) with a channel structure comprising four longitudinal square channels, representing pores, of different diagonal widths, 500, 600, 900, and 1200 μm (termed p500, p600, p900, and p1200, respectively). These were then subjected to chemical and heat treatments to induce bioactivity. Significant osteoinduction was observed in p500 and p600, with the highest observed osteoinduction occurring at 5 mm from the end of the implants. A distance of 5 mm probably provides a favorable balance between blood circulation and fluid movement. Thus, the simple architecture of the implants allowed effective investigation of the influence of the interconnective pore size on osteoinduction, as well as the relationship between bone quantity and its location for different pore sizes.

Tissue response of an osteoinductive bioceramic in bone defect rabbit model

HA/TCP and HA rods (ϕ5 mm×10 mm) were made for implantation in New Zealand white rabbit with different condition. Sixty three rabbit were divided into three groups: group 1 (n=18), group 2 (n=27) and group 3 (n=18). In group 1, 10 mm radius was defected, and one HA/TCP rod was implanted in the muscle a distant away from the bone defect area. In group 2, also, 10 mm radius was defected, one HA rod was implanted in the muscle a distant away from the bone defect area. In group 3, two HA/TCP rods were implanted in the dorsal muscle of the rabbit with bone intact. Histological observation showed that in group 1, some new bone was found only two months after implantation (n=2), and obvious immature woven bone could be observed in these bioceramics from the 3rd month on. However, in group 3, bone began to be found 6 months after implantation (n=2). In group 2, we could not find any bone tissue up to 9 month’s observation. These results suggest that, first, the bone defect model could significantly accelerate bone formation at non-osseous sites in rabbits; second,. HA/TCP bioceramics were confirmed with osteoinductive property while HA bioceramics without osteoinductive property nearly. Thus, bone defect might be a good animal model for further researches for osteoinductive bioceramics.