Rabbit Cranial Defect Model Research Articles

Injectable dual drug-loaded thermosensitive liposome-hydrogel composite scaffold for vascularised and innervated bone regeneration

Adequate blood supply and thorough innervation are essential to the survival of tissue-engineered bones. Though great progress has been created in the application of bone tissue engineering technology to bone defect repair, many challenges remain, such as insufficient vascularisation and deficient innervation in newly regenerated bone. In the present study, we addressed these challenges by manipulating the bone regeneration microenvironment in terms of vascularisation and innervation. We used a novel injectable thermosensitive liposome-hydrogel composite scaffold as a sustained-release carrier for basic fibroblast growth factor (bFGF, which promotes angiogenesis and neurogenic differentiation) and dexamethasone (Dex, which promotes osteogenic differentiation). In vitro biological assessment demonstrated that the composite scaffold had sufficient cell compatibility; it enhanced the capacity for angiogenesis in human umbilical vein endothelial cells, and the capacity for neurogenic/osteogenic differentiation in human bone marrow mesenchymal stem cells. Moreover, the introduction of bFGF/Dex liposome-hydrogel composite scaffold to bone defect sites significantly improved vascularisation and innervated bone regeneration properties in a rabbit cranial defect model. Based on our findings, the regeneration of sufficiently vascularised and innervated bone tissue through a sustained-release scaffold with excellent injectability and body temperature sensitivity represents a promising tactic towards bone defect repair.

A facile Immunoregulatory Constructional Design by Proanthocyanidin Optimizing Directional Chitosan Microchannel.

Improving the interconnected structure and bioregulatory function of natural chitosan is beneficial for optimizing its performance in bone regeneration. Here, a facile immunoregulatory constructional design is proposed for developing instructive chitosan by directional freezing and alkaline salting out. The molecular dynamics simulation confirmed the assembly kinetics and structural features of various polyphenols and chitosan molecules. Along with the in vitro anti-inflammatory, antioxidative, promoting bone mesenchymal stem cell (BMSC) adhesion and proliferation performance, proanthocyanidin optimizing chitosan (ChiO) scaffold presented an optimal immunoregulatory structure with the directional microchannel. Transcriptome analysis in vitro further revealed the cytoskeleton- and immune-regulation effect of ChiO are the key mechanism of action on BMSC. The rabbit cranial defect model (Φ = 10mm) after 12 weeks of implantation confirmed the significantly enhanced bone reconstitution. This facile immunoregulatory directional microchannel design provides effective guidance for developing inducible chitosan scaffolds.

Polysaccharide-Based Composite Hydrogel with Hierarchical Microstructure for Enhanced Vascularization and Skull Regeneration.

Critical-size skull defects caused by trauma, infection, and tumor resection raise great demands for efficient bone substitutes. Herein, a hybrid cross-linked hierarchical microporous hydrogel scaffold (PHCLS) was successfully assembled by a multistep procedure, which involved (i) the preparation of poly(lactic-co-glycolic)/nanohydroxyapatite (PLGA-HAP) porous microspheres, (ii) embedding the spheres in a solution of dopamine-modified hyaluronic acid and collagen I (Col I) and cross-linking via dopamine polyphenols binding to (i) Col I amino groups (via Michael addition) and (ii) PLGA-HAP (via calcium ion chelation). The introduction of PLGA-HAP not only improved the diversity of pore size and pore communication inside the matrix but also greatly enhanced the compressive strength (5.24-fold, 77.5 kPa) and degradation properties to construct a more stable mechanical structure. In particular, the PHCLS (200 mg, nHAP) promoted the proliferation, infiltration, and angiogenic differentiation of bone marrow mesenchymal stem cells in vitro, as well as significant ectopic angiogenesis and mineralization with a storage modulus enhancement of 2.5-fold after 30 days. Meanwhile, the appropriate matrix microenvironment initiated angiogenesis and early osteogenesis by accelerating endogenous stem cell recruitment in situ. Together, the PHCLS allowed substantial skull reconstruction in the rabbit cranial defect model, achieving 85.2% breaking load strength and 84.5% bone volume fractions in comparison to the natural cranium, 12 weeks after implantation. Overall, this study reveals that the hierarchical microporous hydrogel scaffold provides a promising strategy for skull defect treatment.

Nanostructured 3D-Printed Hybrid Scaffold Accelerates Bone Regeneration by Photointegrating Nanohydroxyapatite.

Nanostructured biomaterials that replicate natural bone architecture are expected to facilitate bone regeneration. Here, nanohydroxyapatite (nHAp) with vinyl surface modification is acquired by silicon-based coupling agent and photointegrated with methacrylic anhydride-modified gelatin to manufacture a chemically integrated 3D-printed hybrid bone scaffold (75.6wt% solid content). This nanostructured procedure significantly increases its storage modulus by 19.43-fold (79.2kPa) to construct a more stable mechanical structure. Furthermore, biofunctional hydrogel with biomimetic extracellular matrix is anchored onto the filament of 3D-printed hybrid scaffold (HGel-g-nHAp) by polyphenol-mediated multiple chemical reactions, which contributes to initiate early osteogenesis and angiogenesis by recruiting endogenous stem cells in situ. Significant ectopic mineral deposition is also observed in subcutaneously implanted nude mice with storage modulus enhancement of 25.3-fold after 30 days. Meanwhile, HGel-g-nHAp realizes substantial bone reconstruction in the rabbit cranial defect model, achieving 61.3% breaking load strength and 73.1% bone volume fractions in comparison to natural cranium 15 weeks after implantation. This optical integration strategy of vinyl modified nHAp provides a prospective structural design for regenerative 3D-printed bone scaffold.

Covalently Grafted Biomimetic Matrix Reconstructs the Regenerative Microenvironment of the Porous Gradient Polycaprolactone Scaffold to Accelerate Bone Remodeling.

Integrating a biomimetic extracellular matrix to improve the microenvironment of 3D printing scaffolds is an emerging strategy for bone substitute design. Here, a "soft-hard" bone implant (BM-g-DPCL) consisting of a bioactive matrix chemically integrated on a polydopamine (PDA)-coated porous gradient scaffold by polyphenol groups is constructed. The PDA-coated "hard" scaffolds promoted Ca2+ chelation and mineral deposition; the "soft" bioactive matrix is beneficial to the migration, proliferation, and osteogenic differentiation of stem cells in vitro, accelerated endogenous stem cell recruitment, and initiated rapid angiogenesis in vivo. The results of the rabbit cranial defect model (Φ= 10mm) confirmed that BM-g-DPCL promoted the integration between bone tissue and implant and induced the deposition of bone matrix. Proteomics confirmed that cytokine adhesion, biomineralization, rapid vascularization, and extracellular matrix formation are major factors that accelerate bone defect healing. This strategy of highly chemically bonded soft-hard components guided the construction of the bioactive regenerative scaffold.

Dopamine‐Integrated Nanointerface between Fibrillar Matrix and Hydrophilic Nanohydroxyapatite Regulates Immune Microenvironment to Boost Endogenous Bone Regeneration

AbstractDriving endogenous bone regeneration by cell‐ and factor‐free biomaterials is the most ideal repair strategy. Herein, hybrid interleaved scaffold (HDSH) with nanosized interfacial integration is assembled by organic/inorganic interactive bonding at the nanoscale. With the help of transcriptome and proteome analysis, the regenerative mechanism of this scaffold is elaborated at the molecular level, which confirms that this strategy recreates a suitable immune microenvironment (anti‐inflammatory and M2‐polarizing) and drives functional cell and cytokine adhesion, as well as inchoate vascularization. It greatly enhances endogenous stem cell recruitment, and subsequently initiates robust vasculogenesis and osteogenesis. Significant bony reconstitution in the rabbit cranial defect model (Φ = 10 mm) is observed after 12 weeks, which realizes completely new bone coverage and 79% breaking load strength relative to the natural cranium. By enhancing nano‐sized functional interfacial integration, this strategy can provide effective guidance for developing highly bioactive bone‐regenerative implants.

Development of a Novel Perfusion Rotating Wall Vessel Bioreactor with Ultrasound Stimulation for Mass-Production of Mineralized Tissue Constructs.

As stem cells are considered a promising cell source for tissue engineering, many culture strategies have been extensively studied to generate in vitro stem cell-based tissue constructs. However, most approaches using conventional tissue culture plates are limited by the lack of biological relevance in stem cell microenvironments required for neotissue formation. In this study, a novel perfusion rotating wall vessel (RWV) bioreactor was developed for mass-production of stem cell-based 3D tissue constructs. An automated RWV bioreactor was fabricated, which is capable of controlling continuous medium perfusion, highly efficient gas exchange with surrounding air, as well as low-intensity pulsed ultrasound (LIPUS) stimulation. Embryonic stem cells encapsulated in alginate/gelatin hydrogel were cultured in the osteogenic medium by using our bioreactor system. Cellular viability, growth kinetics, and osteogenesis/mineralization were thoroughly evaluated, and culture media were profiled at real time. The in vivo efficacy was examined by a rabbit cranial defect model. Our bioreactor successfully maintained the optimal culture environments for stem cell proliferation, osteogenic differentiation, and mineralized tissue formation during the culture period. The mineralized tissue constructs produced by our bioreactor demonstrated higher void filling efficacy in the large bone defects compared to the group implanted with hydrogel beads only. In addition, the LIPUS modules mounted on our bioreactor successfully reached higher mineralization of the tissue constructs compared to the groups without LIPUS stimulation. This study suggests an effective biomanufacturing strategy for mass-production of implantable mineralized tissue constructs from stem cells that could be applicable to future clinical practice.

Shapable bulk agarose–gelatine–hydroxyapatite–minocycline nanocomposite fabricated using a mineralizing system aided with electrophoresis for bone tissue regeneration

To develop a shapable bulk antibacterial nanocomposite biomaterial for bone regeneration. A bulk agarose–gelatine hydrogel was mineralized using a hydrogel mineralizing system aided with electrophoresis, and the mineralized hydrogel was loaded with minocycline to obtain the agarose–gelatine–hydroxyapatite–minocycline nanocomposite. The nanocomposite had a large Brunauer–Emmett–Teller surface area of 44.4518 m2 g−1 and a high porosity of 76.9%. Hydroxyapatite crystals were well developed in the hydrogel matrix and exhibited a hybrid structure of microscale and nanoscale motifs. The addition of minocycline resulted in a continuous antibiotic release, inhibiting the growth of Staphylococcus aureus over 2 weeks in vitro. Exposed to rabbit bone marrow mesenchymal stem cells, the nanocomposite revealed good cytocompatibility in vitro. Furthermore, the biomaterial could effectively enhance the bone regeneration in a critical-size rabbit cranial defect model in vivo. These findings depicted that the nanocomposite, with good biocompatibility and good antibacterial property, is a promising candidate for future clinical application in bone tissue engineering or as a prospective bone replacement biomaterial.

Human amniotic mesenchymal stromal cells promote bone regeneration via activating endogenous regeneration.

Rationale: The effectiveness of stem cell based-therapy for bone regeneration has been demonstrated; yet, clinical application of autologous stem cells is still limited by invasive acquisition, long culture processes, and high cost. Besides, it remains controversial whether autologous stem cells could directly participate in tissue repair after differentiation. Thus, increasing allogeneic stem cells have been developed into drugs to indirectly activate endogenous regeneration and induce tissue regeneration. Human amniotic mesenchymal stromal cells (HAMSCs) have been extensively studied, showing multiple regulatory functions, but mechanisms of HAMSCs in promoting bone regeneration are remain unclear.Methods: Proteome profile of HAMSCs and their functions on vascularized bone regeneration were investigated in vitro, while rabbit cranial defect model was used to further detect the effects of bone formation in vivo.Results: HAMSCs secrete many osteogenic, angiogenic, and immunomodulatory cytokines. In vitro, HAMSCs can promote human bone-marrow mesenchymal stromal cells (HBMSCs) migration and osteogenic differentiation; promote the capillary-tube formation of human umbilical vascular endothelial cells (HUVECs), induce HUVECs migration and pro-angiogenic genes expression, and promote M2 macrophage polarization. Further, in vivo studies suggested that transplanted HAMSCs could survive and induce M2 macrophages to secrete bone morphogenetic protein-2 (BMP-2) and vascular endothelial growth factor (VEGF) in rabbits' skull defects at an early stage, and, in turn, promote more new bone formation.Conclusion: HAMSCs have good biocompatibility and paracrine function to promote bone repair by stimulating endogenous regeneration.

Evaluating the bioactivity of a hydroxyapatite-incorporated polyetheretherketone biocomposite

BackgroundPolyetheretherketone (PEEK) exhibits stable chemical properties, excellent biocompatibility, and rational mechanical properties that are similar to those of human cortical bone, but the lack of bioactivity impedes its clinical application.MethodsIn this study, hydroxyapatite (HA) was incorporated into PEEK to fabricate HA/PEEK biocomposite using a compounding and injection-molding technique. The tensile properties of the prepared HA/PEEK composites (HA content from 0 to 40 wt%) were tested to choose an optimal HA content. To evaluate the bioactivity of the composite, the cell attachment, proliferation, spreading and alkaline phosphatase (ALP) activity of MC3T3-E1 cells, and apatite formation after immersion in simulated body fluid (SBF), and osseointegration in a rabbit cranial defect model were investigated. The results were compared to those from ultra-high molecular weight polyethylene (UHMWPE) and pure PEEK.ResultsBy evaluating the tensile properties and elastic moduli of PEEK composite samples/PEEK composites with different HA contents, the 30 wt% HA/PEEK composite was chosen for use in the subsequent tests. The results of the cell tests demonstrated that PEEK composite samples/PEEK composite exhibited better cell attachment, proliferation, spreading, and higher ALP activity than those of UHMWPE and pure PEEK. Apatite islands formed on the HA/PEEK composite after immersion in SBF for 7 days and grew continuously with longer time periods. Animal tests indicated that bone contact and new bone formation around the HA/PEEK composite were more obvious than those around UHMWPE and pure PEEK.ConclusionsThe HA/PEEK biocomposite created by a compounding and injection-molding technique exhibited enhanced osteogenesis and could be used as a candidate of orthopedic implants.

Sustained Delivery of rhBMP-2 by Means of Poly(Lactic-co-Glycolic Acid) Microspheres

Commercially available recombinant human bone morphogenetic protein 2 (rhBMP2) has demonstrated efficacy in bone regeneration, but not without significant side effects. The authors used rhBMP2 encapsulated in poly(lactic-co-glycolic acid) (PLGA) microspheres placed in a rabbit cranial defect model to test whether low-dose, sustained delivery can effectively induce bone regeneration. The rhBMP2 was encapsulated in 15% PLGA using a double-emulsion, solvent extraction/evaporation technique, and its release kinetics and bioactivity were tested. Two critical-size defects (10 mm) were created in the calvaria of New Zealand white rabbits (5 to 7 months of age, male and female) and filled with a collagen scaffold containing either (1) no implant, (2) collagen scaffold only, (3) PLGA-rhBMP2 (0.1 μg per implant), or (4) free rhBMP2 (0.1 μg per implant). After 6 weeks, the rabbits were killed and defects were analyzed by micro-computed tomography, histology, and finite element analysis. The rhBMP2 delivered by means of bioactive PLGA microspheres resulted in higher volumes and surface area coverage of new bone than an equal dose of free rhBMP2 by micro-computed tomography (p=0.025 and p=0.025). Finite element analysis indicated that the mechanical competence using the regional elastic modulus did not differ with rhBMP2 exposure (p=0.70). PLGA-rhBMP2 did not demonstrate heterotopic ossification, craniosynostosis, or seroma formation. Sustained delivery by means of PLGA microspheres can significantly reduce the rhBMP2 dose required for de novo bone formation. Optimization of the delivery system may be a key to reducing the risk for recently reported rhBMP2-related adverse effects.

The effect of rhBMP-2 and PRP delivery by biodegradable β-tricalcium phosphate scaffolds on new bone formation in a non-through rabbit cranial defect model

This study evaluated whether the combination of biodegradable β-tricalcium phosphate (β-TCP) scaffolds with recombinant human bone morphogenetic protein-2 (rhBMP-2) or platelet-rich plasma (PRP) could accelerate bone formation and increase bone height using a rabbit non-through cranial bone defect model. Four non-through cylindrical bone defects with a diameter of 8-mm were surgically created on the cranium of rabbits. β-TCP scaffolds in the presence and absence of impregnated rhBMP-2 or PRP were placed into the defects. At 8 and 16 weeks after implantation, samples were dissected and fixed for analysis by microcomputed tomography and histology. Only defects with rhBMP-2 impregnated β-TCP scaffolds showed significantly enhanced bone formation compared to non-impregnated β-TCP scaffolds (P < 0.05). Although new bone was higher than adjacent bone at 8 weeks after implantation, vertical bone augmentation was not observed at 16 weeks after implantation, probably due to scaffold resorption occurring concurrently with new bone formation.

Development of a guided bone regeneration device using salicylic acid‐poly(anhydride‐ester) polymers and osteoconductive scaffolds

Successful repair of craniofacial and periodontal tissue defects ideally involves a combined therapy that includes inflammation modulation, control of soft tissue infiltration, and bone regeneration. In this study, an anti-inflammatory polymer, salicylic acid-based poly(anhydride-ester) (SAPAE) and a three-dimensional osteoconductive ceramic scaffold were evaluated as a combined guided bone regeneration (GBR) system for concurrent control of inflammation, soft tissue ingrowth, and bone repair in a rabbit cranial defect model. At time periods of 1, 3, and 8 weeks, five groups were compared: (1) scaffolds with a solid ceramic cap (as a GBR structure); (2) scaffolds with no cap; (3) scaffolds with a poly(lactide-glycolide) cap; (4) scaffolds with a slow release SAPAE polymer cap; and (5) scaffolds with a fast release SAPAE polymer cap. Cellular infiltration and bone formation in these scaffolds were evaluated to assess inflammation and bone repair capacity of the test groups. The SAPAE polymers suppressed inflammation and displayed no deleterious effect on bone formation. Additional work is warranted to optimize the anti-inflammatory action of the SAPAE, GBR suppression of soft tissue infiltration, and stimulation of bone formation in the scaffolds and create a composite device for successful repair of craniofacial and periodontal tissue defects.

Negative pressure technology enhances bone regeneration in rabbit skull defects

BackgroundBone is a slowly regenerating tissue influenced by various physiological processes, including proliferation, differentiation, and angiogenesis, under the control of growth factors. Shortening this healing time is an important and popular clinical research focus in orthopedics. Negative pressure can stimulate angiogenesis, improve blood circulation, promote granulation tissue growth and accelerate tissue wound healing. We sought to determine whether negative pressure could reduce bone healing time in a rabbit cranial defect model.MethodsFour symmetrical holes (diameter, 3.5 mm) were drilled into the skulls of 42 New Zealand white rabbits, with two holes in each parietal bone. For each rabbit, the two sides were then randomly assigned into experimental and control groups. Using negative pressure suction tubes, experimental holes were treated with −50 kPa for 15 minutes, four times per day, whereas the control holes remained untreated. After 4 weeks, the negative pressure suction tubes were removed. At 2, 4, 6 and 8 weeks, three-dimensional (3D) reconstruction computed tomography (CT), X-ray radiopacity, and two-photon absorptiometry were used to evaluate new bone formation. Histological changes were determined by hematoxylin and eosin (H.E) staining. At weekly intervals until 6 weeks, the mRNA expression levels of vascular endothelial growth factor (VEGF) and bone morphogenetic protein (BMP)-2 were evaluated by RT-PCR. A paired student’s t-test was employed to compare X-ray radiopacity and bone density measurements between the experimental and control groups.Results3D-reconstruction CT showed that new bone regeneration in the experimental group was greater than that in the control group at 4 and 6 weeks. At these time points, the experimental group presented with higher X-ray radiopacity and increased bone density (P < 0.05) as compared with the control group. Cartilage islands and new bone were observed by H.E staining at 2 weeks in the experimental group. By 6 weeks, the new bone had matured into lamellar bone in the experimental group. RT-PCR results showed that VEGF and BMP-2 were highly expressed in the experimental group as compared with control.ConclusionsIntermittent negative pressure can promote the regeneration of bone possibly by enhancing the expression of VEGF and BMP-2.

Cranial Particulate Bone Graft Ossifies Calvarial Defects by Osteogenesis

Cranial particulate bone graft heals inlay calvarial defects and can be harvested as early as infancy. The purpose of this study was to test the hypothesis that particulate bone promotes ossification primarily by osteogenesis. Freshly harvested particulate bone, devitalized particulate bone, and high-speed drilled bone dust from rabbit calvaria were assayed for metabolic activity (resazurin) and viable osteoblasts (alkaline phosphatase). A rabbit cranial defect model was used to test the effect of devitalizing particulate bone on in vivo ossification. A parietal critical-size defect was created and managed in three ways: (1) no implant (n = 6); (2) particulate bone implant (n = 6); and (3) devitalized particulate bone implant (n = 6). Micro-computed tomographic scanning was used to measure ossification 16 weeks later; histology also was studied. Particulate bone contained more viable cells (0.94 percent transmittance per milligram) compared with devitalized particulate bone (0.007 percent) or bone dust (0.21 percent) (p = 0.01). Particulate bone had greater alkaline phosphatase activity (0.13 μU/μg) than devitalized particulate bone (0.000) or bone dust (0.06) (p = 0.01). Critical-size defects treated with particulate bone had more ossification (99.7 percent) compared with devitalized particulate bone implants (42.2 percent) (p = 0.01); no difference was found between devitalized particulate bone and the control (40.8 percent) (p = 0.9). Particulate bone graft contains living cells, including osteoblasts, that are required to heal critical-size cranial defects. These data support the hypothesis that particulate bone promotes ossification primarily by osteogenesis.

Bioelectric effect of a chitosan bioelectret membrane on bone regeneration in rabbit cranial defects

In this study, the effects of a chitosan bioelectret membrane on bone regeneration in a rabbit cranial defect model were examined. The bioelectret was fabricated by film casting and polarized by grid-controlled corona charging (−1 kV). In vitro, the equivalence surface charge density of the bioelectret gradually reduced under both wet and dry conditions. In vivo, two rectangular transosseous defects of 8 mm × 10 mm were created symmetrically on each rabbit cranial bone. The defects were treated with the bioelectret membrane, with unpolarized membrane, and no membrane, respectively. The rabbits were killed at 4, 8, and 12 weeks postoperatively, and specimens were examined by micro-computed tomography and histological analysis. The bioelectret membrane recipients had a significantly higher volume of newly formed bone and greater rate of material degradation than the unpolarized membrane. Furthermore, the bioelectret membrane induced new bone formation not only around the host bone but also in the center of the defects. The chitosan bioelectret membrane application has an apparent potential in guided bone regeneration applications.

Evaluation of bone replacement materials in a rabbit cranial defect model using micro CT and hard tissue histology

Introduction: For functional and structural restoration of bone deficiencies, various resorbable and nonresorbable alloplastic materials have been introduced, including metals, polymers and ceramics. However, an “optimal” artificial replacement for craniofacial bone has not been found yet and the search for improved reconstruction methods and alternative materials is going on. To assess and compare biocompatibility and osseointegration of these materials, adequate animal test models are indispensable. Methods: In a rabbit cranial defect model, biocompatibility and osseointegration of polymeric and composite bone replacement materials were evaluated at different time points after implantation. Calvaria including implants and surrounding tissue were explanted and embedded in methacrylate resin. The samples were scanned with a nanotom® (phoenix|x-ray) µCT scanner and proceeded for histological examination by sawing-grinding technique. Avizo® Fire (vsg) software was used for visualisation and processing of µCT data. Qualitative and morphometric evaluation of osseointegration and fibrous encapsulation was performed on undecalcified histologic preparations of the explants, and on 3D reconstructions plus virtual slices derived from corresponding 3D µCT datasets. Results: The obtained 3D µCT data enabled a comprehensive qualitative and quantitative assessment of osseointegration and biodegradation of radioopaque composite implants. Prerequisite for visualization and discrimination of materials by µCT is a significant difference of their hounsfield values. Due to this limitation, radiolucent polymeric implant materials and soft tissue could not be distinguished from embedding resin. In contrast, histologic preparations of undecalcified hard tissue and implant materials enabled detailed visualization and examination of all tissues and implant materials. The substantial disadvantage of hard tissue histology was the inevitable loss of information due to small number of slices and large gaps between specimens yielded by this method. Conclusion: To obtain comprehensive and quantifiable information about biodegradation, biocompatibility and osseointegration of alloplastic bone replacement materials, µCT scans as well as histologic evaluation should be performed.

Bone regeneration with glass ceramic implants and calcium phosphate cements in a rabbit cranial defect model

Hydroxyapatite cement (BoneSource®) and brushite calcium phosphate cement (chronOS™ Inject) were tested for fixation of glass ceramic implants (Bioverit®) in experimentally created cranial defects in 24 adult New Zealand White rabbits. Aim of the in vivo study was to assess and compare the biocompatibility and osseointegration of the implanted materials. Macroscopic and histological evaluations were performed 1 month, 3 months, and 6 months postoperatively. All implanted materials were well tolerated by the surrounding tissue. Both bone cements exhibited osteoconductive properties. Differences could be detected regarding to the rates of cement resorption and new bone formation. The brushite cement was resorbed faster than the hydroxyapatite cement. The chronOS™ Inject samples exhibited a higher rate of connective tissue formation and an insufficient osseointegration. BoneSource® was replaced by bone with minimal invasion of connective tissue. New bone formation occurred faster compared to the chronOS™ Inject group. Bioverit® implants fixed with BoneSource® were successfully osseointegrated.

Correlation Between Micro-Computed Tomography and Histomorphometry for Assessment of New Bone Formation in a Calvarial Experimental Model

Conventional histologic or histomorphometric evaluation provides clear evidence of the bone healing process. However, the sample preparation process is tedious and destructive, and the three-dimensional (3D) anisotropic information of the bone trabeculae is compromised. Micro-computed tomography (microCT) has been introduced as an alternative to these traditional evaluation methods. microCT is noninvasive and provides a faster approach to evaluate and quantify cancellous bone. Most previous studies that used microCT have focused on studying trabecular structures of cancellous bone. In this study, we used microCT to analyze the micro-architecture of the regenerated membranous bone using a rabbit cranial defect model. Two 1 cm diameter circular bony defects were created in 12 New Zealand white rabbits. Specimens were harvested at 6 weeks and 12 weeks after surgery and were scanned using a MicroCT machine (Skyscan 1072, Aartselaar, Belgium). The specimens were then sectioned and stained with Goldner's trichrome. Bone volume density (BV/TV), bone surface density (BS/BV), and trabecular thickness (TbTh) were determined from histomorphometric and two-dimensional (2D) and 3D microCT analysis. Pearson's correlation coefficient (gamma), paired t-tests, and intraclass correlation coefficients from measurements between the 2D and 3D microCT and histomorphometry were calculated. There were very strong positive correlations of BV/TV between histomorphometric and 2D or 3D microCT measurements. Correlation between histomorphometric and 2D microCT measurements for BS/BV was moderate, whereas correlation between histomorphometric and 3D microCT measurements was weak. Weak correlations in TbTh among the three methods were found. In conclusion, the present study suggests that, in evaluating micro-architectures in regenerated bones, the correlation between measuring methods vary according to the features measured.

Cranial Defect Reconstruction in an Experimental Model Using Different Mixtures of Bioglass and Autologous Bone

The purpose of this study was to investigate bone healing in a rabbit cranial defect model using different mixtures of bioglass (NovaBone C/M; Porex Surgical, Inc., Newnan, GA) and autologous bone with and without an overlying absorbable plate (Lactosorb; Walter Lorenz Surgical, Jacksonville, FL). Twelve rabbits were divided into three groups, and a 2 cm diameter cranial defect was created in each rabbit. Group I defects were filled with 80% bioglass and 20% autologous bone, group II with 60% bioglass and 40% autologous bone, and group III with 60% bioglass and 40% autologous bone with the addition of an absorbable plate placed directly over the reconstructed defect to help separate it from the overlying soft tissues. Rabbits were euthanized 6 months postoperatively. Histologic examination was then performed. The size of the remaining bone gap, area of reossification within the defect, and percentage of fibrous tissue within the defect were measured. Histologic analyses revealed that group II animals had an increased cross-sectional area of bone formation and decreased bone gap when compared with group I and III animals (P < 0.05). There were no statistical differences between groups I and III with regard to bone formation. This study suggests that when combining bioactive glass and autologous bone for repairing cranial defects, a combination of 60% bioactive glass and 40% bone graft yields superior results to a combination of 80% bioactive glass and 20% bone graft. Placing an absorbable plate over a defect filled with 60% bioactive glass and 40% bone graft inhibits rather than promotes reossification.