Rabbit Cancellous Bone Research Articles

Bone formation in rabbit cancellous bone explant culture model is enhanced by mechanical load

BackgroundWhen studying and designing an artificial bone in vitro with similar features and functionality of natural bone by tissue engineering technology, the culturing environment, especially the mechanical environment is supposed to be an important factor, because a suitable mechanical environment in vitro may improve the adaptability of the planted-in tissue engineering bone in the body. Unfortunately, up to now, the relationship between mechanical stimuli and natural bone growth has not yet been precisely determined, and it is so imperative for a prior study on effect of mechanical loading on growth of the natural bone cultured in vitro.MethodsUnder sterile conditions, explant models of rabbit cancellous bone with 3 mm in thickness and 8 mm in diameter were prepared and cultured in a dynamic loading and circulating perfusion bioreactor system. By Micro-CT scanning, a 3D model for finite element (FEM) analysis was achieved. According to the results of FEM analysis and physiological load bearing capacity of the natural bone, these models were firstly subjected to mechanical load with 1Hz frequency causing average apparent strain of 1000 μϵ, 2000 μϵ, 3000 μϵ and 4000 μϵ respectively for 30 min every day, activities of alkaline phosphatase (AKP) were detected on the 5th and the 14th loading day and on the 14th and the 21st day, mechanical properties, tissue mineral density (TMD) of the bone explant models were investigated and Von-kossa staining and fluorescence double labeling assays were conducted to evaluate whether there were fresh osteoid in the bone explant models. In addition, Western blot, Elisa and Real-time PCR were employed to analyze expression of Collagen-I (COL-1), bone morphogenetic protein-2 (BMP-2) and osteoprotegerin (OPG) protein and RNA.ResultsThe explant models of rabbit cancellous bone prepared under sterile conditions grew well in the bioreactor system. With the increasing culturing time and load levels, bone explant models in groups with 1000 μϵ and 2000 μϵ average apparent strain experienced improving mechanical properties and TMD (P<0.05), and results of Von-kossa staining and fluorescence double labeling also showed apparent fresh osteoid formation. Under the same loading conditions, a up-regulations in protein and RNA of COL-1, BMP-2 and OPG were detected, especially, relative genes notably expressed after 21 days.ConclusionOur study demonstrated that mechanical load could improve function and activity of osteoblasts in explant models of cancellous bone. Through regulations of COL-1, OPG and BMP-2 secreted by osteoblasts, the mechanical load could improve the tissue structural density and stiffness due to formation of fresh osteoid.

In-vitro analysis of the effect of gentamicin and polyhexanide on bone tissue

Though anti-infectives have been used for a long time in surgical procedures, the effect on bone tissue has not been determined for most antibiotics and antiseptics. In our in vitro study, 4x4x8 mm(3) blocks of rabbit cancellous bone tissue were incubated with Ringer's solution, gentamicin and Lavasorb(®) each for time intervals of 15 minutes, 30 minutes, one hour, four hours and eight hours. Samples were examined double blinded through optical and electron microscopy. Tissue degeneration was observed in all samples. It was low in Ringer's solution. Samples with Lavasorb showed a moderate degeneration after 15 and 30 minutes, which was accelerated after one hour. Gentamicin led to a moderate degeneration of bone tissue after 15 and 30 minutes and to a more accelerated degeneration after one hour. The effect of gentamicin on bone tissue was more pronounced than the effect of Lavasorb. This investigation showed that local application of Lavasorb or gentamicin on bone tissue should be restricted to 30 minutes, while Lavasorb showed a better tissue tolerability. This finding could have clinical implications for the management of wounds with open osseous tissue and should be further investigated by in vivo studies.

Positive modulation of osteogenesis‐ and osteoclastogenesis‐related gene expression with strontium‐containing microstructured Ti implants in rabbit cancellous bone

This study investigated the in vivo osteoblast and osteoclast gene expression in the peri-implant bone tissue of the strontium (Sr)-incorporated microstructured Ti implants (SLA/Sr) in order to evaluate whether local Sr delivery to the implant surface as in the form of SrTiO(3) also exerts original bone healing enhancement effect of Sr that take place through the dual modes of action of a stimulation of osteogenesis and an inhibition of osteoclastogenesis. The in vivo mRNA expression of osteoblast- and osteoclast-related genes in the peri-implant bone and removal torque forces of the SLA/Sr implants were compared with a chemically modified super-hydrophilic SLA implants (SLActive®) in rabbit cancellous bone after 2 weeks of implantation. There was no significant difference in the removal torque values between the two groups. Both the torque-tested SLA/Sr and SLActive implants exhibited a considerable quantity of bone attached to the surface. Real-time PCR analysis revealed notably increased mRNA expression of osteoblast genes (Runx2, osterix and osteocalcin) in the peri-implant bone of the SLA/Sr implants compared with the SLActive implants (p < 0.05), whereas osteoclast phenotype gene (TRAP) expression was markedly decreased in the SLA/Sr implant (p < 0.05). The results at the molecular level suggest that local Sr delivery as in the form of the Sr-incorporated Ti oxide layer favors early osseointegration of microrough Ti implants via a positive modulation of bone healing, that is, a promotion of osteoblast differentiation and suppression of osteoclastogenesis, in the bone-implant interface of cancellous bone.

The relative effect of surface strontium chemistry and super‐hydrophilicity on the early osseointegration of moderately rough titanium surface in the rabbit femur

It is unclear whether surface bioactive chemistry or hydrophilicity plays a more dominant role in the osseointegration of micro-structured titanium implants having the same surface topography at the micrometer and submicrometer scales. To understand their comparative effect on enhancing the early osseointegration of micro-rough-surfaced implants, this study compared the bone healing-promoting effect of surface strontium (Sr) chemistry that has been shown in numerous studies to super-hydrophilicity in the early osseointegration of moderately rough-surfaced clinical oral implants (SLA(®) implant) in rabbit cancellous bone. Hydrothermal treatment was performed to incorporate Sr ions into the surface of clinical SLA implants (SLA/Sr implant). The surface characteristics were evaluated by using field emission-scanning electron microscopy, X-ray photoelectron spectroscopy and optical profilometry. Twenty screw implants (10 control and 10 experimental) were placed in the femoral condyles of 10 New Zealand White rabbits. The early osseointegration of the SLA/Sr implant was compared with a chemically modified super-hydrophilic SLA implant (SLActive(®) implant) by histomorphometric and resonance frequency analysis after 2 weeks of implantation. The SLA/Sr and SLActive implants exhibited an identical surface topography and average R(a) values at the micron and submicron scales. The SLA/Sr implant displayed a high amount of surface Sr content (15.6 at.%). There was no significant difference in the implant stability quotient (ISQ) values between the two groups. However, histomorphometric analysis revealed a significantly higher bone-to-implant contact percentage in the SLA/Sr implants compared with the SLActive implants in rabbit cancellous bone (P < 0.01). The results indicate that the surface Sr chemistry surpasses the effect of super-hydrophilicity in promoting the early bone apposition of moderately rough Ti surface in cancellous bone.

Bone Healing Using a Bi-Phasic Ceramic Bone Substitute Demonstrated in Human Vertebroplasty and with Histology in a Rabbit Cancellous Bone Defect Model

A non-exothermic material that demonstrates clinical pain relief comparable to polymethylacrylate (PMMA) for vertebroplasty and promotes bone healing is desirable. The purpose of this investigation is to demonstrate clinical pain score improvement and bone healing following vertebroplasty with a novel bi-phasic ceramic cement. Twenty patients were prospectively treated for compression fractures in a single center in the USA with the injectable bi-phasic ceramic bone substitute. Statistical comparison of pain scores was made during a 12 month follow-up retrospectively against a matched cohort of patients treated with PMMA vertebroplasty by the same neuroradiologist (HPH) in the same setting. The bone remodeling material was also evaluated with histology in a New Zealand white rabbit model. The bi-phasic material demonstrated a pre-operative mean VAS score of 8.5 (± 1.6) with a significant post-operative pain relief mean VAS score of 1.8 (± 2.5) after one week, which was maintained throughout the 12 month follow-up period. These data are in line with the pain scores for the PMMA treated cohort. CT scans six and 12 months after surgery with the bi-phasic cement showed healing of the osteoporotic fractures. In the rabbit model, histology with the study material showed evidence of incorporation, new bone growth and bone healing in a cancellous bone defect. Both the clinical results and the histologic evidence of bone healing and new bone growth support the application of this new bioinjectable material as an alternative to the use of PMMA for vertebroplasty.

Osseointegration of two different phosphate ion‐containing titanium oxide surfaces in rabbit cancellous bone

This study assessed the osseointegration of grit-blasted titanium (Ti) implants with a hydrophilic phosphate ion-incorporated oxide surface in rabbit cancellous bone, and compared its bone healing with commercially available phosphate-incorporated clinical implants obtained by micro-arc oxidation (TiUnite, TU implant). The hydrophilic phosphate-incorporated Ti surface (P implant) was produced by hydrothermal treatment on grit-blasted moderately rough-surfaced clinical implant. The TU surface was used as a control. The surface characteristics were evaluated by field emission-scanning electron microscopy, X-ray photoelectron spectroscopy, optical profilometry, and inductively coupled plasma-atomic emission spectroscopy (ICP-AES). Thirty-two threaded implants with lengths of 10 and 3.3 mm diameter (16 P implants and 16 TU implants) were placed in the femoral condyles of 16 New Zealand White rabbits. Histomorphometric analysis, removal torque tests, and surface analysis of the torque-tested implants were performed 4 weeks after implantation. The P and TU implants displayed micro-rough surface features with similar Ra values at the micron-scale. ICP-AES analysis revealed that both the P and TU implants released phosphate ions into the solution. The torque-tested P and TU implants exhibited a considerable quantity of bone attached to the surface. The P implants exhibited significantly higher bone-implant contact percentages, both in terms of the all threads region and the total lateral length of implants compared with the TU implants (P < 0.01), but no statistical difference was found for the removal torque values. These results suggest that the phosphate-incorporated Ti oxide surface obtained by hydrothermal treatment achieves rapid osseointegration in cancellous bone by increasing the degree of bone-implant contact.

Bone healing of commercial oral implants with RGD immobilization through electrodeposited poly(ethylene glycol) in rabbit cancellous bone

Immobilization of RGD peptides on titanium (Ti) surfaces enhances implant bone healing by promoting early osteoblastic cell attachment and subsequent differentiation by facilitating integrin binding. Our previous studies have demonstrated the efficacy of RGD peptide immobilization on Ti surfaces through the electrodeposition of poly(ethylene glycol) (PEG) (RGD/PEG/Ti), which exhibited good chemical stability and bonding. The RGD/PEG/Ti surface promoted differentiation and mineralization of pre-osteoblasts. This study investigated the in vivo bone healing capacity of the RGD/PEG/Ti surface for biomedical application as a more osteoconductive implant surface in dentistry. The RGD/PEG/Ti surface was produced on an osteoconductive implant surface, i.e. the grit blasted micro-rough surface of a commercial oral implant. The osteoconductivity of the RGD/PEG/Ti surface was compared by histomorphometric evaluation with an RGD peptide-coated surface obtained by simple adsorption in rabbit cancellous bone after 2 and 4weeks healing. The RGD/PEG/Ti implants displayed a high degree of direct bone apposition in cancellous bone and achieved greater active bone apposition, even in areas of poor surrounding bone. Significant increases in the bone to implant contact percentage were observed for RGD/PEG/Ti implants compared with RGD-coated Ti implants obtained by simple adsorption both after 2 and 4weeks healing (P<0.05). These results demonstrate that RGD peptide immobilization on a Ti surface through electrodeposited PEG may be an effective method for enhancing bone healing with commercial micro-rough surface oral implants in cancellous bone by achieving rapid bone apposition on the implant surface.

Osseointegration of commercial microstructured titanium implants incorporating magnesium: a histomorphometric study in rabbit cancellous bone

Recent studies have suggested that magnesium (Mg) ions exert a beneficial effect on implant osseointegration. This study assessed the osseointegration of nanoporous titanium (Ti) surface incorporating the Mg produced by hydrothermal treatment in rabbit cancellous bone to determine whether this surface would further enhance bone healing of moderately rough-surfaced implants in cancellous bone, and compared the result with commercially available micro-arc oxidized Mg-incorporated implants. The Mg-incorporated Ti surfaces (RBM/Mg) were obtained by hydrothermal treatment using an alkaline Mg-containing solution on grit-blasted moderately rough (RBM) implants. Untreated RBM and recently introduced Mg-incorporated microporous Ti implants produced by micro-arc oxidation (M) were used controls in this study. The surface characteristics were evaluated by scanning electron microscopy, X-ray photoelectron spectroscopy and optical profilometry. Twenty-four threaded implants with a length of 10 mm (eight RBM implants, eight RBM/Mg implants and eight M implants) were placed in the femoral condyles of 12 New Zealand White rabbits. Histomorphometric analysis was performed 4 weeks after implantation. Hydrothermally treated and untreated grit-blasted implants displayed almost identical surface morphologies and R(a) values at the micron-scale. The RBM/Mg implants exhibited morphological differences compared with the RBM implants at the nano-scale, which displayed nanoporous surface structures. The Mg-incorporated implants (RBM/Mg and M) exhibited more continuous bone apposition and a higher degree of bone-to-implant contact (BIC) than the untreated RBM implants in rabbit cancellous bone. The RBM/Mg implants displayed significantly greater BIC% than untreated RBM implants, both in terms of the all threads region and the total lateral length of implants (P<0.05), but no statistical differences were found between the RBM/Mg and M implants except BIC% values in total lateral length. These results indicate that a nanoporous Mg-incorporated surface may be effective in enhancing the osseointegration of moderately rough grit-blasted implants by increasing the degree of bone-implant contact in areas of cancellous bone.

Abstract No. 191: Bone healing demonstrated in human vertebroplasty using a bi-phasic ceramic bone substitute with supporting histology in a rabbit cancellous bone defect model

Abstract No. 191: Bone healing demonstrated in human vertebroplasty using a bi-phasic ceramic bone substitute with supporting histology in a rabbit cancellous bone defect model

Bone regeneration potential of a soybean-based filler: experimental study in a rabbit cancellous bone defects

Autologous and allogenic bone grafts are considered as materials of choice for bone reconstructive surgery, but limited availability, risks of transmittable diseases and inconsistent clinical performances have prompted the development of alternative biomaterials. The present work compares the bone regeneration potential of a soybean based bone filler (SB bone filler) in comparison to a commercial 50:50 poly(D: ,L: lactide-glycolide)-based bone graft (Fisiograft((R)) gel) when implanted into a critical size defect (6-mm diameter, 10-mm length) in rabbit distal femurs. The histomorphometric and microhardness analyses of femoral condyles 4, 8, 16 and 24 weeks after surgery showed that no significant difference was found in the percentage of both bone repair and bone in-growth in the external, medium and inner defect areas. The SB filler-treated defects showed significantly higher outer bone formation and microhardness results at 24 weeks than Fisiograft((R)) gel (P < 0.05). Soybean-based biomaterials clearly promoted bone repair through a mechanism of action that is likely to involve both the scaffolding role of the biomaterial for osteoblasts and the induction of their differentiation.

Enhanced osteoconductivity of micro‐structured titanium implants (XiVE S CELLplus™) by addition of surface calcium chemistry: a histomorphometric study in the rabbit femur

This study evaluated the osseointegration in rabbit cancellous bone of titanium (Ti) implants with a micro-topographically complex surface structure produced by grit-blasting/acid-etching with or without the addition of surface calcium ion (Ca) chemistry. Micro-structured Ti implants (XiVE S CELLplus screw implant, Dentsply Friadent GmbH) were hydrothermally treated in an alkaline Ca-containing solution to produce a nano-structured Ca-incorporated oxide surface layer. The surface characteristics were evaluated by scanning electron microscopy and stylus profilometry before and after Ca surface treatment. Twenty implants (10 control and 10 experimental) were placed in the femoral condyles of 10 New Zealand White rabbits. Histomorphometric analysis was performed 6 weeks after implantation. Ca-incorporated and untreated control implants showed similar surface morphologies and surface roughness values at the micron scale. Untreated micro-structured Ti implants achieved a high degree of bone-to-implant contact (BIC), and Ca incorporation further increased BIC% (P<0.05). Active new bone apposition was found on surfaces of Ca-incorporated implants in areas of loose trabeculae. The nano-structured Ca-incorporated oxide surface significantly enhanced osteoconductivity of micro-structured Ti implants in rabbit cancellous bone. Results indicate that this surface produced by simple hydrothermal treatment may be effective in improving the osseointegration of implants with micro-topographically complex surface structures in areas of loose cancellous bone.

Is hydroxyapatite cement an alternative for allograft bone chips in bone grafting procedures? A mechanical and histological study in a rabbit cancellous bone defect model.

To evaluate in vivo performance of hydroxyapatite cement (HAC) as a porous bone graft substitute, HAC was mixed (1:1 ratio) with either porous calcium-phosphate granules (80% tricalcium phosphate, 20% hydroxyapatite) or defatted morsellized cancellous bone (MCB) allograft and implanted bilaterally in cylindrical drill holes in distal femurs of rabbits. Groups with empty defects and impacted MCB were used for reference. After 8 weeks, one femur from each pair was examined histologically. All contralateral specimens and Time-0 specimens were used for mechanical indentation tests. Histology showed that some empty defects were filled with newly formed osteopenic bone after 8 weeks. The impacted MCB showed remodeling into new vital bone. Incorporation of the HAC/MCB composite was incomplete, whereas minimal new bone ingrowth was found in the HAC/granule composites. Though not different from each other, both composites were significantly stronger than empty defects, incorporated impacted MCB, and intact cancellous bone. At Time 0, the mechanical behavior of impacted MCB was similar to both HAC composites. In conclusion, composites of HAC and porous biomaterials can maintain relatively high strength over 8 weeks in vivo, but their incorporation into a new bony structure is slower than impacted MCB. The HAC/MCB composite showed favorable incorporation behavior.

Effects of Surface Roughness and Maximum Load on the Mechanical Properties of Cancellous Bone Measured by Nanoindentation

AbstractNanoindentation was used to assess the mechanical properties of lamellar and interlamellar tissue in dehydrated rabbit cancellous bone. The effects of surface roughness and maximum nanoindentation load on the measured mechanical properties were examined in two samples of differing surface roughness using maximum loads ranging from 250-3000 μN. As the ratio of indentation depth to surface roughness decreased below approximately 3:1, the variability in material properties increased substantially. At low loads, the indentation modulus of the lamellar bone was approximately 20% greater than that of the interlamellar bone, while at high loads the measured properties of both layers converged to an intermediate value. Relatively shallow indentations made on smooth surfaces revealed significant differences in the properties of lamellar and interlamellar bone that are consistent with microstructural observations of lamellar bone as more mineralized than interlamellar bone.

Integration properties of bone substitute materials. Experimental studies on animals

In order to avoid the potential risks of disease transmission in allograft surgery, numerous substitute materials have been described. As the biological response to implant materials is different, we undertook the following study to assess type and amount of bone ingrowth in CaP-ceramics. 105 cylindrical bone defects with a diameter of 5.4 mm were created surgically in the femoral condyles of 53 skeletal mature NZW rabbits. The defects were filled with crushed coralline hydroxyapatite (HA) implants (n = 21), synthetically produced hydroxyapatite (n = 21) and surface-modified alpha-Tricalciumphosphate (TCP) grains (n = 21). 21 defects were left empty and other drill holes were filled with rabbit cancellous bone cylinders (n = 21) after 3 months of cryopreservation at -78 degrees C without sterilization. Following observation periods of 2, 4, 6, 8, 12, 26 and 52 weeks the femoral condyles were harvested for histological evaluation and quantitative analysis of bone ingrowth. Woven bone formation at implant periphery can be observed in all substances as early as 2 weeks postoperatively. At 4-week-intervals cryopreserved allografts show new bone apposition on surfaces of necrotic trabeculae and graft-host junctions by a predominantly osteoblastic reaction at the periphery of all cylinders, while in HA- and TCP-grains early bone formation in the center of drill holes is detectable as well. There is a direct contact between HA-/TCP-particles and newly formed bone without fibrous tissue formation at the implant surfaces. Central new bone formation in rabbit allografts can be observed after 6 to 8 weeks together with a secondary osteoclastic resorption of necrotic transplant trabeculae. The result of this remodeling process is a complete degradation of transplant cylinders with reorganization of vital trabeculae oriented in a mature pattern after 12 to 26 weeks. In contrast the HA- and TCP-implants did not show any signs of resorption.

Fat in bone xenografts: Importance of the purification procedures on cleanliness, wettability and biocompatibility

Xenografting is a promising alternative to allografts and autografts. The remaining lipids in bone are known to influence the biocompatibility. A comparative study of wettability was done on standardized blocks of two biomaterials. A highly purified and defatted bovine bone graft (T650) was found to retain more water (2.06 g/block) than a less defatted biomaterial (T360, 0.3 g/block). Wettability, observed in the laboratory, may reflect an important in vivo property: the rapidity for extracellular fluids and blood cells to invade the graft and carry bone forming cells. When implanted in rabbit cancellous bone, T650 appears to be osteoconductive in a manner that allows trabecular architecture to be restored within 6 months.

In vivo Chondrogenesis and Histochemical Appearance of Dermatan Sulphate in Rabbit Cancellous Bone

In vivo Chondrogenesis and Histochemical Appearance of Dermatan Sulphate in Rabbit Cancellous Bone