Biodegradable and bioactive properties of a novel bone scaffold coated with nanocrystalline bioactive glass for bone tissue engineering

The development of the new technologies of bone tissue engineering requires the production of bioactive and biodegradable macroporous scaffolds. Hydroxyapatite (HA) ceramics are useful bone substitutes, but they degrade minimally. Tricalcium phosphates also show poor ability of Ca-P formation both in-vitro and in-vivo, although they are degradable. The present study introduces a biodegradable, bioactive, and macroporous scaffold with suitable mechanical properties. The prepared hydroxyapatite scaffold was coated with a nanocrystalline bioactive glass layer to be subsequently sintered at different temperatures. The bioactivity and degradability of the coated scaffolds were investigated by standard procedures. The ability to induce Ca-P formation in SBF (simulated body fluid) was also investigated semi-quantitatively. BS1 scaffolds (scaffolds sintered at 800 °C with a holding time of 2 h) showed remarkable bioactivity and degradability simultaneously. Formation of a nanocrystalline phase (Si₂PO₇) during the sintering considerably decreased the capability of BS1 scaffolds for Ca-P formation and the rate of degradation but enhanced their mechanical properties. The BS1 scaffolds showed not only significant bioactivity but also good degradability and suitable mechanical property.     

A novel sheep vertebral bone defect model for injectable bioactive vertebral augmentation materials

New injectable bone substitutes have been developed that are, unlike polymethylmethacrylate, biologically active and have an osteogenic effect leading to osteogenesis and bone remodeling for vertebroplasty or kyphoplasty. In this study, we developed a sheep vertebral bone defect model to evaluate the new bioactive materials and assessed the feasibility of the model in vivo. Bone voids were experimentally created on lumbar vertebrae L2–L5 with L1 and L6 left intact as a normal control in mature sheep. The defect vertebrae L2–L5 in each sheep were randomized to receive augmentation with calcium phosphate cement (CPC) or sham. Vertebrae (L1–L6) were collected after 2 and 24 weeks of the cement augmentation and their strength and stiffness, as well as osseointegration activity and biodegradability, were evaluated. Finally, CPC significantly improved the strength and stiffness of vertebrae but did not yet restore it to the normal level at 24 weeks. Osteogenesis occurred at a substantially high level after 24 weeks of CPC augmentation or sham. Therefore, the sheep vertebral model with one void, 6.0 mm in diameter and 15.0 mm in depth, is replicable and can be used for evaluating the new injectable bioactive materials in vertebral augmentation or reconstruction.     

In vitro and in vivo biological responses to a novel radiopacifying agent for bone cement.

Iodixanol (IDX) and iohexol (IHX) have been investigated as possible radiopacification agents for polymethylmethacrylate (PMMA) bone cement, to replace the currently used barium sulphate and zirconia. IDX and IHX are both water-soluble iodine-based contrast media and for the last 20 years have been used extensively in clinical diagnostic procedures such as contrast media enhanced computed tomography, angiography and urography. One of the major reasons to remove the current radiopacifying agents is their well-documented cytotoxicity and their potential to increase bone resorption. Using in vitro bone resorption assays, the effect of PMMA particles plus IDX or IHX to induce osteoclast formation and lacunar resorption on dentine slices has been investigated. These responses have been compared with the in vitro response to PMMA particles containing the conventional radiopacifying agents, that is, barium sulphate and zirconia. In parallel, the in vivo reaction, in terms of new bone formation, to particles of these materials has been tested using a bone harvest chamber in rabbit tibiae. In vitro cell culture showed that PMMA containing IHX resulted in significantly less bone resorption than PMMA containing the conventional opacifiers. In vivo testing, however, showed no significant differences between the amounts of new bone formed around cement samples containing the two iodine-based opacifying agents in particulate form, although both led to fewer inflammatory cells than particles of PMMA containing zirconia. Our results suggest that a non-ionic radiopacifier could be considered as an alternative to the conventional radiopacifying agents used in biomaterials in orthopaedic surgery.     

Preparation and characterization of a novel bioactive bone cement: Glass based nanoscale hydroxyapatite bone cement

A novel type of glass-based nanoscale hydorxypatite (HAP) bioactive bone cement (designed as GBNHAPC) was synthesized by adding nanoscale hydroxyapatite (HAP) crystalline (20–40 nm), into the self-setting glass-based bone cement (GBC). The inhibition rate of nanoscale HAP and micron HAP on osteosarcoma U2-OS cells was examined. The effects of nanoscale HAP on the crystal phase, microstructure and compressive strength of GBNHAPC were studied respectively. It was concluded that nanoscale HAP could inhibit the cell proliferation, while micron HAP could not, and that nanoscale HAP could be dispersed in the cement evenly and the morphology did not change significantly after a longer immersion time. XRD and FTIR results show nanoscale HAP did not affect the setting reaction of the cement. Furthermore, GBNHAPC had a higher compressive strength (92 MPa) than GBC. It was believed that GBNHAPC might be a desirable biomaterial that could not only fill bone defects but also inhibit cancer cell growth.     

Hydrophilic matrices to be used as bioactive and degradable bone cements

Two different hydrophilic systems were investigated regarding their suitability to be used as enzymatically degradable and highly bioactive bone cements. They contained either acrylic acid (AA) or 2-hydroxyethyl methacrylate (HEMA) as the hydrophilic monomer. Swelling, degradation, mechanical and bioactivity tests were employed to characterise their behaviour. Although both of the systems were very hydrophilic, only the one containing HEMA was able to form an apatite-like layer on its surface. Moreover, this system could be degraded by amylolytic enzymes at a rate easily controlled by the incorporation of different amounts of enzyme to the formulation, as shown by the evolution of the mechanical properties, weight loss and glucose concentration in the solution. These results show these novel systems have a great potential to induce bone ingrowth inside the pores created during the degradation of the material, therefore establishing a strong interface with the tissue.     

A novel injectable borate bioactive glass cement for local delivery of vancomycin to cure osteomyelitis and regenerate bone

Osteomyelitis (bone infection) is often difficult to cure. The commonly-used treatment of surgical debridement to remove the infected bone combined with prolonged systemic and local antibiotic treatment has limitations. In the present study, an injectable borate bioactive glass cement was developed as a carrier for the antibiotic vancomycin, characterized in vitro, and evaluated for its capacity to cure osteomyelitis in a rabbit tibial model. The cement (initial setting time = 5.8 ± 0.6 min; compressive strength = 25.6 ± 0.3 MPa) released vancomycin over ~25 days in phosphate-buffered saline, during which time the borate glass converted to hydroxyapatite (HA). When implanted in rabbit tibial defects infected with methicillin-resistant Staphylococcus aureus (MRSA)-induced osteomyelitis, the vancomycin-loaded cement converted to HA and supported new bone formation in the defects within 8 weeks. Osteomyelitis was cured in 87 % of the defects implanted with the vancomycin-loaded borate glass cement, compared to 71 % for the defects implanted with vancomycin-loaded calcium sulfate cement. The injectable borate bioactive glass cement developed in this study is a promising treatment for curing osteomyelitis and for regenerating bone in the defects following cure of the infection.     

On the production of novel zirconia-reinforced bioactive glass porous structures for bone repair

Journal of Materials Science - The objective of this study was to develop a replica method for producing zirconia-reinforced bioactive glass (ZRBG) porous structures for bone...     

羟磷灰石骨水泥载体缓释生物活性因子的有效性研究

探讨羟磷灰石骨水泥(HAC)作为生物活性因子缓释载体的有效性。实验组分为(1)局部用药组:HAC中加入去甲万古霉素,植入家兔胫骨上端;(2)全身用药对照组:通过耳缘静脉注射去甲万古霉素,植入家兔胫骨的 HAC 中不加药;术后分时段对动物血药浓度和骨药浓度测定。(3)BMP组:HAC中加入 rh BMP 2后置入家兔背阔肌肌袋,术后 4 周观察异位诱导成骨效果。实验结果显示局部用药组各时段的血药浓度均低于全身用药对照组,而局部用药组的骨药浓度显著超过全身用药组,即使到第 2 周其骨药浓度仍有3.96μg/mg。HAC/rhBMP 2复合体植入家兔背阔肌肌袋后第4周即可观察到复合体表面形成包绕的新生骨质。这表明 HAC作为药物和 BMP 缓释载体显示出良好的缓释特性。     

A novel photoreactive amphiphile of nitrophenylazide for immobilization of bioactive proteins

Monolayers of an amphiphilic nitrophenylazide (ANPA) derivative were prepared and their ability to immobilize a protein molecule by a photochemical binding reaction was demonstrated as a novel means to constitute a biofunctional membrane. Molecular orientations and photoreactions in the ANPA monolayers were investigated by Fourier transform IR spectroscopy. Rapid photolysis of ANPA was observed which reflects decomposition of the azide group to a reactive nitrene radical intermediate. As an enzyme, glucose oxidase (GOD) was immobilized on a monomolecular film of ANPA, coated on a substrate, by means of enzyme adsorption at the solid-solution interface and subsequent photolysis of ANPA. Radioisotope labelling experiments revealed that the enzyme was immobilized at a high surface concentration which corresponds to a closely packed monolayer of GOD. The specific activity of the immobilized enzyme was estimated to be high compared with those of other systems involving the adsorption of this enzyme. The usefulness of the present technique for fabricating biofunctional monolayer assemblies is discussed.     

Nanoporous,bioactive and cytocompatible TiO2 encapsulated Ti particles as bone augmentation material

Bone impaction grafting is a surgical technique used for the restoration of bone stock loss with impaction of autograft or allograft bone particles. Porous Ti particles are deformable, like bone particles, and offer better primary stability. In this study, spherical Ti particles were subjected to H₂O₂ solution treatment at 70 °C for 3 h and heat treated at different temperatures in the range of 400–800 °C. FE-SEM observation showed that Ti particle form highly porous network structure and these porous network structures were confirmed to be hydrogen titanate by Raman analysis. Subsequent heat treatment at temperature ranges of 400–800 °C showed the gradual transformation of hydrogen titanate network to anatase and finally rutile phase of TiO₂. The network structure appeared to be compacted by the heat treatment due to water removal and ultimately take the particulate morphology above 800 °C. Thus formed TiO₂ encapsulated Ti particles showed bioactivity in terms of deposition of apatite layer from simulated body fluid in the range of 400–600 °C. The cytocompatibility studies using osteoblast-like cells, MG63 showed good cell viability as well as adhesion for all Ti particles. Present results indicates that bioactive TiO₂ encapsulated Ti particles could be a candidate material to be useful as bone or dental cavity filler or bone cement for total hip replacement materials.     

In vitro degradation of novel bioactive polycaprolactone—20% tricalcium phosphate composite scaffolds for bone engineering

Our group recently fabricated novel 3D polycaprolactone—20% tricalcium phosphate (PCL-TCP) composite scaffolds for applications in bone engineering. The bioactivity of such synthetic biomaterials can be evaluated by examining its ability to initiate the formation of apatite on its surface when immersed in simulated body fluids (SBF). In this study, the in vitro degradation behaviors of these scaffolds were systematically monitored for varying time periods of 1, 7, 14, 21 and 28 days post-immersion in SBF at 37 °C. Weight loss and water absorption of the samples indicated that PCL-TCP scaffolds were only slowly degraded. Biochemical assays and pH measurements revealed that hydroxyapatite, the main inorganic constituent of bone, commenced to form on the surface of the scaffolds after 17 days of immersion in SBF. Von Kossa assays demonstrated that calcium deposits increased progressively on the surface of the scaffolds after soaking in SBF for 2 weeks. Scanning electron microscopy verified the surface crystallization of the apatite layer formed over the entire period of time. In conclusion, the synergy of PCL with TCP in a composite scaffold confers both bioresorbability as well as bioactivity that offer an exciting approach for bone regeneration purposes.     

Effect of inorganic and organic bioactive signals decoration on the biological performance of chitosan scaffolds for bone tissue engineering

The present work is focused on the design of a bioactive chitosan-based scaffold functionalized with organic and inorganic signals to provide the biochemical cues for promoting stem cell osteogenic commitment. The first approach is based on the use of a sequence of 20 amino acids corresponding to a 68–87 sequence in knuckle epitope of BMP-2 that was coupled covalently to the carboxyl group of chitosan scaffold. Meanwhile, the second approach is based on the biomimetic treatment, which allows the formation of hydroxyapatite nuclei on the scaffold surface. Both scaffolds bioactivated with organic and inorganic signals induce higher expression of an early marker of osteogenic differentiation (ALP) than the neat scaffolds after 3 days of cell culture. However, scaffolds decorated with BMP-mimicking peptide show higher values of ALP than the biomineralized one. Nevertheless, the biomineralized scaffolds showed better cellular behaviour than neat scaffolds, demonstrating the good effect of hydroxyapatite deposits on hMSC osteogenic differentiation. At long incubation time no significant difference among the biomineralized and BMP-activated scaffolds was observed. Furthermore, the highest level of Osteocalcin expression (OCN) was observed for scaffold with BMP2 mimic-peptide at day 21. The overall results showed that the presence of bioactive signals on the scaffold surface allows an osteoinductive effect on hMSC in a basal medium, making the modified chitosan scaffolds a promising candidate for bone tissue regeneration.     

A novel bio-inorganic bone implant containing deglued bone, chitosan and gelatin

With the aim of developing an ideal bone graft, a new bone grafting material was developed using deglued bone, chitosan and gelatin. Deglued bone (DGB) which is a by-product of bone glue industries and has the close crystallographic similarities of hydroxyapatite was used as main component in the preparation of bone implant. Chitosan was prepared from the exoskeleton of prawn (Pinaeus indicus, family Crustaceae) which is a by-product of seafood industries. Chitosan gives toughness to the product and do not allow the DGB particles to wither away when the implant is placed in the defect. Gelatin was used as binder for the preparation of DGB-chitosan composite. The DGB, chitosan and DGB-chitosan-gelatin composite, which were prepared in the laboratory, were analysed for their physicochemical properties by infrared spectroscopy, X-ray diffraction and scanning electron microscopy studies.     

Clustered DNA damages as dosemeters for ionising radiation exposure and biological responses.

Clustered DNA damages--two or more lesions (oxidised bases. abasic sites, or strand breaks) within a few DNA helical turns on opposing strands--are induced in DNA in solution and in vivo in human cells by ionising radiation. They have been postulated to be difficult to repair, and thus of potentially high biological significance. Since the total of clustered damages produced by ionising radiation is at about 3 to 4 times higher levels than double-strand breaks and are apparently absent in unirradiated cells, levels of clustered damages present immediately alter radiation exposure could serve as sensitive dosemeters of radiation exposure. Since some clusters may not be repairable and may accumulate in cells, they might also be useful as integrating dosemeters of biological effects of radiation damage.     

Lithium fluoride as a novel X-ray image detector for biological mu-world capture

X-ray microradiographs of small biological objects, such as animals and plant materials at micrometric resolution, are currently performed by various methods, all of which are limited by the resolution or the dynamic range of the image detectors. Here a novel X-ray image detector is discussed, in which the previous limitations have been overcome. A film of lithium fluoride salt is used as a detector, in which the stored biological image is read by observing the optically stimulated visible luminescence of the active color centers, efficiently produced by the X-rays.     

Poly (l-lactide-co-Є caprolactone) microspheres laden with bioactive glass-ceramic and alendronate sodium as bone regenerative scaffolds

Microspheric scaffolds of poly-(lactide-co-caprolactone) loaded with alendronate sodium, a family precursor of bisphosphonate drug and bioactive glass-ceramic (BGS) were prepared for the treatment of osteoporosis like bone defects with the rationale of getting a combined effect/concurrent advantage of osteoclast apoptosis as well as the augmentation of bone regeneration. The porous microspheres were generated by oil in water/solvent evaporation technique. The distribution of bioactive glass-ceramic was evidenced by the microcomputed tomography (μ-CT) and scanning electron microscopy analyses. The microspheres were evaluated for their in vitro cytocompatibility using L929 cell line and were found to be noncytotoxic. The osteoinductivity of the scaffold was assessed by its response in simulated body fluid and observed an excellent hydroxy carbonate apatite (HCA) layer formation on the surface which revealed the bone bonding and bone regeneration capability of the scaffold. The cell adhesion studies was performed with L-929 cell line and a marking cell growth on the surface as well as in the pores of the bioactive glass-ceramic as well as bioactive glass-ceramic cum drug incorporated microspheres was evidenced by the Confocal laser scanning microscopy (CLSM) investigation. No cell adhesion was observed onto the surface of the bare microspheres prepared by the copolymer alone where as the bioactive glass-ceramic and drug cum bioactive glass-ceramic loaded microspheres were found to promote the cell adhesion. The viability of the adhered cells on the microspheres was checked by flourescein diacetate (FDA) staining and it was observed that the adhered cells were viable and metabolically active. The release of the drug, alendronate sodium, directly into the problem site makes the presently prepared microsphere superior to the oral variety of drug available which is associated with oral discomfort and low bioavailability.     

Development of degradable and bioactive composite as bone implants by incorporation of mesoporous bioglass into poly(l-lactide)

Bioactive composites containing mesoporous bioglass (MBG) and poly(l-lactide) (PLLA) for bone regeneration were fabricated by solution casting method. The results showed that the compressive strength and hydrophilicity of the MBG/PLLA composites significantly improved with the increase of MBG content. In addition, the weight loss ratio of the composites in Tris–HCl solution was obviously enhanced with the increase of MBG content. Moreover, the composite containing MBG could compensate for the decrease of pH value by neutralizing the acidic products from PLLA degradation in the Tris–HCl solution. Furthermore, the MBG/PLLA composites could induce apatite formation on their surfaces after soaked into simulated body fluid (SBF), indicating good bioactivity. In cell culture experiments, the results showed that the composite could enhance cell attachment, proliferation and alkaline phosphatase activity (ALP) of MC3T3-E1 cells, and the improvements were dependent on the MBG content in the composites. In short, the MBG/PLLA biocomposites with improved properties of hydrophilicity, degradability, bioactivity, neutralizing acidic degradable products and good cytocompatibility would be a promising orthopedic implant material for bone repair application.