Homogeneous chitosan/carbonate apatite/citric acid nanocomposites prepared through a novel in situ precipitation method

Homogeneous nanocomposites composed of carbonate apatite and chitosan in the presence of citric acid were synthesized by a novel in situ precipitation method. The morphological and componential properties of composites were investigated. The carbonate apatite particulates, in sizes of about 50–100 nm, were distributed within the network chitosan hydrogel homogeneously, moreover, inorganic particles could be controlled by the size of networks of organic matrix which was mediated with crosslink degree of chitosan. Through highly magnified TEM observation, it can be shown that inorganic particles were composed of more fine sub-particles whose diameters were between 2 and 5 nm in size without regular crystallographic orientation. The concept of multiple-order template mediation was brought forward for the first time. A novel hierarchical porous nanocomposite scaffold was also prepared by multilevel freeze-drying technique and its mechanical performance had obvious increase compared with neat chitosan. These results provided an efficient approach toward new biomimetic tissue scaffold for the biomedical applications with enhanced intensity/bioactivity and controlled resorption rates.     

Novel biomimetic hydroxyapatite/alginate nanocomposite fibrous scaffolds for bone tissue regeneration

Hydroxyapatite/alginate nanocomposite fibrous scaffolds were fabricated via electrospinning and a novel in situ synthesis of hydroxyapatite (HAp) that mimics mineralized collagen fibrils in bone tissue. Poorly crystalline HAp nanocrystals, as confirmed by X-ray diffractometer peak approximately at 2θ = 32° and Fourier transform infrared spectroscopy spectrum with double split bands of PO₄(v ₄) at 564 and 602 cm^?1, were induced to nucleate and grow at the [–COO^?]–Ca²⁺–[–COO^?] linkage sites on electrospun alginate nanofibers impregnated with PO₄ ^3? ions. This novel process resulted in a uniform deposition of HAp nanocrystals on the nanofibers, overcoming the severe agglomeration of HAp nanoparticles processed by the conventional mechanical blending/electrospinning method. Preliminary in vitro cell study showed that rat calvarial osteoblasts attached more stably on the surface of the HAp/alginate scaffolds than on the pure alginate scaffold. In general, the osteoblasts were stretched and elongated into a spindle-shape on the HAp/alginate scaffolds, whereas the cells had a round-shaped morphology on the alginate scaffold. The unique nanofibrous topography combined with the hybridization of HAp and alginate can be advantageous in bone tissue regenerative medicine applications.     

Synthesis of mesoporous structured hydroxyapatite particles using yeast cells as the template

In this study, hydroxyapatite (HAp) particles with mesoporous structure have been synthesized from calcium hydroxide and di-ammonium hydrogen phosphate using yeast cells as the template. The characterization methods such as X-ray diffraction (XRD), Fourier transform infrared spectrograph (FTIR), N₂ adsorption–desorption isotherms (NADI), transmission electron microscopy (TEM) and field emission scanning electron microscopy (FESEM) were used for determination of the particles structure (particle size, structural evolution and morphology). The results show that HAp particles with mesoporous structure could be produced. The size of HAp particles was approximately hundreds of nanometer. The pore width of HAp particles was in the range of 2.0–40 nm and the maximum centered around 4.5 nm.     

Development of nano-sized hydroxyapatite reinforced composites for tissue engineering scaffolds

Nano-sized hydroxyapatite (nanoHA) reinforced composites, mimicking natural bone, were produced. Examination by transmission electron microscopy revealed that the nanoHA particles had a rod-like morphology, 20–30 nm in width and 50–80 nm in length. The phase composition of hydroxyapatite was confirmed by X-ray diffraction. The nanoHA particles were incorporated into poly-2-hydroxyethylmethacrylate (PHEMA)/polycaprolactone (PCL) matrix to make new nanocomposites: nanoHA-PHEMA/PCL. Porous nanocomposite scaffolds were then produced using a porogen leaching method. The interconnectivity of the porous structure of the scaffolds was revealed by non-destructive X-ray microtomography. Porosity of 84% was achieved and pore sizes were approximately around 300–400 μm. An in vitro study found that the nanocomposites were bioactive as indicated by the formation of a bone-like apatite layer after immersion in simulated body fluid. Furthermore, the nanocomposites were able to support the growth and proliferation of primary human osteoblast (HOB) cells. HOB cells developed a well organized actin cytoskeletal protein on the nanocomposite surface. The results demonstrate the potential of the nanocomposite scaffolds for tissue engineering applications for bone repair.     

Poly(lactide-co-glycolide)/hydroxyapatite nanofibrous scaffolds fabricated by electrospinning for bone tissue engineering

Poly(lactide-co-glycolide) (PLGA) nanofibrous composite scaffolds having nano-hydroxyapatite particles (HAp) in the fibers were prepared by electrospinning of PLGA and HAp with an average diameter of 266.6 ± 7.3 nm. Microscopy and spectroscopy characterizations confirmed integration of the crystalline HAp in the scaffolds. Agglomerates gradually appeared and increased on the fiber surface along with increase of the HAp concentration. In vitro mineralization in a 5 × simulated body fluid (SBF) revealed that the PLGA/HAp nanofibrous scaffolds had a stronger biomineralization ability than the control PLGA scaffolds. Biological performance of the nanofibrous scaffolds of the control PLGA and PLGA with 5 wt% HAp (PLGA/5HAp) was assessed by in vitro culture of neonatal mouse calvaria-derived MC3T3-E1 osteoblasts. Both types of the scaffolds could support cell proliferation and showed sharp increase of viability until 7 days, but the cells cultured on the PLGA/5HAp nanofibers showed a more spreading morphology. Despite the similar level of the cell viability and cell number at each time interval, the alkaline phosphatase secretion was significantly enhanced on the PLGA/5HAp scaffolds, indicating the higher bioactivity of the as-prepared nano-HAp and the success of the present method for preparing biomimetic scaffold for bone regeneration.     

A novel approach of homogenous inorganic/organic composites through in situ precipitation in poly-acrylic acid gel

A new in situ precipitation technique was developed to promote high-affinity nucleation and growth of calcium phosphate in the polymer hydrogel. Poly-acrylic acid/ hydroxyapatite (HAP) composites have been prepared using template-driven reaction. Nano-sized hydroxyapatite particles were distributed within organic template homogenously, furthermore, inorganic particles were fine and uniform. A kind of specific product with the characteristic of fractal was also obtained. During the composite process, 3D network of organic matrices played an important role in the superfine interaction of HAP and hydrogel through the compartment-effect of interior of hydrogel. This method provides an efficient approach toward inorganic/organic nanocomposites with high-uniformity decentralization for biomimetic replant applications. This paper discussed the mechanism of compartment-effect, and the concept of in situ precipitation in gel was brought forward. SEM and TEM were employed in the analysis of the morphological characteristic of uniform inorganic/organic composites and inorganic minerals separated from organic template. SEM-associated EDS area analysis and TEM-associated EDS analysis were employed in the analysis of component characteristic of inorganic/organic composite and inorganic particles. X-ray diffraction was employed in the analysis of precipitation phase crystal structure of inorganic component.     

Development of porous HAp and β-TCP scaffolds by starch consolidation with foaming method and drug-chitosan bilayered scaffold based drug delivery system

The inability to maintain high concentrations of antibiotic at the site of infection for an extended period of time along with dead space management is still the driving challenge in treatment of osteomyelitis. Porous bioactive ceramics such as hydroxyapatite (HAp) and beta-tri calcium phosphate (β-TCP) were some of the alternatives to be used as local drug delivery system. However, high porosity and high interconnectivity of pores in the scaffolds play a pivotal role in the drug release and bone resorption. Ceftriaxone is a cephalosporin that has lost its clinical popularity. But has recently been reported to exhibit better bactericidal activity in vitro and reduced probability of resistance development, in combination with sulbactam, a β-lactamase inhibitor. In this article, a novel approach of forming HAp and pure β-TCP based porous scaffolds by applying together starch consolidation with foaming method was used. For the purpose, pure HAp and β-TCP were prepared in the laboratory and after thorough characterization (including XRD, FTIR, particle size distribution, etc.) the powders were used for scaffold fabrication. The ability of these scaffolds to release drugs suitably for osteomyelitis was studied in vitro. The results of the study indicated that HAp exhibited better drug release profile than β-TCP when drug was used alone indicating the high influence of the carrier material. However, this restriction got relaxed when a bilayered scaffold was formed using chitosan along with the drug. SEM studies along with EDAX on the drug-chitosan bilayered scaffold showed closest apposition of this combination to the calcium phosphate surface.     

Glass–ceramic scaffolds containing silica mesophases for bone grafting and drug delivery

Glass–ceramic macroporous scaffolds were prepared using glass powders and polyethylene (PE) particles of two different sizes. The starting glass, named as Fa-GC, belongs to the system SiO₂–P₂O₅–CaO–MgO–Na₂O–K₂O–CaF₂ and was synthesized by a traditional melting-quenching route. The glass was ground and sieved to obtain powders of specific size which were mixed with PE particles and then uniaxially pressed in order to obtain crack-free green samples. The compact of powders underwent a thermal treatment to remove the organic phase and to sinter the Fa-GC powders. Fa-GC scaffolds were characterized by means of X-Ray Diffraction, morphological observations, density measurements, image analysis, mechanical tests and in vitro tests. Composite systems were then prepared combining the drug uptake-delivery properties of MCM-41 silica micro/nanospheres with the Fa-GC scaffold. The system was prepared by soaking the scaffold into the MCM-41 synthesis batch. The composite scaffolds were characterized by means of X-Ray Diffraction, morphological observations, mechanical tests and in vitro tests. Ibuprofen was used as model drug for the uptake and delivery analysis of the composite system. In comparison with the MCM-41-free scaffold, both the adsorption capacity and the drug delivery behaviour were deeply affected by the presence of MCM-41 spheres inside the scaffold.     

Studies on the preparation and structure of polyacrylamide/α-zirconium phosphate nanocomposites

In this paper, a novel polyacrylamide(PAM)/α-zirconium phosphate(α-ZrP) nanocomposite was successfully synthesized by exfoliation-adsorption and in-situ intercalative polymerization. The microstructure of PAM/α-ZrP nanocomposites was confirmed by X-ray diffraction measurement, transmission electron microscopy (TEM), high resolution electron microscopy (HRTEM). The results suggested that the α-ZrP lamellae were dispersed well in PAM matrix, which indicated the formation of the exfoliated nanocomposites in the low inorganic loading of α-ZrP (≤5 wt%). With the increase of the inorganic loadings, the intercalated structure of PAM/α-ZrP nanocomposites was dominant with the d-spacings of about 1.50–1.58 nm corresponding to the inorganic loadings in the range of 10–20 wt%. Moreover, besides the electrostatic adsorption, it was also found that there may be some weak effect such as hydrogen bonding or protonation between the host and guest investigated using fourier transform infrared spectroscopy (FT-IR) and thermogravimetric (TG )analysis, which resulted in the enhancement of the thermal properties on the decomposition process of PAM/α-ZrP nanocomposites by the retardant effect of the exfoliated or intercalated α-ZrP nanometer lamellae.     

The osteogenic differentiation of dog bone marrow mesenchymal stem cells in a thermo-sensitive injectable chitosan/collagen/β-glycerophosphate hydrogel: in vitro and in vivo

Type I collagen was added to the composite chitosan solution in a ratio of 1:2 to build a physical cross-linked self-forming chitosan/collagen/β-GP hydrogel. Osteogenic properties of this novel injectable hydrogel were evaluated. Gelation time was about 8 min which offered enough time for handling a mixture containing cells and the subsequent injection. Scanning electronic microscopy (SEM) observations indicated good spreading of bone marrow mesenchymal stem cells (BMSCs) in this hydrogel scaffold. Mineral nodules were found in the dog-BMSCs inoculated hydrogel by SEM after 28 days. After subcutaneous injection into nude mouse dorsum for 4 weeks, partial bone formation was observed in the chitosan/collagen/β-GP hydrogel loaded with pre-osteodifferentiated dog-BMSCs, which indicated that chitosan/collagen/β-GP hydrogel composite could induce osteodifferentiation in BMSCs without exposure to a continual supply of external osteogenic factors. In conclusion, the novel chitosan/collagen/β-GP hydrogel composite should prove useful as a bone regeneration scaffold.     

Surface modification of poly(l-lactide) electrospun fibers with nanocrystal hydroxyapatite for engineered scaffold applications

The hydrophobicity of the poly(l-lactide) (PLLA) surface was modified by incorporating hydroxyapatite (HAp) nanocrystalline particles during the electrospinning process for the engineered scaffold applications. The HAp nanocrystals were synthesized with 30 nm in diameter and 100–120 nm in length, which subsequently formed micrometer-sized agglomerates in the range of 2.5 μm. The synthesized HAp agglomerates were electrospun in the PLLA solution, and the HAp nanocrystals were desirably exposed on the surface of the electrospun PLLA fibers to give higher surface energy and lower contact angles with water. The surface-exposed hydrophilic HAp nanocrystals substantially increased the precipitation of various salts on the HAp/PLLA fiber surfaces in a buffer solution due to the hydrophilic nature and ionic affinity of HAp. Finally, the developed HAp/PLLA fibers desirably sustained the fibrous structural integrity during the accelerated-aging test in water, which was not the case with the pristine PLLA fibers.     

醇化改性蓖麻油基聚氨酯/n-HA复合支架材料的结构及力学性能

有机/无机材料的两相界面作用和均匀复合是影响复合支架材料性能的主要因素。本研究通过工艺改进, 对聚氨酯软段成分进行改性得到醇化蓖麻油, 经原位聚合发泡制备出多孔复合支架, 较好地实现了纳米羟基磷灰石(n-HA)颗粒的均匀分散。结果表明, 改性聚氨酯基体有效增加了羟基值, 其与极性n-HA两相界面复合良好, 无明显界面分相和颗粒团聚发生。支架孔径分布均匀, 但支架材料的孔隙率、孔径大小和结晶度略有减小。红外光谱和X射线衍射分析表明, n-HA和醇化蓖麻油基聚氨酯基体的分子间存在丰富的氢键和化学键, 促进了无机-有机相的相容性和稳定性。醇化改性和纳米无机粒子添加对材料性能的协同作用有效改善了支架的力学性能, 复合支架的压缩强度和模量均大幅增长。该种n-HA/聚氨酯复合支架有望用于进一步的骨再生和骨组织工程研究。     

Mineralization of osteoblasts with electrospun collagen/hydroxyapatite nanofibers

Regeneration of fractured or diseased bones is the challenge faced by current technologies in tissue engineering. The major solid components of human bone consist of collagen and hydroxyapatite. Collagen (Col) and hydroxyapatite (HA) have potential in mimicking natural extracellular matrix and replacing diseased skeletal bones. More attention has been focused on HA because of its crystallographic structure similar to inorganic compound found in natural bone and extensively investigated due to its excellent biocompatibility, bioactivity and osteoconductivity properties. In the present study, electrospun nanofibrous scaffolds are fabricated with collagen (80 mg/ml) and Col/HA (1:1). The diameter of the collagen nanofibers is around 265 ± 0.64 nm and Col/HA nanofibers are 293 ± 1.45 nm. The crystalline HA (29 ± 7.5 nm) loaded into the collagen nanofibers are embedded within nanofibrous matrix of the scaffolds. Osteoblasts cultured on both scaffolds and show insignificant level of proliferation but mineralization was significantly (p < 0.001) increased to 56% in Col/HA nanofibrous scaffolds compared to collagen. Energy dispersive X-ray analysis (EDX) spectroscopy results proved the presence of higher level of calcium and phosphorous in Col/HA nanocomposites than collagen nanofibrous scaffolds grown osteoblasts. The results of the present study suggested that the designed electrospun nanofibrous scaffold (Col/HA) have potential biomaterial for bone tissue engineering.     

Construction of collagen II/hyaluronate/chondroitin-6-sulfate tri-copolymer scaffold for nucleus pulposus tissue engineering and preliminary analysis of its physico-chemical properties and biocompatibility

To construct a novel scaffold for nucleus pulposus (NP) tissue engineering, The porous type II collagen (CII)/hyaluronate (HyA)–chondroitin-6-sulfate (6-CS) scaffold was prepared using 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC) and N-hydroxysuccinimide (NHS) cross-linking system. The physico-chemical properties and biocompatibility of CII/HyA–CS scaffolds were evaluated. The results suggested CII/HyA–CS scaffolds have a highly porous structure (porosity: 94.8 ± 1.5%), high water-binding capacity (79.2 ± 2.8%) and significantly improved mechanical stability by EDC/NHS crosslinking (denaturation temperature: 74.6 ± 1.8 and 58.1 ± 2.6°C, respectively, for the crosslinked scaffolds and the non-crosslinked; collagenase degradation rate: 39.5 ± 3.4 and 63.5 ± 2.0%, respectively, for the crosslinked scaffolds and the non-crosslinked). The CII/HyA–CS scaffolds also showed satisfactory cytocompatibility and histocompatibility as well as low immunogenicity. These results indicate CII/HyA–CS scaffolds may be an alternative material for NP tissue engineering due to the similarity of its composition and physico-chemical properties to those of the extracellular matrices (ECM) of native NP.     

Polyampholytes superabsorbent nanocomposites with excellent gel strength

A series of novel polyampholyte superabsorbent nanocomposites with excellent gel strength were synthesized by in situ solution polymerization in aqueous solution. Acrylic acid and acryloyloxyethyl trimethyl ammonium chloride (DAC) were employed as ionic monomers and montmorillonite (MMT) was used as inorganic component. The addition of cationic component could supply the positive charge in the network of nanocomposite and promote the formation of nanostructure of composites due to the interaction between DAC and clay platelets. The performance of polyampholyte nanocomposites were investigated and the result showed that the gel strength of nanocomposite hydrogel in distilled water and 0.9 wt% NaCl solution could reach 198.85 and 204.23 mJ/g, respectively, which were 13 times of the gel strength of matrix. The investigation of swelling behaviors showed that the nanocomposites had particular swelling behaviors of polyampholytes hydrogel in solution with different pH values and concentration of NaCl.     

Preparation of high strength macroporous hydroxyapatite scaffold

Hydroxyapatite (HAp) powder was prepared from CaNO₃·4H₂O and (NH₄)₂HPO₄ by wet-chemical method and has phase stable up to 1250 °C. High strength macroporous HAp–naphthalene (HN) and HAp–naphthalene–benzene (HNB) scaffolds were fabricated by adapting sintering method. The resulting HAp scaffolds have porosity about 60 vol.% with compressive strength of ~ 11 MPa and average pore diameter in the range of ~ 125 μm. The incorporation of benzene in HN scaffold reduces the strength whereas enhanced both the porosity and pore size distribution. XRD, FTIR, SEM and mercury porosimeter techniques were used to study the phase purity, morphology, pore size and pore size distribution of scaffold. The study compared the effect of concentration of naphthalene on strength, porosity and pore size distribution on both HN and HNB scaffold. In-vitro bioactivity studies on HN and HNB scaffolds show the nucleation of spherical carbonated apatite particles on the surface in SBF solution.     

Biocompatible glass–ceramic materials for bone substitution

A new bioactive glass composition (CEL2) in the SiO₂–P₂O₅–CaO–MgO–K₂O–Na₂O system was tailored to control pH variations due to ion leaching phenomena when the glass is in contact with physiological fluids. CEL2 was prepared by a traditional melting-quenching process obtaining slices that were heat-treated to obtain a glass-ceramic material (CEL2GC) that was characterized thorough SEM analysis. Pre-treatment of CEL2GC with SBF was found to enhance its biocompatibility, as assessed by in vitro tests. CEL2 powder was then used to synthesize macroporous glass–ceramic scaffolds. To this end, CEL2 powders were mixed with polyethylene particles within the 300–600 μm size-range and then pressed to obtain crack-free compacted powders (green). This was heat-treated to remove the organic phase and to sinter the inorganic phase, leaving a porous structure. The biomaterial thus obtained was characterized by X-ray diffraction, SEM equipped with EDS, density measurement, image analysis, mechanical testing and in vitro evaluation, and found to be a glass–ceramic macroporous scaffold with uniformly distributed and highly interconnected porosity. The extent and size-range of the porosity can be tailored by varying the amount and size of the polyethylene particles.     

Preparation and characterization of hydroxyapatite/collagen nanocomposite gel

The self-organized hydroxyapatite/colagen (HAp/Col) nanocomposite fiber (79.6/20.4 weight ratio) was synthesized by a co-precipitation method using Ca(OH)2, H3PO4, and Col as starting substances. The gelation of the nanocomposite is essential in the application of the scaffold for bone tissue engineering. We successfully prepared HAp/Col nanocomposite gels by a facile novel method using a sodium phosphate buffer at pH 6.8. The water-insoluble nanocomposite was homogeneously dispersed in the buffer to form a viscous mixture, and gels were obtained after incubating of the mixture at 37 degrees C. The mechanical strength of the gels was analyzed against the incubation time. The demineralized gel with EDTA had the typical nanostructure of native type I Col fibers from the results of scanning electron microscopy (SEM) and atomic force microscopy (AFM); the dense network of type I Col nano-fibers below 100 nm in diameter, and the periodic pattern of 68.8+/-4.4 nm (mean +/- SD) along the fibers were observed. The gelation of the HAp/Col nanocomposite in the buffer is attributed to the physical cross-linking through entanglement of the reconstituted Col fibrils.     

Electrorheological properties of polypyrrole–SnO<Subscript>2</Subscript>–methylcellulose nanocomposite suspensions

Inorganic-conducting polymer particles were prepared to enhance physical and chemical properties by forming hybrid nanocomposites, which would improve the electrorheological (ER) effect of their suspensions. Polypyrrole (PPy)–SnO₂–methylcellulose nanocomposite particles were synthesized by controlling the ratio of pyrrole, SnO₂, and methylcellulose during the polymerization. The ER and dielectric properties of the PPy–SnO₂–methylcellulose nanocomposite suspensions were investigated. The ER response increases with the increase in the SnO₂/pyrrole ratio and also depends on the amount of methylcellulose amount during the polymerization, showing a maximum ER behavior.     

Aqueous ferrofluids as templates for magnetic hydroxyapatite nanocomposites

Poly (vinyl) alcohol stabilized aqueous ferrofluids (PVA-ff) were used as nanotemplates for the crystallization of calcium hydroxyapatite (HAp). Four sets of PVA-ff-HAp nanocomposites were synthesized using 20, 40, 60 and 80 ml of PVA-ff for the same initial constituents of HAp. Various physico-chemical analyses suggest that the HAp lattice structure accommodates PVA-ff to a certain extent, beyond which the magnetic intra-molecular interactions predominate and PVA-ff starts to be pushed out of the HAp matrix. The in situ incorporation of PVA-ff during HAp synthesis results in a novel magnetic biomaterial with potential applications as targeted delivery vehicles.