Investigations of gold nanoparticles-mediated carbon nanotube reinforced hydroxyapatite composite for bone regenerations

An excellent combination of biomaterials permits prompt features and high-throughput investigations in various fields, particularly in the biomedical applications. This article investigates the bone regeneration ability and compatibility of the Gold (Au) Nanoparticles (NPs) medicated carbon nanotube reinforced hydroxyapatite (HAP) composite. The morphologies of the synthesized Au NPs, HAP and HAP/CNT, and HAP/CNT-Au composites vary suggestively with modifying the components and the final composite showing as bone mimic extracellular matrix morphology. The structure, phase, and composition of the as-synthesized HAP were studied by FTIR, XRD, EDAX, and TEM techniques. The materials' biocompatibility was investigated in the Stimulated Body Fluid (SBF) solution, which resulted in the composite having good biocompatibility, bioactivity nature and hydroxyapatite layer formed on the composite surface. The composite shows good viability with Adipose Tissue-derived Stem Cells (ADSC) to cell growth and cell proliferation in the biological evaluation. It represented the composites having a good ability for cell formation development. Since the HAP/CNT-Au composite are useful in medicinal applications such as orthopedic and orthodontic repair/regenerations after the evaluations of animal and clinical investigations.     

Ultralong Hydroxyapatite Nanowire/Collagen Biopaper with High Flexibility,Improved Mechanical Properties and Excellent Cellular Attachment

Highly flexible hydroxyapatite/collagen (HAP/Col) composite membranes are regarded to be significant for guided bone regeneration application owing to their similar chemical composition to that of natural bone, excellent bioactivity and good osteoconductivity. However, the mechanical strength of the HAP/Col composite membranes is usually weak, which leads to difficult surgical operations and low mechanical stability during the bone healing process. Herein, highly flexible ultralong hydroxyapatite nanowires/collagen (UHANWs/Col) composite biopaper sheets with weight fractions of UHANWs ranging from 0 to 100 % are facilely synthesized. The UHANWs are able to weave with each other to construct a three‐dimensional fabric structure in the collagen matrix, providing a strong interaction between UHANWs and an intermolecular force between UHANWs and the collagen matrix. The as‐prepared UHANWs/Col composite biopaper exhibits improved mechanical properties and high flexibility. More importantly, the as‐prepared highly flexible 70 wt % UHANWs/Col composite biopaper exhibits an excellent cytocompatibility and outstanding cellular attachment performance as compared with the pure collagen and 70 wt % HAP nanorods/Col membranes. In consideration of its superior mechanical properties and outstanding cellular attachment performance, the as‐prepared UHANWs/Col composite biopaper is promising for applications in various biomedical fields such as guided bone regeneration.     

Nanostructured Materials for Skeletal Repair

The treatment of bone and cartilage defects with bioengineered constructs of artificial scaffolds and autogenous cells became the main challenge of contemporary regenerative medicine. Early defect repair may prevent secondary injury. Recent studies could prove that bone and cartilage cells are sensitive to microscale and nanoscale patterns of surface topography and chemical structure. Nanostructured materials provide an environment for tissue regeneration mimicking the physiological range of extracellular matrix. The article reviews several studies substantiating the superiority of nanostructured materials for bone and cartilage repair along with own results on cell attachment.     

Bioactive Phenylboronic Acid-Functionalized Hyaluronic Acid Hydrogels Induce Chondro-Aggregates and Promote Chondrocyte Phenotype

Hydrogels are extensively investigated as biomimetic extracellular matrix (ECM) scaffolds in tissue engineering. The physiological properties of ECM affect cellular behaviors, which is an inspiration for cell-based therapies. Photocurable hyaluronic acid (HA) hydrogel (AHAMA-PBA) modified with 3-aminophenylboronic acid, sodium periodate, and methacrylic anhydride simultaneously is constructed in this study. Chondrocytes are then cultured on the surface of the hydrogels to evaluate the effect of the physicochemical properties of the hydrogels on modulating cellular behaviors. Cell viability assays demonstrate that the hydrogel is non-toxic to chondrocytes. The existence of phenylboronic acid (PBA) moieties enhances the interaction of chondrocytes and hydrogel, promoting cell adhesion and aggregation through filopodia. RT-PCR indicates that the gene expression levels of type II collagen, Aggrecan, and Sox9 are significantly up-regulated in chondrocytes cultured on hydrogels. Moreover, the mechanical properties of the hydrogels have a significant effect on the cell phenotype, with soft gels (≈2 kPa) promoting chondrocytes to exhibit a hyaline phenotype. Overall, PBA-functionalized HA hydrogel with low stiffness exhibits the best effect on promoting the chondrocyte phenotype, which is a promising biomaterial for cartilage regeneration.     

In vitro investigation of cartilage regeneration properties of polymeric ceramic hybrid composite

There would be a major effect on the cartilage regeneration characteristics of ceramic material in a substrate implant requiring biologically active biomaterials and the reinforcement phase. At this moment, we produced collagen-hyaluronic acid @ hydroxyapatite-halloysite nanotube-single walled carbon nanotube composites, which is a successful technique for making a scaffold with a superior counter for cartilage property. FTIR, XRD, and SEM-EDAX were used to perform morphological and structural studies. The prepared composite's surface feature was investigated and discovered by HRTEM-SAED analysis, and it observed porous nature. The simulated body fluids (SBF) assessment of the materials was noticed their bioactivity and chondrocytes to determine their biocompatibility. Hybrid composite displayed promise for cartilage tissue engineering despite mesenchymal stem cells compatibility effect and magnificently demonstrated an antibacterial effect without antibiotics. The live/dead cells analysis shows that the composite can significantly improve mesenchymal stem cells, and the composite has the potential ability for cartilage regeneration. The above characteristics make the material quite interesting and important in the area for regenerative medicinal uses.     

Porous composite scaffolds of hydroxyapatite/silk fibroin via two‐step method

A two‐step method was used to fabricate the hydroxyapatite (HAP)/silk fibroin (SF) scaffolds, i.e. the nano‐sized HAP/SF composite powders were prepared by co‐precipitation, which were then blended with SF solution to fabricate the HAP/SF composite scaffolds. The obtained scaffolds showed a 3D porous structure. The porosity was higher than 90% with the average macropore size of 214.2 µm. Moreover, the nano‐sized HAP/SF composite powders were uniformly dispersed in the silk fibroin matrix, which provided the scaffolds enhanced compressive properties. The cell culture assay showed that the scaffolds fabricated by the two‐step method could improve the cell proliferation and osteogenic differentiation when compared with those prepared by the conventional one‐step blending method. The results suggested that the two‐step method could promote the uniform dispersion of HAP in the SF matrix and efficient combination between the HAP and the matrix, which may provide a potential application in the composite scaffold preparation for tissue engineering. Copyright © 2009 John Wiley & Sons, Ltd.     

Ultrasound augmenting injectable chemotaxis hydrogel for articular cartilage repair in osteoarthritis

The increasing incidence of osteoarthritis (OA) seriously affects life quality, posing a huge socioeconomic burden. Tissue engineering technology has become a hot topic in articular cartilage repair as one of the key treatment methods to alleviate OA. Hydrogel, one of the most commonly used scaffold materials, can provide a good extracellular matrix microenvironment for seed cells such as bone marrow mesenchymal stem cells (BMSCs), which can promote cartilage regeneration. However, the low homing rate of stem cells severely limits their role in promoting articular cartilage regeneration. Stromal cell-derived factor-1α (SDF-1α) plays a crucial role in the activation, mobilization, homing, and migration of MSCs. Herein, a novel injectable chemotaxis hydrogel, composed of chitosan-based injectable hydrogel and embedding SDF-1α-loaded nanodroplets (PFP@NDs-PEG-SDF-1α) was designed and fabricated. The ultrasound was then used to augment the injectable chemotaxis hydrogel and promote the homing migration of BMSCs for OA cartilage repair. The effect of ultrasound augmenting injectable PFP@NDs-PEG-SDF-1α/hydrogel on the migration of BMSCs was verified in vitro and in vivo, which remarkably promotes stem cell homing and the repair of cartilage in the OA model. Therefore, the treatment strategy of ultrasound augmenting injectable chemotaxis hydrogel has a bright potential for OA articular cartilage repair.     

Development of injectable high molecular weight hyaluronic acid hydrogels for cartilage regeneration

The study focuses on developing hyaluronic acid (1200 kilo Dalton) hydrogels for cartilage regeneration. In spite of being highly biocompatible; a large amount of water absorption and easily degrading nature restricts the use of hyaluronic acid in the field of tissue regeneration. This can be rectified by crosslinking hyaluronic acid with a crosslinking agent such as divinyl sulfone; which results in a biocompatible hydrogel with superior rheological properties. Different amounts of divinyl sulfone have been used for crosslinking hyaluronic acid to get three types of hydrogels with differing properties. Swelling studies, rheology analysis, enzymatic degradation and scanning electron microscopic analysis were conducted on all the different types of hydrogels prepared. Viscoelastic properties of the hydrogel were analyzed so that a hydrogel with better elastic property and stability is obtained. Scanning electron microscopy was used to study the morphology of the HA hydrogels. The cytotoxicity testing was conducted to prove the non-toxic nature of the hydrogels and cell culture studies using adipose mesenchymal stem cells showed better adhesion and proliferation properties in all the three hydrogels. Thus hyaluronic acid hydrogel makes a promising material for cartilage regeneration.     

A Multi‐Functional Silicon Nanoparticle Designed for Enhanced Osteoblast Calcification and Related Combination Therapy

Implant materials applied in bone defect commonly focus on the inducement of bone regeneration and neglect to cure complications including bacterial infection and inflammation, which may result in delayed unions or even amputation. In this study, a microporous silica nanoparticle‐poly(N‐isopropylacrylamide‐b‐(2‐(dimethylamino)ethyl methacrylate) is synthesized for loading DXMS and the ECM‐derived peptide (Sequence: Succinic acid‐GTPGPQGIAGQRGVV) in order to enhance the osteoblast calcification and relieve related symptoms. Positively charged PDMA blocks endow the nanoparticle with the antimicrobial property. Moreover, the combination of DXMS makes it have the ability of anti‐inflammation and promoting calcification formation. Furthermore, incorporation of the peptide leads to a significant improvement of mineralization and alkaline phosphatase expression in the preosteoblast. After intramuscular implantation in mice for four weeks, the results indicate the composite nanoparticle can promote ectopic bone formation. These combined properties make the composite silicon nanoparticle a promising osteogenic drug appropriate for further study in bone repair and related combination therapy.     

Preparation,Characterization, and In Vitro Biological Evaluation of PLGA/Nano-Fluorohydroxyapatite (FHA) Microsphere-Sintered Scaffolds for Biomedical Applications

In this research, the novel three-dimensional (3D) porous scaffolds made of poly(lactic-co-glycolic acid) (PLGA)/nano-fluorohydroxyapatite (FHA) composite microspheres was prepared and characterize for potential bone repair applications. We employed a microsphere sintering method to produce 3D PLGA/nano-FHA scaffolds composite microspheres. The mechanical properties, pore size, and porosity of the composite scaffolds were controlled by varying parameters, such as sintering temperature, sintering time, and PLGA/nano-FHA ratio. The experimental results showed that the PLGA/nano-FHA (4:1) scaffold sintered at 90 °C for 2 h demonstrated the highest mechanical properties and an appropriate pore structure for bone tissue engineering applications. Furthermore, MTT assay and alkaline phosphatase activity (ALP activity) results ascertained that a general trend of increasing in cell viability was seen for PLGA/nano-FHA (4:1) scaffold sintered at 90 °C for 2 h by time with compared to control group. Eventually, obtained experimental results demonstrated PLGA/nano-FHA microsphere-sintered scaffold deserve attention utilizing for bone tissue engineering.     

Improving mechanical and biological properties of macroporous HA scaffolds through composite coatings

Interconnected porous hydroxyapatite (HA) scaffolds are widely used for bone repair and replacement, owing to their ability to support the adhesion, transfer, proliferation and differentiation of cells. In the present study, the polymer impregnation approach was adopted to produce porous HA scaffolds with three-dimensional (3D) porous structures. These scaffolds have an advantage of highly interconnected porosity (≈85%) but a drawback of poor mechanical strength. Therefore, the as-prepared HA scaffolds were lined with composite polymer coatings in order to improve the mechanical properties and retain its good bioactivity and biocompatibility at the same time. The composite coatings were based on poly(d,l-lactide) (PDLLA) polymer solutions, and contained single component or combination of HA, calcium sulfate (CS) and chondroitin sulfate (ChS) powders. The effects of composite coatings on scaffold porosity, microstructure, mechanical property, in vitro mineralizing behavior, and cell attachment of the resultant scaffolds were investigated. The results showed that the scaffolds with composite coatings resulted in significant improvement in both mechanical and biological properties while retaining the 3D interconnected porous structure. The in vitro mineralizing behaviors were mainly related to the compositions of CS and ChS powders in the composite coatings. Excellent cell attachments were observed on the pure HA scaffold as well as the three types of composite scaffolds. These composite scaffolds with improved mechanical properties and bioactivities are promising bone substitutes in tissue engineering fields.     

FTIR and NMR study of poly(lactide-co-glycolide) and hydroxyapatite implant degradation under in vivo conditions

The influence of hydroxyapatite (HAP) addition on the rate and mechanism of lactide-co-glycolide copolymer (PGLA) degradation after implantation (in vivo study) was analyzed and compared with the process taking place during in vitro studies. Structural and phase changes of poly(lactide-co-glycolide) and its composite with hydroxyapatite were determined using IR and NMR spectroscopy.Degradation of PGLA and PGLA + HAP composite in biological environment proceeds faster than under in vitro condition. Concentration of glycolidyl units in the copolymer chain decreases and that of lactidyl units increases during in vivo degradation both, in PGLA and in PGLA + HAP composite. However, in the case of the composite the decrease of glycolidyl units concentration is slower and after 6 weeks of degradation the contents of lactidyl and glycolidyl units remain stable. On the other hand, PGLA + HAP composite degrades faster than pure PGLA. The addition of HAP nanoparticles distinctly accelerates degradation of PGLA copolymer which is probably connected with the increase of hydrophilicity of the composite and inhibition of semi-crystalline lactidyl domains formation during the degradation process. Observation of the bone tissue after implantation of PGLA + HAP allows to conclude that the degradation of the composite occurs simultaneously with the implant replacement by the bone cells.     

Bioinspired dual dynamic network hydrogels promote cartilage regeneration through regulating BMSC chondrogenic differentiation

How to improve the therapeutic efficacy of cell delivery during mechanical injection has been a great challenge for tissue engineering. Here, we present a facile strategy based on dynamic chemistry to prepare injectable hydrogels for efficient stem cell delivery using hyaluronic acid (HA) and poly(γ-glutamic acid) (γ-PGA). The combination of the guest–host (GH) complexation and dynamic hydrazone bonds enable the HA/γ-PGA hydrogels with physical and chemical dual dynamic network and endow hydrogels a stable structure, rapid self-healing ability, and injectability. The mechanical properties, self-healing ability, and adaptability can be programmed by changing the ratio of GH network to hydrazine bond cross-linked network. Benefitting from the dynamic cross-linking networks, mild preparation process, and cytocompatibility of HA/γ-PGA hydrogels, bone marrow mesenchymal stem cells (BMSCs) show high cell viability in this system following mechanical injection. Moreover, HA/γ-PGA hydrogels can promote BMSC proliferation and upregulate the expression of cartilage-critical genes. Notably, in a rabbit auricular cartilage defect model, BMSC-laden HA/γ-PGA hydrogels can effectively promote cartilage regeneration. Together, we propose a general strategy to develop injectable self-healing HA/γ-PGA hydrogels for effective stem cell delivery in cartilage tissue engineering.     

Solid-state NMR spectroscopic study of the structure of grafted layers of phosphonic acid ester-modified hydroxyapatite

Interaction of hydroxyapatite (HAP) surface with phosphonic acid esters is studied under different conditions. The urgency of the study is due to the fact that HAP is one of the main components of bone tissue and is widely applied in the production of materials designed for filling bone defects. Modification of the surface properties of HAP may result in the development of materials with increased biocompatibility. It is revealed that, in the absence of solvents or in the medium of an aprotic solvent, phosphonic acid esters interact with HAP surface to yield pyrophosphate groups. The interaction with the surface does not take place in the presence of protic solvents.     

Curcumin‐loaded biodegradable polyurethane scaffolds modified with gelatin using 3D printing technology for cartilage tissue engineering

We described the curcumin‐loaded biodegradable polyurethane (PU) scaffolds modified with gelatin based on three‐dimensional (3D) printing technology for potential application of cartilage regeneration. The printing solution of poly(ε‐caprolactone) (PCL) triol (polyol) and hexamethylene diisocyanate (HMDI) in 2,2,2‐trifluoroethanol was printed through a nozzle in dimethyl sulfoxide phase with or without gelatin. The weight ratio of HMDI against PCL triol was varied as 3, 5, and 7 in order to evaluate its effect on the mechanical properties and biodegradation rate. A higher ratio of HMDI resulted in higher mechanical properties and a lower biodegradation rate. The use of gelatin increased the mechanical properties, biodegradation rate, and curcumin release due to the surface cross‐linking, nanoporous structure, and surface hydrophilicity of the scaffolds. In vitro study revealed that the released curcumin enhanced the proliferation and differentiation of chondrocyte. The 3D‐printed biodegradable PU scaffold modified with gelatin should thus be considered as a potential candidate for cartilage regeneration.     

MC3T3-E1 cell adhesion to hydroxyapatite with adsorbed bone sialoprotein, bone osteopontin, and bovine serum albumin

Native bone tissue is composed of a complex matrix of collagen, non-collagenous proteins, and hydroxyapatite (HAP). Bone sialoprotein (BSP) and bone osteopontin (OPN) are members of the non-collagenous protein family termed the SIBLING (small integrin-binding ligand, N-linked glycoproteins) proteins, which are primarily found in mineralized tissues. Previously, OPN was shown to exhibit a preferential orientation for MC3T3-E1 cell adhesion when it was specifically bound to collagen, while the MC3T3-E1 cell adhesion was shown to be dependant on the conformational flexibility of BSP specifically bound to collagen. Additionally, OPN was shown to play a greater role than BSP for cell binding to collagen. In this work, the orientations and conformations of BSP and OPN specifically bound to HAP are probed under similar conditions. Radiolabeled adsorption isotherms were obtained for BSP and OPN on HAP formed from a simulated body fluid, and the results show that HAP has the capacity to bind significantly more BSP than OPN. An in vitro MC3T3-E1 cell adhesion assay was then performed to compare the cell binding ability of adsorbed BSP and OPN specifically bound to HAP. It was found that there is a preference for cell binding to HAP with adsorbed BSP as compared to OPN, but not to a statistically significant level. However, the maximum cell binding was observed on HAP substrates with adsorbed heat denatured bovine serum albumin (BSA). The influence of BSA on cell binding was shown to be concentration dependant and it is believed that the adsorbed BSA modulates the proliferation state of the bound cells.     

HAP/P(MMA-St)复合微球的制备及其吸附性能

以微孔纳米羟基磷灰石（HAP）为无机载体,甲基丙烯酸甲酯（MMA）和苯乙烯（St）为聚合单体,采用悬浮聚合法制备了HAP/P(MMA-St)复合微球。研究了影响球体粒度及其分布的主要因素和微球的吸附性能。结果表明,表面活性剂用量（质量分数,下同）为1‰、MMA用量为2%,HAP用量为30%,转速为300r/min时复合微球的合格球收率最高为86.4%。当微球中HAP的含量为36.07%时,HAP/P(MMA-St)微球对牛血清蛋白的最大吸附量Qe=18.70 mg/g,比未加HAP时,增加了4.65 mg/g。     

Engineered 3D-Printed Polyvinyl Alcohol Scaffolds Incorporating β-Tricalcium Phosphate and Icariin Induce Bone Regeneration in Rat Skull Defect Model

The skull defects are challenging to self-heal, and autologous bone graft repair has numerous drawbacks. The scaffolds for the rapid and effective repair of skull defects have become an important research topic. In this study, polyvinyl alcohol (PVA)/β-tricalcium phosphate(β-TCP) composite scaffolds containing icariin (ICA) were prepared through direct-ink three-dimensional (3D) printing technology. β-TCP in the composite scaffold had osteoconductive capability, and the ICA molecule had osteoinductive capacity. The β-TCP and ICA components in the composite scaffold can enhance the capability to repair skull defects. We show that ICA exhibited a slow-release behaviour within 80 days. This behaviour helped the scaffold to continuously stimulate the formation of new bone. The results of in vitro cell compatibility experiments showed that the addition of ICA molecules contributed to the adhesion and proliferation of MC-3T3-E1 cells. The level of alkaline phosphatase secretion demonstrated that the slow release of ICA can promote the osteogenic differentiation of MC-3T3-E1 cells. The introduction of ICA molecules accelerated the in situ bone regeneration in in vivo. It is concluded that the 3D-printed PVA scaffold with β-TCP and ICA has a wide range of potential applications in the field of skull defect treatment.     

Interaction between low molecular weight organic acids and hydroxyapatite with different degrees of crystallinity

In this study, two types of hydroxyapatite (HAP) with different degrees of crystallinity were prepared by a sol-gel method and a chemical precipitation method. Influences of crystallinity on the adsorption and dissolution properties of HAP, and the release of phosphorus (total phosphorus) during the adsorption of organic acid were investigated. Results showed that crystallinity had a great effect on the adsorption capacity and dissolution properties of HAP, as well as the adsorption mechanisms of organic acids on HAP surfaces. The poorly crystallized (the degree of crystallinity X_c = 0.23) HAP adsorbed greater amounts of oxalic, citric, or malic acid than the well crystallized (X_c = 0.86) HAP, and the former could release more phosphorus in the presence of organic acids. The adsorption capacity of oxalic acid was much higher than citric and malic acids on both the well and the poorly crystallized HAP, which was due to the strong coordination of oxalic acid with calcium on HAP surface, and that physical adsorption was more inclined to dominate the adsorption of malic or citric acid on the well crystallized HAP. These findings might be of importance in understanding the effects of crystallinity and organic acid binding on the dissolution of calcium phosphates and the adsorption characteristics of HAP.     

One-Step Synthesis of Magnetic Nanocomposite with Embedded Biologically Active Substance

Magnetic nanocomposites based on hydroxyapatite were prepared by a one-step process using the hydrothermal coprecipitation method to sinter iron oxides (Fe₃O₄ and γ-Fe₂O₃). The possibility of expanding the proposed technique for the synthesis of magnetic composite with embedded biologically active substance (BAS) of the 2-arylaminopyrimidine group was shown. The composition, morphology, structural features, and magnetic characteristics of the nanocomposites synthesized with and without BAS were studied. The introduction of BAS into the composite synthesis resulted in minor changes in the structural and physical properties. The specificity of the chemical bonds between BAS and the hydroxyapatite-magnetite core was revealed. The kinetics of the BAS release in a solution simulating the stomach environment was studied. The cytotoxicity of (HAP)FexOy and (HAP)FexOy + BAS composites was studied in vitro using the primary culture of human liver carcinoma cells HepG2. The synthesized magnetic composites with BAS have a high potential for use in the biomedical field, for example, as carriers for magnetically controlled drug delivery and materials for bone tissue engineering.