骨组织工程用纳米羟基磷灰石/ 壳聚糖多孔支架材料的制备及性能表征

以16.7%(质量分数)的柠檬酸水溶液作溶剂,通过粒子沥滤法制备了 n HA/CS多孔材料,并对其进行了IR、XRD、SEM、孔隙率及力学性能测试。结果表明n HA/CS复合材料中羟基磷灰石呈弱结晶状态,复合前后两组分的化学组成未发生显著变化,但两相间发生了相互作用。多孔材料呈高度多孔结构,孔壁上富含微孔,孔间贯通性高;复合材料/致孔剂质量比为1时,多孔材料的孔隙率为 53%,其抗压强度可达17 MPa左右,可以满足组织工程支架材料的要求。     

Preparation of poly(3-hydroxybutyrate)/nano-hydroxyapatite composite scaffolds for bone tissue engineering

Poly(3-hydroxybutyrate)/nano-hydroxyapatite (PHB/nHA) composite scaffolds were fabricated via powder mixing, compression moulding, and particle leaching technique. The scaffolds had high porosity with interconnected porous architecture, a favorable structure for cell attachment and new bone tissue ingrowth. A homogeneous dispersion and a uniform distribution of HA nanoparticles in the polymer matrix were obtained. The scaffolds exhibited improved compressive modulus and compressive strength, which were all in the range of compressive modulus and compressive strength of cancellous bone. In addition, the use of toxic organic solvents was eliminated. Thus, the fabricated PHB/nHA composite scaffolds tend to be promising for application in bone tissue engineering.     

Fabrication and in vitro evaluation of a sponge-like bioactive-glass/gelatin composite scaffold for bone tissue engineering

In this work a bioactive composite scaffold, comprised of bioactive-glass and gelatin, is introduced. Through direct foaming a sponge-like composite of a sol–gel derived bioactive-glass (70S30C; 70% SiO₂, 30% CaO) and porcine gelatin was developed for use as a biodegradable scaffold for bone tissue engineering. The composite was developed to provide a suitable alternative to synthetic polymer based scaffolds, allowing directed regeneration of bone tissue. The fabricated scaffold was characterised through X-ray microtomography, scanning electron and light microscopy demonstrating a three dimensionally porous and interconnected structure, with an average pore size (170 μm) suitable for successful cell proliferation and tissue ingrowth. Acellular bioactivity was assessed through apatite formation during submersion in simulated body fluid (SBF) whereby the rate and onset of apatite nucleation was found to be comparable to that of bioactive-glass. Modification of dehydrothermal treatment parameters induced varying degrees of crosslinking, allowing the degradation of the composite to be tailored to suit specific applications and establishing its potential for a wide range of applications. Use of genipin to supplement crosslinking by dehydrothermal treatment provided further means of modifying degradability. Biocompatibility of the composite was qualified through successful cultures of human dental pulp stem cells (HDPSCs) on samples of the composite scaffold. Osteogenic differentiation of HDPSCs and extracellular matrix deposition were confirmed through positive alkaline phosphatase staining and immunohistochemistry.     

In vitro evaluation of electrospun PCL/nanoclay composite scaffold for bone tissue engineering

Polycaprolactone (PCL) is a widely accepted synthetic biodegradable polymer for tissue engineering, however its use in hard tissue engineering is limited because of its inadequate mechanical strength and low bioactivity. In this study, we used halloysite nanoclay (NC) as an inorganic filler material to prepare PCL/NC fibrous scaffolds via electrospinning technique after intercalating NC within PCL by solution intercalation method. The obtained nanofibrous mat was found to be mechanically superior to PCL fibrous scaffolds. These scaffolds allowed greater protein adsorption and enhanced mineralization when incubated in simulated body fluid. Moreover, our results indicated that human mesenchymal stem cells (hMSCs) seeded on these scaffolds were viable and could proliferate faster than in PCL scaffolds as confirmed by fluorescence and scanning electron microscopic observations. Further, osteogenic differentiation of hMSCs on nanoclay embedded scaffolds was demonstrated by an increase in alkaline phosphatase activity when compared to PCL scaffold without nanoclay. All of these results suggest the potential of PCL/NC scaffolds for bone tissue engineering.     

磷酸钙骨水泥/明胶复合组织工程支架材料的制备及其结构与性能

磷酸钙骨水泥组织工程支架材料具有良好的生物相容性和骨传导性,是一种良好的骨组织工程支架材料,但是这种材料存在力学性能差的缺点,限制了它的应用.本文采用生物相容性良好的可降解明胶材料与磷酸钙骨水泥支架进行复合,制备出的明胶/磷酸钙骨水泥复合支架材料,其压缩强度可达3.7MPa,比复合前磷酸钙支架材料的强度提高了37倍,而且材料具有良好的柔韧性,适合用作为非承重部位骨组织缺损修复用组织工程支架材料.     

Chitosan-collagen/organomontmorillonite scaffold for bone tissue engineering

A novel composite scaffold based on chitosan-collagen/organomontmo-rillonite (CS-COL/OMMT) was prepared to improve swelling ratio, biodegradation ratio, biomineralization and mechanical properties for use in tissue engineering applications. In order to expend the basal spacing, montmorillonite (MMT) was modified with sodium dodecyl sulfate (SDS) and was characterized by XRD, TGA and FTIR. The results indicated that the anionic surfactants entered into interlayer of MMT and the basal spacing of MMT was expanded to 3.85 nm. The prepared composite scaffolds were characterized by FTIR, XRD and SEM. The swelling ratio, biodegradation ratio and mechanical properties of composite scaffolds were also studied. The results demonstrated that the scaffold decreased swelling ratio, degradation ratio and improved mechanical and biomineralization properties because of OMMT.     

Bioactive nanocomposite PLDL/nano-hydroxyapatite electrospun membranes for bone tissue engineering

New nanocomposite membranes with high bioactivity were fabricated using the electrospinning. These nanocomposites combine a degradable polymer poly(l/dl)-lactide and bone cell signaling carbonate nano-hydroxyapatite (n-HAp). Chemical and physical characterization of the membranes using scanning electron microscopy, Fourier transform infrared spectroscopy and the wide angle X-ray diffraction evidenced that nanoparticles were successfully incorporated into the fibers and membrane structure. The incorporation of the n-HAp into the structure increased significantly the mineralization of the membrane in vitro. It has been demonstrated that after a 3-day incubation of composite membrane in the Simulated Body Fluid a continuous compact apatite layer was formed. In vitro experiments demonstrated that the incorporation of n-HAp significantly improved cell attachment, upregulated cells proliferation and stimulated cell differentiation quantified using Alkaline Phosphatase and OsteoImage tests. In conclusion, the results demonstrated that the addition of n-HAp provided chemical cues that were a key factor that regulated osteoblastic differentiation.     

Fabrication of nano-structured electrospun collagen scaffold intended for nerve tissue engineering

Nerve tissue engineering is one of the most promising methods in nerve tissue regeneration. The development of blended collagen and glycosaminoglycan scaffolds can potentially be used in many soft tissue engineering applications. In this study an attempt was made to develop two types of random and aligned electrospun, nanofibrous scaffold using collagen and a common type of glycosaminoglycan. Ion chromatography test, MTT and attachment assays were conducted respectively to trace the release of glycosaminoglycan, and to investigate the biocompatibility of the scaffold. Cell cultural tests showed that the scaffold acted as a positive factor to support connective tissue cell outgrowth. The positive effect of fiber orientation on cell outgrowth organization was traced through SEM images. Porosity percentage calculation and tensile strength measurement of the webs specified analogous properties to the native neural matrix tissue. These results suggested that nanostructured porous collagen-glycosaminoglycan scaffold is a potential cell carrier in nerve tissue engineering.     

可缓释药物的明胶/磷酸钙骨水泥复合组织工程支架材料

采用向孔隙中灌注含聚乳酸聚乙醇酸共聚物（PLGA）载药微球的明胶溶液的方法制备了具有药物缓释功能的明胶/磷酸钙骨水泥复合组织工程支架。用扫描电子显微镜观察了微球和支架的形貌特征,用万能材料试验机测定了支架材料的抗压强度,用紫外-可见分光光度计分析了复合支架的释药率。结果表明,灌注明胶对多孔磷酸钙骨水泥支架起到显著的增强作用,抗压强度达2.42 MPa。复合支架携载硫酸庆大霉素, 具有良好的药物缓释功能,缓释时间可达30天以上,使支架在修复骨缺损的同时能消除炎症反应,成为一种集骨修复和治疗于一体的新型组织工程支架材料,具有良好的应用前景。      

骨组织工程用支架材料研究进展

骨组织工程用支架材料是骨组织工程研究的主要内容之一 ,按来源可将支架分为天然支架材料和合成支架材料 ,本文综述了骨组织工程用支架材料的研究和应用。     

Electrospun composites of PHBV,silk fibroin and nano-hydroxyapatite for bone tissue engineering

Electrospinning of fibrous scaffolds containing nano-hydroxyapatite (nHAp) embedded in a matrix of functional biomacromolecules offers an attractive route to mimicking the natural bone tissue architecture. Functional fibrous substrates will support cell attachment, proliferation and differentiation, while the role of HAp is to induce cells to secrete extracellular matrix (ECM) for mineralization to form bone. Electrospinning of biomaterials composed of polyhydroxybutyrate-co-(3-hydroxyvalerate) with 2% valerate fraction (PHBV), nano-hydroxyapatite (nHAp), and Bombyx mori silk fibroin essence (SF), Mw = 90KDa, has been achieved for nHAp and SF solution concentrations of 2 (w/vol) % each and 5 (w/vol) % each. The structure and properties of the nanocomposite fibrous membranes were investigated by means of Scanning Electron Microscopy in combination with Energy Dispersive X-Ray Analysis (SEM/EDX), Fourier Transformed Infrared Spectroscopy (FT-IR), uniaxial tensile and compressive mechanical testing, degradation tests and in vitro bioactivity tests. SEM images showed smooth, uniform and continuous fibre deposition with no bead formation, and fibre diameters of between 10 and 15 μm. EDX and FT-IR confirmed the presence of nHAp and SF. After one month in deionised water, tests showed less than 2% weight loss with the samples retaining their fibrous morphology, confirming that this material biodegrades slowly. After 28 days of immersion in Simulated Body Fluid (SBF) an apatite layer was visible on the surface of the fibres, proving their bioactivity. Preliminary in vitro biological assessment showed that after 1 and 3 days in culture, cells were attached to the fibres, retaining their morphology while presenting a flattened appearance and elongated shape on the surface of fibres. Young's modulus was found to increase from 0.7 kPa (± 0.33 kPa) for electrospun samples of PHBV only to 1.4 kPa (± 0.54 kPa) for samples with 2 (w/vol) % each of nHAp and SF. Samples prepared with 5 (w/vol) % each of nHAp and SF did not show a similar improvement.     

Needle-like nano hydroxyapatite/poly(l-lactide acid) composite scaffold for bone tissue engineering application

In this paper, a new nano-hydroxyapatite / poly (l-lactide acid) (nHAP/PLLA) composite scaffold comprising needle-like nHAP particles was prepared. In the first step, the identification and morphology of chemically synthesized HAP particles were determined by XRD, EDX, FTIR and SEM analyses. The needle-like nHAP particles with an average size of approximately 30–60 nm in width and 100–400 nm in length were found similar to needle-like bone nano apatites in terms of chemical composition and morphology. In the second step, nHAP and micro-sized HAP (mHAP) particles were used to fabricate HAP filled PLLA (HAP/PLLA) composites scaffolds using solid–liquid phase separation method. The porosity of scaffolds was up to 85%, and their average macropore diameter was in the range of 64–175 µm. FTIR and XRD analyses showed the presence of molecular interactions and chemical linkages between HAP particles and PLLA matrix. The compressive strength of nanocomposite scaffolds could high up to 8.46 MPa while those of pure PLLA and microcomposite scaffolds were 1.79 and 4.61 MPa, respectively. The cell affinity and cytocompatibility of the nanocomposite scaffold were found to be higher than those of pure PLLA and microcomposite scaffolds. Based on the results, the newly developed nHAP/PLLA composite scaffold is comparable with cancellous bone in terms of microstructure and mechanical strength, so it may be a suitable alternative for bone tissue engineering applications.     

A smart bilayer scaffold of elastin-like recombinamer and collagen for soft tissue engineering

Elastin-like recombinamers (ELRs) are smart, protein-based polymers designed with desired peptide sequences using recombinant DNA technology. The aim of the present study was to produce improved tissue engineering scaffolds from collagen and an elastin-like protein tailored to contain the cell adhesion peptide RGD, and to investigate the structural and mechanical capacities of the resulting scaffolds (foams, fibers and foam-fiber bilayer scaffolds). The results of the scanning electron microscopy, mercury porosimetry and mechanical testing indicated that incorporation of ELR into the scaffolds improved the uniformity and continuity of the pore network, decreased the pore size (from 200 to 20 μm) and the fiber diameter (from 1.179 μm to 306 nm), broadened the pore size distribution (from 70–200 to 4–200 μm) and increased their flexibility (from 0.007 to 0.011 kPa⁻¹). Culture of human fibroblasts and epithelial cells in ELR-collagen scaffolds showed the positive contribution of ELR on proliferation of both types of cells.     

一种新型的骨组织工程用支架材料的制备与性能表征

以硼硅酸盐玻璃粉为原料,采用有机泡沫浸渍工艺,制备了高孔隙率的网眼多孔支架.应用XRD、SEM及ICP-AES等对硼酸盐生物玻璃粉末在生理模拟液中的降解性能、生物活性等进行了测试分析.结果表明,硼硅酸盐生物玻璃的降解性和生物活性与材料的组成配比有关,因此,可以通过调整玻璃的组成有效控制材料的降解性和表面形成的羟基磷灰石晶体的形态.硼硅酸盐生物活性玻璃作为硬组织工程支架材料的研究具有重要的意义和广泛的应用前景.     

A biomimetic three-dimensional woven composite scaffold for functional tissue engineering of cartilage

Tissue engineering seeks to repair or regenerate tissues through combinations of implanted cells, biomaterial scaffolds and biologically active molecules. The rapid restoration of tissue biomechanical function remains an important challenge, emphasizing the need to replicate structural and mechanical properties using novel scaffold designs. Here we present a microscale 3D weaving technique to generate anisotropic 3D woven structures as the basis for novel composite scaffolds that are consolidated with a chondrocyte-hydrogel mixture into cartilage tissue constructs. Composite scaffolds show mechanical properties of the same order of magnitude as values for native articular cartilage, as measured by compressive, tensile and shear testing. Moreover, our findings showed that porous composite scaffolds could be engineered with initial properties that reproduce the anisotropy, viscoelasticity and tension-compression nonlinearity of native articular cartilage. Such scaffolds uniquely combine the potential for load-bearing immediately after implantation in vivo with biological support for cell-based tissue regeneration without requiring cultivation in vitro.     

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.     

羟基丁酸与羟基辛酸共聚体骨组织工程支架的初步研究

采用粒子滤出/冷冻干燥复合法制备羟基丁酸与羟基辛酸共聚体[P(HB-HO)]多孔支架,并进行扫描电镜观察、孔隙率和力学性能检测以及体外降解和细胞相容性实验.结果表明,P(HB-HO)多孔支架孔隙分布均匀,连通性好,孔隙率为50%～90%时,抗压强度在1.7～6.2MPa之间,十二周体外降解率约为20%;与P(HB-H0)复合培养的小鼠成骨样细胞MC3T3-E1黏附率高,生长状态良好.证明P(HB-HO)具有良好的理化性能和细胞相容性,有望成为一种具有临床应用价值的骨组织工程支架材料.     

Application of strontium-doped calcium polyphosphate scaffold on angiogenesis for bone tissue engineering

The key factor for regenerating large segmental bone defects through bone tissue engineering is angiogenesis in scaffolds. Attempts to overcome this problem, it is a good strategy to develop a new scaffold with bioactivity to induce angiogenesis in bone tissue engineering. In our previous research, the ability of strontium-doped calcium polyphosphate (SCPP) to stimulate the release of angiogenic growth factors from cultured osteoblasts was studied. This study was performed to determine the ability of SCPP to induce angiogenesis within in vitro co-culture model of human umbilical vein endothelial cells (HUVEC) and osteoblasts co-cultured. The bioactivity of developed scaffolds to induce angiogenesis in vivo was also researched in this paper. Co-cultured model has been developed in vitro and then cultured with SCPP scaffold as well as calcium polyphosphate (CPP) scaffold and hydroxylapatite (HA) scaffold. The results showed that the optimal ratio of HUVEC and osteoblasts co-cultured model for in vitro angiogenesis was 5:1. The model could maintain for more than 35 days when cultured with the scaffold and show the best activity at 21st day. Compared with those in CPP and HA scaffold, the formation of tube-like structure and the expression of platelet endothelial cell adhesion molecule in co-cultured model is better in SCPP scaffold. The in vivo immunohistochemistry staining for VEGF also showed that SCPP had a potential to promote the formation of angiogenesis and the regeneration of bone. SCPP scaffold could be served as a potential biomaterial with stimulating angiogenesis in bone tissue engineering and bone repair.     

Preparation of flexible bone tissue scaffold utilizing sea urchin test and collagen

Gonads of sea urchin are consumed in Japan and some countries as food and most parts including its tests are discarded as marine wastes. Therefore, utilization of them as functional materials would reduce the waste as well as encourage Japanese fishery. In this study, magnesium containing calcite granules collected from sea urchin tests were hydrothermally phosphatized and the obtained granules were identified as approximately 82% in mass of magnesium containing β-tricalcium phosphate and 18% in mass of nonstoichiometric hydroxyapatite, i.e., a biphasic calcium phosphate, maintaining the original porous network. Shape-controlled scaffolds were fabricated with the obtained biphasic calcium phosphate granules and collagen. The scaffolds showed good open porosity (83.84%) and adequate mechanical properties for handling during cell culture and subsequent operations. The MG-63 cells showed higher proliferation and osteogenic differentiation in comparison to a control material, the collagen sponge with the same size. Furthermore, cell viability assay proved that the scaffolds were not cytotoxic. These results suggest that scaffold prepared using sea urchin test derived calcium phosphate and collagen could be a potential candidate of bone void fillers for non-load bearing defects in bone reconstruction as well as scaffolds for bone tissue engineering.     

Development and cell response of a new biodegradable composite scaffold for guided bone regeneration

Composites of biodegradable polymers with different calcium phosphate ceramics and glasses, have been developed as scaffolds for applications in bone-tissue engineering. In this work, phosphate glass particles have been incorporated into the polymer, poly(95L/5DL) lactic acid (PLA) and porous structures were elaborated. Their porosity, compressive mechanical properties and biological response were evaluated. Interconnected structures with evenly distributed pores and a porosity as high as 97% were obtained. The incorporation of glass particles into the polymer showed to have a positive effect in the mechanical properties of the foams. Indeed, the compressive modulus increased from 74.5 to 120 KPa and the compressive strength from 17.5 to 20.1 KPa for the PLA and the PLA/glass foams, respectively. The biological response was evaluated by means of the MTT test, the materials resulted to be noncytotoxic.