Fabrication of Nanofiber Reinforced Protein Structures For Tissue Engineering

Extracellular matrices and degradable nanofibers are two very promising materials in the field of tissue engineering; however both of these structures face limitations as tissue engineering scaffolds. Extracellular matrices, such as collagen, gelatin, and laminin, have excellent biocompatibility and allow cell in growth and survival, but structural weakness makes them difficult to handle and greatly limits their uses. Degradable nanofibers support cell attachment and can provide structural support and directional guidance, but individual degradable nanofibers are fragile and have a tendency to form dense fiber bundles which limit cell penetration into the spaces between the nanofibers, especially in the case of aligned nanofibers. To overcome these difficulties, degradable loose nanofibers were embedded in protein matrix in an attempt to fabricate a hybrid scaffold with improved properties, such as improved strength, guidance, spacing among nanofibers, etc. Polycaprolactone (PCL) was used as a model material for degradable nanofibers. Gelatin was employed as a model protein for matrix structure formation. Thin hybrid films (average thickness = 2.78 um) were fabricated by wetting the loose aligned undirectional nanofiber arrays or loose aligned bi-directional nanofiber grids with a gelatin aqueous solution, which also allows for live cell loading into the nanofiber-protein composite if cell are premixed with protein solution or on the surface of the films. Gelatin film alone without nanofiber reinforcement is difficult to handle due to the weakness of the thin membrane. Gelatin films with a fiber density as low as 3% v/v were structurally robust enough for handling, and manipulation into complex shapes. Mechanical testing confirmed that the addition of nanofibers enhanced the strength of gelatin films, in both dry and hydrated state. In vitro testing confirmed that nanofiber reinforced films were biocompatible and provided cells with directional guidance. Results demonstrate the promise of gelatin/PCL nanofiber composites as a tissue scaffolding material.     

羟基磷灰石纳米纤维增强甲基丙烯酸酐改性明胶复合水凝胶的制备及性能

采用溶剂热法制备了具有超高长径比的羟基磷灰石(HAP)纳米纤维,并将其与甲基丙烯酸酐改性明胶(GelMA)结合,利用紫外光交联制备了HAP纳米纤维/GelMA复合水凝胶。通过SEM、XRD、力学测试、溶胀测试、降解测试、细胞培养等对HAP纳米纤维/GelMA复合水凝胶进行结构表征和性能测试。SEM断面观察表明,HAP纳米纤维/GelMA水凝胶呈三维孔隙贯通的多孔结构。力学实验表明,HAP纳米纤维能有效增强水凝胶的弹性模量,且随着HAP纳米纤维添加量的增加,力学性能增强效果越明显。溶胀实验表明,当HAP纳米纤维质量分数为5.2wt%~14.2wt%时,HAP纳米纤维复合水凝胶的溶胀率变化不明显,当质量分数为18.2wt%时,溶胀率降低。降解实验表明,HAP纳米纤维的加入能有效保持水凝胶结构形态,使其更加稳定可控。细胞包裹培养实验表明,HAP纳米纤维/GelMA复合水凝胶能为细胞提供良好的三维生长环境,表现出优良的生物相容性。本实验制备的HAP纳米纤维/GelMA复合水凝胶在组织工程领域有着良好的应用前景。      

纤维素纳米纤丝-碳纳米管/聚乙烯醇-硼酸盐复合导电水凝胶

利用纤维素纳米纤丝(Cellulose nanofibers,CNFs)搭载碳纳米管(Carbon nanotubes,CNTs),在水相中将CNF-CNT复合物均匀分散于聚乙烯醇-硼酸盐(PVA-B)基体中,制备具有立体网络结构的CNF-CNT/PVA-B复合导电水凝胶,旨在提高其动态黏弹性、力学强度和导电性能。结果表明:CNF-CNT/PVA-B内部呈现微米级蜂窝状多孔结构,CNFs与CNTs组成的立体网络在显著提高CNF-CNT/PVA-B力学强度和黏弹性的同时还赋予其导电功能。CNTs含量由0增至0.5wt%时,CNF-CNT/PVA-B的抗压强度和弹性模量分别达到24kPa和53kPa,最大和高频稳态剪切模量分别达到7 028Pa和6 945Pa,电导率达到0.8×10-1 S·cm-1。     

Fabrication of chitosan/poly(ε-caprolactone) composite hydrogels for tissue engineering applications

The aim of this study was to fabricate three-dimensional (3D) porous chitosan/poly(ε-caprolactone) (PCL) hydrogels with improved mechanical properties for tissue engineering applications. A modified emulsion lyophilisation technique was developed to produce 3D chitosan/PCL hydrogels. The addition of 25 and 50 wt% of PCL into chitosan substantially enhanced the compressive strength of composite hydrogel 160 and 290%, respectively, compared to pure chitosan hydrogel. The result of ATR–FTIR imaging corroborated that PCL and chitosan were well mixed and physically co-existed in the composite structures. The composite hydrogels were constructed of homogenous structure with average pore size of 59.7 ± 14 μm and finer pores with average size of 4.4 ± 2 μm on the wall of these larger pores. The SEM and confocal laser scanning microscopy images confirmed that fibroblast cells were attached and proliferated on the 3D structure of these composite hydrogels. The composite hydrogels acquired in this study possessed homogeneous porous structure with improved mechanical strength and integrity. They may have a high potential for the production of 3D hydrogels for tissue engineering applications.     

Facile preparation of low swelling,high strength,self-healing and pH-responsive hydrogels based on the triple-network structure

A polyacrylic acid(PAA)/gelatin(Gela)/polyvinyl alcohol(PVA)hydrogel was prepared by copolymerization,cooling,and freezing/thawing methods.This triplenetwork(TN)structure hydrogel displayed superior mechanical properties,low swelling ratio and self-healing properties,The superior mechanical properties are attributed to the triple helix association of Gela and PVA crystallites by reversible hydrogen bonding.The characterization results indicated that the fracture stress and the strain were 808 kPa and 370% respectively,while the compression strength could reach 4443 kPa and the compressive modulus was up to 39 MPa under the deformation of 90%.The hydrogen bonding in PVA contributed to maintain and improve the self-healing ability of hydrogels.Every type of hydrogels exhibited a higher swelling ratio under alkaline conditions,and the swelling ratios of PAA,PAA/PVA and PAA/Gela hydrogels were 27.71,12.30 and 9.09,respectively.The PAA/Gela/PVA TN hydrogel showed the lowest swelling ratio(6.57)among these hydrogels.These results indicate that the novel TN hydrogels possess good environmental adaptability and have potential applications in the biomedical engineering and sensor field.     

Synthesis, characterization, and in vitro evaluation of a hydrogel-based corneal onlay

Blindness due to opacity of the cornea is treated by corneal transplantation with donor tissue. Due to the limited supply of suitable donor corneas, the need for synthetic corneal equivalents is clear. Herein we report the design and in vitro characterization of a hydrogel-based implant; this implant will serve as a permanent, transparent, space-filling onlay with a two-layer design that mimics the native corneal stratification to support surface epithelialization and foster integration with the surrounding tissue. The top layer of the implant was composed of a 2-hydroxyethylmethacrylate hydrogel containing methacrylic acid as the co-monomer (HEMA-co-MAA) with tunable dimensions and compressive modulus ranging from 700-1000 kPa. The bottom layer, which constitutes the bulk of the implant and is designed to provide integration with the corneal stroma, is a dendrimer hydrogel with high water content and compressive modulus ranging from 500-1200 kPa. Both hydrogels were found to possess optical and diffusion properties similar to those of the human cornea. In addition, composite implants with uniform and structurally sound interfaces were formed when the gels were sequentially injected and cross-linked in the same mold. HEMA-co-MAA hydrogels were covalently modified with type I collagen to enable corneal epithelial cell adhesion and spreading that was dependent upon the collagen coating density but independent of hydrogel stiffness. Similarly, dendrimer hydrogels supported the adhesion and spreading of corneal fibroblasts upon modification with the adhesion ligand arginine-glycine-aspartic acid (RGD). Fibroblast adhesion was not dependent upon dendrimer hydrogel stiffness for the formulations studied and, after in vitro culture for 4 weeks, fibroblasts remained able to adhere to and conformally coat the hydrogel surface. In conclusion, the tunable physical properties and structural integrity of the laminated interface suggests that this design is suitable for further study. The judicious tuning of material properties and inclusion of bioactive moieties is a promising strategy for promotion of implant epithelialization and tissue integration.     

High performance polyimide composite films prepared by homogeneity reinforcement of electrospun nanofibers

Highly aligned polyimide (PI) and PI nanocomposite fibers containing carbon nanotubes (CNTs) were produced by electrospinning. Scanning electron microscopy showed the electrospun nanofibers were uniform and almost free of defects. Transmission electron microscopy indicated that the CNTs were finely dispersed and highly oriented along the CNT/PI nanofiber axis at a relatively low concentration. The as-prepared well-aligned electrospun nanofibers were then directly used as homogeneity reinforcement to enhance the tensile strength and toughness of PI films. The neat PI nanofiber reinforced PI films showed good transparency, decreased bulk density and significantly improved mechanical properties. Compared with neat PI film prepared by solution casting, the tensile strength and elongation at break for the PI film reinforced with 2 wt.% CNT/PI nanofibers were remarkably increased by 138% and 104%, respectively. The significant increases in the overall mechanical properties of the nanofibers reinforced polyimide films can be ascribed to good compatibility between the electrospun nanofibers and the matrix as well as high nanofiber orientation in the matrix. Our study demonstrates a good example for fabricating high performance and high toughness polyimide nanocomposites by using this facile homogeneity self-reinforcement method.     

Microstructural and mechanical characteristics of PHEMA-based nanofibre-reinforced hydrogel under compression

Natural network-structured hydrogels (e.g. bacterial cellulose (BC)) can be synthesised with specific artificial hydrogels (e.g. poly(2-hydroxyethyl methacrylate) (PHEMA)) to form a tougher and stronger nanofibre-reinforced composite hydrogel, which possesses micro- and nano-porous structure. These synthetic hydrogels exhibit a number of advantages for biomedical applications, such as good biocompatibility and better permeability for molecules to pass through. In this paper, the mechanical properties of this nanofibre-reinforced hydrogel containing BC and PHEMA have been characterised in terms of their tangent modulus and fracture stress/strain by uniaxial compressive testing. Numerical simulations based on Mooney-Rivlin hyperelastic theory are also conducted to understand the internal stress distribution and possible failure of the nanofibre-reinforced hydrogel under compression. By comparing the mechanical characteristics of BC, PHEMA, and PHEMA-based nanofibre reinforced hydrogel (BC-PHEMA) under the compression, it is possible to develop a suitable scaffold for tissue engineering on the basis of fundamental understanding of mechanical and fracture behaviours of nanofibre-reinforced hydrogels.     

多价金属离子增强琼脂-聚丙烯酸复合双网络水凝胶及其自修复性能

使水凝胶同时具备高强度和自修复性能,采用"一锅法"制备了多价金属离子Ca~(2+)、Al~(3+)、Fe~(3+)增强琼脂(Agar)-聚丙烯酸(PAAc)复合双网络水凝胶Ca~(2+)/Agar-PAA、Al~(3+)/Agar-PAA和Fe~(3+)/Agar-PAAc双网络水凝胶。研究不同多价金属离子对Agar-PAAc双网络水凝胶的增强作用及Fe~(3+)/Agar-PAAc的自修复自愈合性能。结果表明,Fe~(3+)/Agar-PAAc的拉伸强度(320.7kPa)和断裂伸长率(1 130%)分别为Ca~(2+)/Agar-PAAc的12倍和9倍,Fe~(3+)/Agar-PAAc的断裂伸长率与Al~(3+)/Agar-PAAc相当,但拉伸强度为Al~(3+)/Agar-PAAc的5倍。对样品实施定应变拉伸使内部发生破坏,被破坏的Fe~(3+)/Agar-PAAc在Fe~(3+)溶液中浸泡30 min,修复率达到100%。被破坏的Fe~(3+)/Agar-PAAc在50℃环境中处理15min,修复率达到100%。切断的Fe~(3+)/Agar-PAAc凝胶断面被固定在一起,在密封环境中放置48小时,试样断面可自愈合,且可拉伸至约8倍初始长度。Fe~(3+)/AgarPAAc双网络水凝胶具有优异力学性能和自修复性能,不施加温度、化学物质等外界刺激,实现两重网络均具有自修复性能的效果。     

Electrospun nanofiber belts made from high performance copolyimide

Electrospun nanofibers based on copolyimides were made, aiming at finding a promising method for improving the mechanical properties of electrospun polyimide nanofibers. The copolyimide had a backbone consisting of 3,3',4,4'-biphenyl-tetracarboxylic dianhydride (BPDA), biphenylamide (BPA) and 4,4'-oxydianiline (ODA) residues. The structure and composition of the copolyimide was controlled by the ratio of rigid BPA and flexible ODA moieties. The electrospun copolyimide nanofibers were collected in the form of a belt using a rotating disc with a rim of 8?mm width. Scanning electron microscopy (SEM), infrared (IR) spectroscopy, x-ray scattering and tensile testing, dynamic mechanical analysis (DMA) and thermogravimetric analysis (TGA) were used to characterize the nanofiber belts. The nanofibers had a diameter range from 80 to 300?nm and were well aligned in the belts. The thermal stability of the nanofiber belts was over 460?°C. The tensile test showed that the copolyimide nanofiber belts had much better mechanical properties than either of the flexible and rigid homo-polyimide (homo-PI) nanofiber belts. The tensile strength, modulus and elongation to break of the copolyimide nanofiber belt with BPA/ODA ratio of 40/60 are respectively 1.1 ± 0.1?GPa, 6.2 ± 0.7?GPa and 20.8 ± 1.2%, compared to 459 ± 36?MPa, 2.1 ± 0.3?GPa and 41.3 ± 2.2% for BPDA/ODA homo-PI as well as 384 ± 18?MPa, 11.5 ± 0.6?GPa and 3.9 ± 0.1% for BPDA/BPA homo-PI. The important feature is that the electrospun polymer nanofibers can be made very strong by using copolyimides as spinning materials.     

Reduced Graphene Oxide‐GelMA Hybrid Hydrogels as Scaffolds for Cardiac Tissue Engineering

Biomaterials currently used in cardiac tissue engineering have certain limitations, such as lack of electrical conductivity and appropriate mechanical properties, which are two parameters playing a key role in regulating cardiac cell behavior. Here, the myocardial tissue constructs are engineered based on reduced graphene oxide (rGO)‐incorporated gelatin methacryloyl (GelMA) hybrid hydrogels. The incorporation of rGO into the GelMA matrix significantly enhances the electrical conductivity and mechanical properties of the material. Moreover, cells cultured on composite rGO‐GelMA scaffolds exhibit better biological activities such as cell viability, proliferation, and maturation compared to ones cultured on GelMA hydrogels. Cardiomyocytes show stronger contractility and faster spontaneous beating rate on rGO‐GelMA hydrogel sheets compared to those on pristine GelMA hydrogels, as well as GO‐GelMA hydrogel sheets with similar mechanical property and particle concentration. Our strategy of integrating rGO within a biocompatible hydrogel is expected to be broadly applicable for future biomaterial designs to improve tissue engineering outcomes. The engineered cardiac tissue constructs using rGO incorporated hybrid hydrogels can potentially provide high‐fidelity tissue models for drug studies and the investigations of cardiac tissue development and/or disease processes in vitro.     

高强度耐低温纳米纤维素/聚乙烯醇导电复合水凝胶制备及其在柔性传感中的应用

纳米纤维素具有大长径比、较高的弹性模量与比表面积及丰富的表面官能团,是一种优良的纳米增强材料。首先以纳米纤维素(CNFs)为分散介质辅助分散MXene纳米片层,制备CNF-MXene纳米复合物,并通过FTIR与XPS分析CNFs与MXene的相互作用。以此复合物为增强填料,聚乙烯醇(PVA)为基底,制备CNF-MXene/PVA复合水凝胶,进一步通过KOH溶液处理,提高复合水凝胶的力学性能,并赋予复合水凝胶优异的离子导电性。该复合水凝胶表现出优异的力学性能,其拉伸强度与断裂伸长率分别达到255.9 kPa与1098.2%,还具有高电导率(2.38 S/m)、一定的抗冻性能与灵敏的应变/压力响应性。基于该复合水凝胶组装的应变/压力柔性传感器,由于具有极低的检测极限质量(100 mg)与极快的响应时间(225 ms),可以监控脉搏跳动与喉咙发声微小震动引起的压力变化。因此,该复合水凝胶基柔性传感器非常有希望应用于未来新一代可穿戴电子、人机交互等领域。      

Surface modification of three-dimensional Ca-P/PHBV nanocomposite scaffolds by physical entrapment of gelatin and its in vitro biological evaluation

The properties of bone tissue engineering scaffolds such as architecture, porosity, mechanical properties and surface properties have significant effects on cellular response and play an important role in bone regeneration. In this study, three-dimensional nanocomposite scaffolds consisting of calcium phosphate (Ca-P) nanoparticles and poly(hydroxybutyrate-co-hydroxyvalerate) (PHBV) copolymer with controlled external and internal architectures were successfully produced via selective laser sintering (SLS), one of the versatile rapid prototyping techniques. The Ca-P/PHBV nanocomposite scaffolds had a porosity of (61.75±1.24)%, compressive strength of (2.16±0.21) MPa and Young’s modulus of (26.98±2.29) MPa. The surface modification of scaffolds by gelatin was achieved through physical entrapment. The amount of entrapped gelatin could be controlled by varying the solvent composition and reaction time. The surface modification improved the hydrophilicity of scaffolds but did not significantly affect the surface morphology and mechanical properties. Osteoblast-like cells (SaOS-2) were cultured on scaffolds with and without gelatin surface modification. The majority of SaOS-2 cells were viable and proliferated in both types of scaffolds for up to 14 d in culture, as indicated by MTT assay and live and dead assay. Surface modification significantly increased cell proliferation for surface modified scaffolds, which could be due to the improvement in hydrophilicity of the scaffolds.     

In vitro evaluation of electrospun gelatin–glutaraldehyde nanofibers

The gelatin–glutaraldehyde (gelatin–GA) nanofibers were electrospun in order to overcome the defects of ex-situ crosslinking process such as complex process, destruction of fiber morphology and decrease of porosity. The morphological structure, porosity, thermal property, moisture absorption and moisture retention performance, hydrolytic resistance, mechanical property and biocompatibility of nanofiber scaffolds were tested and characterized. The gelatin–GA nanofiber has nice uniform diameter and more than 80% porosity. The hydrolytic resistance and mechanical property of the gelatin–GA nanofiber scaffolds are greatly improved compared with that of gelatin nanofibers. The contact angle, moisture absorption, hydrolysis resistance, thermal resistance and mechanical property of gelatin–GA nanofiber scaffolds could be adjustable by varying the gelatin solution concentration and GA content. The gelatin–GA nanofibers had excellent properties, which are expected to be an ideal scaffold for biomedical and tissue engineering applications.     

Composite hydrogels for intervertebral disc prostheses

Different PHEMA/PCL semi-IPNs hydrogels and their relative composite systems reinforced with PET fibres have been investigated for potential use as intervertebral disc prostheses. Compression properties and water absorption were evaluated. Uniaxial compression tests on the swollen samples showed an increase of the modulus and maximum stress with increasing content of PCL and PET fibres. In particular, the composite PHEMA/PCL hydrogels showed compression properties similar to those expressed by the canine intervertebral discs in different spinal locations. The equilibrium water content of modified semi-IPNs decreased as function of the PCL and PET fibres. These tests indicate that the use of composite hydrogels as disc prostheses is very promising because it is possible to combine transport and mechanical properties which are crucial for the performance of the intervertebral disc.This paper was accepted for publication after the 1995 Conference of the European Society of Biomaterials, Oporto, Portugal, 10–13 September.     

聚乙二醇改性纳米纤维素/聚乙烯醇复合水凝胶的制备及性能

通过酸碱处理和机械研磨结合的方法制备纳米纤维素(CNFs),并利用冻融循环法分别制备了聚乙烯醇(PVA)和纳米纤维素/聚乙烯醇(CNFs/PVA)复合水凝胶,以及聚乙二醇(PEG)改性PVA和CNFs/PVA复合水凝胶。考察不同配方下复合水凝胶的微观形貌变化,并对复合水凝胶的溶胀性能、压缩强度及热稳定性能进行研究。结果表明,CNFs与PEG对PVA水凝胶的微观形貌均有改善作用,加入PEG后形成的PEG/PVA凝胶产生明显的三维网络结构。当PEG与CNFs同时加入到PVA凝胶后形成的CNFs-PEG/PVA凝胶具有均匀的互穿孔洞结构,此时复合水凝胶的孔隙率最高((67.5±4.3)%),溶胀度最好(980%),且压缩强度较PVA水凝胶也有所提升。PEG对复合凝胶的热稳定性无影响,而加入CNFs后,CNFs-PEG/PVA复合凝胶的初始热分解温度从235℃上升至300℃,显著提高了PVA凝胶的热稳定性。     

利用静电纺丝技术制备纳米黏土/聚乳酸复合纳米纤维与其表征

利用静电纺丝技术制备了纳米黏土/聚乳酸(PLA)复合纳米纤维,并将该复合纳米纤维收集成无纺布薄膜,采用SEM和TEM观察了复合纳米纤维的微观形貌和结构,分别利用XRD和TGA测试了复合纳米纤维的结晶行为及热学行为,并分析了复合纳米纤维薄膜的拉伸力学性能随纳米黏土含量的变化关系。结果表明:当PLA含量为10wt%、纳米黏土含量为1wt%、CHCl3与DMF体积比为3∶1溶剂条件下,所制备的纳米黏土/PLA复合纳米纤维的细度和均匀性均得到改善;XRD测试结果表明,纳米黏土成功附着在PLA中。TGA和力学测试结果表明,纳米黏土/PLA复合纳米纤维的热稳定性和力学性能相对于纯PLA纤维有较大幅度提高,当纳米黏土含量为1wt%时,其初始分解温度提高了60℃,拉伸强度、断裂伸长率和弹性模量分别提高了111.3%、74.9%和20.0%。     

细菌纤维素/聚谷氨酸复合水凝胶的辐照制备及性能

采用Co~(60)-γ射线辐照交联法制备细菌纤维素/聚谷氨酸(BC/PGA)复合水凝胶。采用红外光谱和扫描电子显微镜等对复合水凝胶的结构进行表征,研究了BC引入对复合水凝胶的凝胶分数、热失重、溶胀性能、压缩性能和流变性能的影响,并利用CCK-8法对复合水凝胶进行了细胞毒性评价。研究结果表明,辐照作用下BC纳米纤维和PGA形成双交联复合凝胶网络,BC可有效增加复合水凝胶的压缩强度、储能模量(G')和凝胶分数,降低复合水凝胶的平衡溶胀度。50kGy辐照剂量下,相对于纯PGA水凝胶,复合水凝胶压缩强度增大5倍,G'增大10倍。同时复合水凝胶无细胞毒性,可安全应用于生物医学领域。     

Cellulose nanocomposite biopolymer foam--hierarchical structure effects on energy absorption

Starch is an attractive biofoam candidate as replacement of expanded polystyrene (EPS) in packaging materials. The main technical problems with starch foam include its hygroscopic nature, sensitivity of its mechanical properties to moisture content, and much lower energy absorption than EPS. In the present study, a starch-based biofoam is for the first time able to reach comparable mechanical properties (E = 32 MPa, compressive yield strength, 630 kPa) to EPS at 50% relative humidity and similar relative density. The reason is the nanocomposite concept in the form of a cellulose nanofiber network reinforcing the hygroscopic amylopectin starch matrix in the cell wall. The biofoams are prepared by the freezing/freeze-drying technique and subjected to compressive loading. Cell structure is characterized by FE-SEM of cross sections. Mechanical properties are related to cell structure and cell wall nanocomposite composition. Hierarchically structured biofoams are demonstrated to be interesting materials with potential for strongly improved mechanical properties.     

TiO2改性PC纳米纤维增强PMMA透光复合材料

利用同轴共纺技术制备出壳（聚甲基丙烯酸甲酯,PMMA）－芯（聚碳酸酯,PC）复合纳米纤维,再通过热压将壳层熔融后得到PC纳米纤维增强PMMA透光复合材料. 分别在复合纳米纤维的壳层或芯层中添加不同含量的纳米二氧化钛（TiO₂）粒子,观察纳米颗粒在复合纤维不同结构中的分布,并分析其含量及分布状态对透光复合材料的可见光透过率、紫外光透过率以及力学性能的影响. 研究结果表明,分布在壳层的纳米TiO₂可明显提高复合材料的紫外光屏蔽性,拉伸性能得到增强,但是透光率有所下降;分布在芯层的纳米TiO₂对复合材料的透光率影响较小,而对拉伸性能的提高较引入壳层的效果更为显著.