激光熔融静电纺丝法制备PLLA/PCL/nHA复合纤维支架及细胞相容性评价

利用激光熔融静电纺丝技术制备了PLLA/PCL及PLLA/PCL/nHA复合纤维,热压后形成层压复合纤维支架。利用扫描电镜对纤维支架进行了表征,同时对其进行了亲水性的测试,最后通过倒置荧光显微镜和MTT实验对复合纤维支架的细胞相容性进行了评价。研究结果表明,层压复合纤维支架的直径和孔结构具有多样性,nHA能够提高PLLA/PCL层压纤维支架的亲水性,改善支架的细胞相容性,增加细胞的附着能力,提高细胞的存活率。     

聚己内酯/明胶电纺纳米纤维支架的制备与细胞相容性研究

构建了聚己内酯/明胶(PCL/GE)电纺纳米纤维支架,考察了该支架在模拟体液中的降解速率及其细胞相容性。结果表明:相比于PCL纤维膜,支架表面具有良好的亲水性,且PCL/GE复合纤维支架降解速率得到极大提高;大鼠脂肪来源间充质干细胞(marrow stromal cells,MSCs)在纤维支架表面粘附力强,生长、增殖情况良好。     

静电纺丝制备小直径血管支架及其血液相容性的研究

以甲酸为溶剂,将具有特定信号识别功能的RGD重组蛛丝蛋白(pNSR32)与聚己内酯(PCL)和壳聚糖(CS)共混,采用静电纺丝技术制备复合纳米纤维小直径血管支架,扫描电子显微镜(SEM)观察支架材料的微观结构,并通过溶血率、动态凝血时间、血小板黏附以及复钙化凝血时间实验评价支架材料的血液相容性.研究表明,(1) 所制备的pNSR32/PCL/CS复合纳米纤维支架内径为4mm,长度可达8cm,并且纤维连续,孔隙相通,孔隙率达85%以上,支架具有三维多孔网状结构,有利于种子细胞在支架上的黏附、增殖和迁移;(2)复合支架材料的溶血率＜5%,黏附的血小板少且不变形,复钙化凝血时间达293s,与单独PCL相比延长94s,说明该支架材料具有良好的血液相容性,有望成为一种新型的小直径组织工程血管支架.     

壳聚糖/羟基磷灰石表面修饰聚己内酯多孔骨支架的制备及性能

采用选择性激光烧结技术构建多孔聚己内酯(PCL)骨支架,用原位合成的方法制得壳聚糖/羟基磷灰石(CS/HA)悬浮液,并采用真空浸泡、低速离心和冷冻凝胶的方法使CS/HA黏附在PCL支架的表面,以改善骨支架的生物相容性和细胞增殖活性。通过X射线衍射(XRD)和扫描电子显微镜(SEM)观测复合支架的物相和形貌,测量支架的压缩强度和杨氏模量,测量支架表面的水接触角,并通过体外细胞实验研究复合支架的生物学性能。实验结果表明,原位合成的方法制得了羟基磷灰石(HA);CS/HA凝胶与PCL骨支架表面黏附良好;CS/HA改善了PCL支架表面的亲水性,提升了骨支架的生物相容性和细胞增殖活性。     

磷灰石-硅灰石生物活性玻璃陶瓷表面接枝多肽改性研究

为在磷灰石-硅灰石生物活性玻璃陶瓷(Apatite-Wollastonite Bioactive Glass-Ceramic, AW)表面引入能够促进细胞粘附的RGD(精氨酸-甘氨酸-天冬氨酸)多肽以提高其生物活性, 采用低温等离子法在材料的表面引入活性氨基基团, 并通过浸渍法使氨基基团与多肽发生反应。采用XRD、XPS、ATR-FTIR对AW的相组成及表面改性特性进行表征, 确认通过低温等离子法在AW表面接上氨基, RGD多肽分子与氨基反应以化学键合的形式接枝到材料表面(RGD-AW), 实现了在AW表面接枝生物大分子的改性。将改性前后的材料分别与类成骨细胞(MG63细胞)混合培养并使用荧光显微镜、SEM及MTT等测试方法对材料的细胞生物学性能进行了表征。细胞实验结果表明: 接枝RGD多肽分子的材料在细胞培养的早期阶段比AW更有利于细胞的粘附及铺展。     

纳米非计量磷灰石/聚合物复合支架的制备和性能研究

采用快速成形法制备了孔径和孔隙率可控、大孔互相贯通的纳米缺钙羟基磷灰石(cd-HA)与聚己内酯(PCL)复合材料多孔支架,并对复合支架的微结构进行了表征.通过细胞培养和体内动物实验研究了该支架的生物学性能.结果表明:复合材料的亲水性和细胞粘附率随磷灰石含量增加而提高;成骨细胞在复合支架上的增殖明显高于纯PCL;μ-CT和组织学分析结果显示,新骨在支架的表面形成并长入其中.相互贯通的多孔支架促进了细胞的增殖和新骨长入支架内部.cd-HA/PCL复合材料支架具有很好的生物相容性,在组织工程领域中有潜在的应用前景.     

静电自组装表面修饰HA/PA6膜:表征及其细胞相容性研究

通过表面改性引入活性生物分子可以用来增强材料表面的生物相容性。采用静电自组装技术将明胶和聚乙烯亚胺引入羟基磷灰石/聚酰胺6(HA/PA6)材料表面,最终获得大分子修饰的复合材料。利用水接触角测量(WCA)、X射线光电子能谱(XPS)和原子力显微镜(AFM)测试手段对表面改性前后HA/PA6膜的表面化学、亲水性和表面形貌进行表征。并研究了其对细胞活性的影响。结果表明,改性后膜表面的亲水性增强,粗糙度增大。体外细胞实验表明,明胶涂层后的HA/PA6表面细胞更利粘附、铺展和生长。该固定方法模拟细胞外基质组成制备出的生物活性膜能进一步满足生物医学工程需求。     

聚酰胺66膜表面明胶固定化研究

生物材料可通过表面改性引入活性基团来增强其生物相容性.采用紫外辐照方法,以亚铁离子辅助引发接枝聚合,将羧基引入聚酰胺66(PA66)膜表面,再用水溶性碳化二亚胺(WSC)作为缩合剂,进一步将明胶固定在聚酰胺膜表面,最终获得大分子修饰的聚酰胺材料.利用衰减全反射-傅里叶红外光谱(ATR-FTIR)、X射线光电子能谱(XPS)、扫描电子显微镜(SEM)和水接触角测量(WCA)等测试手段对PA66膜和表面改性PA66膜进行了表征.结果表明,在40℃条件下,光氧化的最佳时间为60min.接枝聚合后,膜的亲水性增强,水接触角由纯聚酰胺的67.5°变为固定明胶分子后的60°,并且膜表面的粗糙度增加.表面改性PA66膜可促进细胞活性,适用于组织工程中.     

组织工程支架材料生物功能化的研究进展

组织工程的目标之一就是制备出能引导细胞分化并产生具有一定功能再生组织的支架材料,支架材料生物功能化能够有效促进细胞的生物活动,是组织再生的关键。本文从提高支架材料生物功能化入手,说明纳米材料是蛋白和细胞吸附的最佳选择,重点分析了化学改性、引入RGD多肽片断、多肽-聚合物支架及ECM基支架材料四种提高支架材料生物功能化的方法,并对支架材料生物功能化的未来发展趋势进行了展望。     

静电纺丝天然-人工合成聚合物复合纳米纤维及其对细胞黏附、增殖和迁移的影响

生物材料组成成分对细胞生物功能有不同的影响。利用静电纺丝技术制备了基于聚己内酯(PCL,polycaprolactone)的不同天然蛋白、多糖(丝素蛋白(SF,silk fibroin)、透明质酸(HA,hyaluronicacid))的混合组分纳米纤维,采用了扫描电镜和接触角对纳米纤维进行基础表征。同时,进一步考察了纳米纤维作为组织工程支架的可行性。研究结果表明SF组分能增加材料的可纺性,有利于细胞的前期黏附,并能够促进细胞增殖。HA组分可以改善材料的亲水性,增加细胞伪足并促进细胞迁移。重要的是,PCL/SF/HA纳米纤维能同时结合SF和HA的优点,有望在组织工程领域得到应用。     

Investigations on the in vitro bioactivity of swift heavy oxygen ion irradiated hydroxyapatite

Poly(propylene fumarate) (PPF) is an ultraviolet-curable and biodegradable polymer with potential applications for bone regeneration. In this study, we designed and fabricated three-dimensional (3D) porous scaffolds based on a PPF polymer network using micro-stereolithography (MSTL). The 3D scaffold was well fabricated with a highly interconnected porous structure and porosity of 65%. These results provide a new scaffold fabrication method for tissue engineering. Surface modification is a commonly used and effective method for improving the surface characteristics of biomaterials without altering their bulk properties that avoids the expense and long time associated with the development of new biomaterials. Therefore, we examined surface modification of 3D scaffolds by applying accelerated biomimetic apatite and arginine-glycine-aspartic acid (RGD) peptide coating to promote cell behavior. The apatite coating uniformly covered the scaffold surface after immersion for 24 h in 5-fold simulated body fluid (5SBF) and then the RGD peptide was applied. Finally, the coated 3D scaffolds were seeded with MC3T3-E1 pre-osteoblasts and their biologic properties were evaluated using an MTS assay and histologic staining. We found that 3D PPF/diethyl fumarate (DEF) scaffolds fabricated with MSTL and biomimetic apatite coating can be potentially used in bone tissue engineering.     

Effects of surface modification on the mechanical and structural properties of nanofibrous poly(ε-caprolactone)/forsterite scaffold for tissue engineering applications

Composite scaffolds consisting of polymers reinforced with ceramic nanoparticles are widely applied for hard tissue engineering. However, due to the incompatible polarity of ceramic nanoparticles with polymers, they tend to agglomerate in the polymer matrix which results in undesirable effects on the integral properties of composites. In this research, forsterite (Mg₂SiO₄) nanoparticles was surface esterified by dodecyl alcohol and nanofibrous poly(ε-caprolactone)(PCL)/modified forsterite scaffolds were developed through electrospinning technique. The aim of this research was to investigate the properties of surface modified forsterite nanopowder and PCL/modified forsterite scaffolds, before and after hydrolytic treatment, as well as the cellular attachment and proliferation. Results demonstrated that surface modification of nanoparticles significantly enhanced the tensile strength and toughness of scaffolds upon 1.5- and 4-folds compared to unmodified samples, respectively, due to improved compatibility between matrix and filler. Hydrolytic treatment of scaffolds also modified the bioactivity and cellular attachment and proliferation due to greatly enhanced hydrophilicity of the forsterite nanoparticles after this process compared to surface modified samples. Results suggested that surface modification of forsterite nanopowder and hydrolytic treatment of the developed scaffolds were effective approaches to address the issues in the formation of composite fibers and resulted in development of bioactive composite scaffolds with ideal mechanical and structural properties for bone tissue engineering applications.     

三维打印羟基磷灰石晶须增强复合骨修复支架

利用三维打印技术成功制备羟基磷灰石晶须(HAPw)增强的聚己内酯(PCL)复合骨修复支架。通过改变三维打印的挤出速度和挤出气压, 使不同含量HAPw均能在PCL基材中一致排列并均匀分布。PCL支架的机械强度随HAPw含量增加显著提高, 添加33wt%HAPw使PCL支架强度提升了高达3倍。此外, HAPw使PCL支架表面与水的接触角从近100º降低至约50º, 有效改善了细胞表面粘附。经过体外人类骨髓间充质干细胞(hBMSC)在支架上的培养实验, 发现添加HAPw的复合支架具有更好的生物相容性, 能够有效促进hBMSC的增殖生长, 且HAPw-PCL复合支架上细胞具有更高的碱性磷酸酶(ALP)活性和OCN、RUNX2等相关成骨基因表达, 显示出hBMSCs向成骨方向更好的分化及成骨活性。     

Robotic dispensing of composite scaffolds and in vitro responses of bone marrow stromal cells

The development of bioactive scaffolds with a designed pore configuration is of particular importance in bone tissue engineering. In this study, bone scaffolds with a controlled pore structure and a bioactive composition were produced using a robotic dispensing technique. A poly(ε-caprolactone) (PCL) and hydroxyapatite (HA) composite solution (PCL/HA = 1) was constructed into a 3-dimensional (3D) porous scaffold by fiber deposition and layer-by-layer assembly using a computer-aided robocasting machine. The in vitro tissue cell compatibility was examined using rat bone marrow stromal cells (rBMSCs). The adhesion and growth of cells onto the robotic dispensed scaffolds were observed to be limited by applying the conventional cell seeding technique. However, the initially adhered cells were viable on the scaffold surface. The alkaline phosphatase activity of the cells was significantly higher on the HA–PCL than on the PCL and control culture dish, suggesting that the robotic dispensed HA–PCL scaffold should stimulate the osteogenic differentiation of rBMSCs. Moreover, the expression of a series of bone-associated genes, including alkaline phosphatase and collagen type I, was highly up-regulated on the HA–PCL scaffold as compared to that on the pure PCL scaffold. Overall, the robotic dispensed HA–PCL is considered to find potential use as a bioactive 3D scaffold for bone tissue engineering. Seok-Jung Hong and Ishik Jeong contributed equally.     

Polymer Powder Processing of Cryomilled Polycaprolactone for Solvent‐Free Generation of Homogeneous Bioactive Tissue Engineering Scaffolds

Synthetic polymers used in tissue engineering require functionalization with bioactive molecules to elicit specific physiological reactions. These additives must be homogeneously dispersed in order to achieve enhanced composite mechanical performance and uniform cellular response. This work demonstrates the use of a solvent‐free powder processing technique to form osteoinductive scaffolds from cryomilled polycaprolactone (PCL) and tricalcium phosphate (TCP). Cryomilling is performed to achieve micrometer‐sized distribution of PCL and reduce melt viscosity, thus improving TCP distribution and improving structural integrity. A breakthrough is achieved in the successful fabrication of 70 weight percentage of TCP into a continuous film structure. Following compaction and melting, PCL/TCP composite scaffolds are found to display uniform distribution of TCP throughout the PCL matrix regardless of composition. Homogeneous spatial distribution is also achieved in fabricated 3D scaffolds. When seeded onto powder‐processed PCL/TCP films, mesenchymal stem cells are found to undergo robust and uniform osteogenic differentiation, indicating the potential application of this approach to biofunctionalize scaffolds for tissue engineering applications.     

3D cell growth and proliferation on a RGD functionalized nanofibrillar hydrogel based on a conformationally restricted residue containing dipeptide

Three-dimensional (3D) hydrogels incorporating a compendium of bioactive molecules can allow efficient proliferation and differentiation of cells and can thus act as successful tissue engineering scaffolds. Self-assembled peptide-based hydrogels can be worthy candidates for such applications as peptides are biocompatible, biodegradable and can be easily functionalized with desired moieties. Here, we report 3D growth and proliferation of mammalian cells (HeLa and L929) on a dipeptide hydrogel chemically functionalized with a pentapeptide containing Arg-Gly-Asp (RGD) motif. The method of functionalization is simple, direct and can be adapted to other functional moieties as well. The functionalized gel was noncytotoxic, exhibited enhanced cell growth promoting properties, and promoted 3D growth and proliferation of cells for almost 2 weeks, with simultaneous preservation of their metabolic activities. The presence of effective cell growth supporting properties in a simple and easy to functionalize dipeptide hydrogel is unique and makes it a promising candidate for tissue engineering and cell biological applications.     

Hydroxyapatite porous scaffold engineered with biological polymer hybrid coating for antibiotic Vancomycin release

The purpose of this study is to improve hydroxyapatite (HA) porous scaffolds via coating with biological polymer-HA hybrids for use as wound healing and tissue regeneration. Highly porous HA scaffolds, fabricated by a polyurethane foam reticulate method, were coated with hybrid coating solution, consisting of poly(-caprolactone) (PCL), HA powders, and the antibiotic Vancomycin. The PCL to HA ratio was fixed at 1.5 and the drug amounts were varied [drug/(PCL + HA) = 0.02 and 0.04]. For the purpose of comparison, bare HA scaffold without the hybrid coating layer was also loaded with Vancomycin via an immersion-adsorption method. The hybrid coating structure and morphology were observed with Fourier transformed infrared (FT-IR) spectroscopy and scanning electron microscopy (SEM). The effects of the hybrid coating on the compressive mechanical properties and the in vitro drug release of the scaffolds were investigated in comparison with bare HA scaffold. The PCL-HA hybrid coating altered the scaffold pore structure slightly, resulting in thicker stems and reduced porosity. With the hybrid coating, the HA scaffold responded to an applied compressive stress more effectively without showing a brittle failure. This was attributed to the shielding and covering of the framework surface by the coating layer. The encapsulated drugs within the coated scaffold was released in a highly sustained manner as compared to the rapid release of drugs directly adsorbed on the pure HA scaffold. These findings suggest that the coated HA scaffolds expand their applicability in hard tissue regeneration and wound healing substitutes delivering bioactive molecules.     

Bio-functionalized PCL nanofibrous scaffolds for nerve tissue engineering

Surface properties of scaffolds such as hydrophilicity and the presence of functional groups on the surface of scaffolds play a key role in cell adhesion, proliferation and migration. Different modification methods for hydrophilicity improvement and introduction of functional groups on the surface of scaffolds have been carried out on synthetic biodegradable polymers, for tissue engineering applications. In this study, alkaline hydrolysis of poly (ε-caprolactone) (PCL) nanofibrous scaffolds was carried out for different time periods (1 h, 4 h and 12 h) to increase the hydrophilicity of the scaffolds. The formation of reactive groups resulting from alkaline hydrolysis provides opportunities for further surface functionalization of PCL nanofibrous scaffolds. Matrigel was attached covalently on the surface of an optimized 4 h hydrolyzed PCL nanofibrous scaffolds and additionally the fabrication of blended PCL/matrigel nanofibrous scaffolds was carried out. Chemical and mechanical characterization of nanofibrous scaffolds were evaluated using attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy, contact angle, scanning electron microscopy (SEM) and tensile measurement. In vitro cell adhesion and proliferation study was carried out after seeding nerve precursor cells (NPCs) on different scaffolds. Results of cell proliferation assay and SEM studies showed that the covalently functionalized PCL/matrigel nanofibrous scaffolds promote the proliferation and neurite outgrowth of NPCs compared to PCL and hydrolyzed PCL nanofibrous scaffolds, providing suitable substrates for nerve tissue engineering.     

The influence of plasma technology coupled to chemical grafting on the cell growth compliance of 3D hydroxyapatite scaffolds

The development of advanced materials with biomimetic features in order to elicit desired biological responses and to guarantee tissue biocompatibility is recently gaining attention for tissue engineering applications. Bioceramics, such as hydroxyapatite-based biomaterials are now used in a number of different applications throughout the body, covering all areas of the skeleton, due to their biological and chemical similarity to the inorganic phases of bones. When bioactive sintered hydroxyapatite (HA) is desired, biomolecular modification of these materials is needed. In the present work, we investigated the influence of plasma surface modification coupled to chemical grafting on the cell growth compliance of HA 3D scaffolds.     

Poly (lactic-co-glycolic acid) as a controlled release delivery device

Poly (lactic-co-glycolic acid) (PLGA) is a biodegradable polymer used to make resorbable sutures, and is also used in other applications in tissue engineering. Being an artificial polymer, its degradation rate can be tailored to suit its application. It can be easily moulded into structures with suitable mechanical strength and degrades into relatively harmless products in the body. Its adjustable degradation rate also makes it a potentially excellent controlled release delivery device. However, the functionalization of PLGA with bioactive molecules usually requires extensive chemical modification. Chemical modification may compromise the mechanical strength of PLGA and inactivate the bioactive molecules. In this paper, a study is done to investigate the coating of an angiogenic factor on unmodified PLGA suture substrates for the differentiation of human mesenchymal stem cells (hMSC) into endothelial cells (EC). The results show that the method used to anchor vascular endothelial growth factor (VEGF) onto the PLGA surface can enable the gradual release of VEGF from the substrate into solution to induce the differentiation of hMSCs into ECs. Thus, this method can potentially be used to coat PLGA materials like sutures, meshes and scaffolds, rendering them functional as effective controlled release delivery devices for a wide range of bioactive molecules.