Bone tissue reactions to biomimetic ion-substituted apatite surfaces on titanium implants

The aim of this study was to evaluate the bone tissue response to strontium- and silicon-substituted apatite (Sr-HA and Si-HA) modified titanium (Ti) implants. Sr-HA, Si-HA and HA were grown on thermally oxidized Ti implants by a biomimetic process. Oxidized implants were used as controls. Surface properties, i.e. chemical composition, surface thickness, morphology/pore characteristics, crystal structure and roughness, were characterized with various analytical techniques. The implants were inserted in rat tibiae and block biopsies were prepared for histology, histomorphometry and scanning electron microscopy analysis. Histologically, new bone formed on all implant surfaces. The bone was deposited directly onto the Sr-HA and Si-HA implants without any intervening soft tissue. The statistical analysis showed significant higher amount of bone–implant contact (BIC) for the Si-doped HA modification (P = 0.030), whereas significant higher bone area (BA) for the Sr-doped HA modification (P = 0.034), when compared with the non-doped HA modification. The differences were most pronounced at the early time point. The healing time had a significant impact for both BA and BIC (P < 0.001). The present results show that biomimetically prepared Si-HA and Sr-HA on Ti implants provided bioactivity and promoted early bone formation.     

Biomimetic poly(glycerol sebacate) (PGS) membranes for cardiac patch application

In this study biomimetic poly(glycerol sebacate) PGS matrix was developed for cardiac patch application. The rationale was that such matrices would provide conducive environment for the seeded cells at the interphase with PGS. From the microstructural standpoint, PGS was fabricated into dense films and porous PGS scaffolds. From the biological aspect, biomimetic PGS membranes were developed via covalently binding peptides Tyr-Ile-Gly-Ser-Arg (YIGSR) and Gly-Arg-Gly-Asp-Ser-Pro (GRGDSP), corresponding to the epitope sequences of laminin and fibronectin, respectively onto the surface. To improve and enhance homogenous binding of peptides onto the PGS surface, chemical modification of its surface was carried out. A sequential regime of alkaline hydrolysis with 0.01 M NaOH for 5 min and acidification with 0.01 M HCl for 25 s was optimal. More COOH chemical group was exposed without causing deleterious effect on the bulk properties of the polymer as revealed by the physicochemical analysis carried out. HPLC analysis, chemical imaging and ToF-SIMS were able to establish the successful homogenous functionalization of PGS membranes with the peptides. Finally, the developed biomimetic membranes supported the adhesion and growth of rat and human cardiac progenitor cells.     

Biomechanical evaluation of cell-loaded and cell-free hydroxyapatite implants for the reconstruction of segmental bone defects

Porous hydroxyapatite (HA) scaffoldings are currently used in tissue engineering for bone reconstruction. When this osteoconductive biomaterial is combined with osteoprogenitor cells, it acquires osteoinductive features which accelerate and improve bone formation in vivo. The aim of our study was to assess the mechanical properties of HA–bone complexes undergoing indentation tests, and relate stiffness to composition and structure as examined by micro X-ray. To this purpose, 35-mm tibia diaphyseal resections were performed in sheep. Gaps were filled using porous HA cylinders. Implants were loaded with autologous bone marrow stromal cells (BMSC); cell-free cylinders were used as control. After 8 weeks, bone tissue was found within the internal macropores of cell-loaded HA carriers, and in control implants, bone formation was mostly limited to the outer surface. As assessed by indentation testing the stiffness values of bone–HA composites were halfway between those of HA scaffoldings and tibia bone. Cell-loaded implants were stiffer than cell-free ones. In a cell-loaded implant we also analyzed the variation of stiffness along the main axis of the tibia.     

羟基磷灰石球形颗粒表面微形貌构建及其对干细胞生物学行为的调控

通过调节溶胶-凝胶体系中羟基磷灰石(HA)粉末和甲壳素(Chitin)的质量比, 制备具有不同表面微形貌的HA球形颗粒。扫描电子显微镜(SEM)表征结果显示: 随着HA/Chitin质量比从4/1增加到35/1, 球形颗粒的表面微形貌发生了明显变化, 由粗糙渐趋平滑, 微米级皱褶逐渐减少至消失, 微孔隙率从(35%±0.8%)减少到(10.4%±0.7%)。体外细胞培养的结果表明具有微米级皱褶, 微孔隙率较高的粗糙表面具有引导干细胞铺展和增殖的作用, 微孔隙率低的平滑表面则具有引导干细胞轴向延伸及骨向分化的趋势。同时, HA球形颗粒表面微形貌对干细胞表面特征性抗原标志物的表达具有调控作用。     

Novel nano-composite biomimetic biomaterial allows chondrogenic and osteogenic differentiation of bone marrow concentrate derived cells

In clinical orthopedics suitable materials that induce and restore biological functions together with the right mechanical properties are particularly needed for the regeneration of osteochondral lesions. For this purpose, the ideal scaffold should possess the right properties with respect to degradation, cell binding, cellular uptake, non-immunogenicity, mechanical strength, and flexibility. In addition, it should be easy to handle and serve as a template for chondrocyte and bone cells guiding both cartilage and bone formation. The aim of the present study was to estimate the chondrogenic and osteogenic capability of bone marrow concentrated derived cells seeded onto a novel nano-composite biomimetic material. These properties have been evaluated by means of histological, immunohistochemical and electron microscopy analyses. The data obtained demonstrated that freshly harvested cells obtained from bone marrow were able, once seeded onto the biomaterial, to differentiate either down the chondrogenic and osteogenic pathways as evaluated by the expression and production of specific matrix molecules. These findings support the use, for the repair of osteochondral lesions, of this new nano-composite biomimetic material together with bone marrow derived cells in a “one step” transplantation procedure.     

生物材料细胞粘附肽仿生修饰的研究进展

RGD(Arg-G1y-Asp)短肽序列是一种细胞粘附肽,能被细胞膜上的整合素识别,参与细胞与基质间的粘附.为改善合成生物材料缺乏细胞识别信号的缺点,可将含RGD序列肽经本体或表面修饰引入材料,使材料具有良好的细胞亲和性.综述了采用含RGD肽对各种合成生物材料进行仿生修饰的研究进展.     

SLA表面PDA/CPP复合涂层的制备及其性能研究

在大颗粒喷砂酸蚀(SLA)工艺形成的三维多级嵌套孔洞结构纯钛表面, 利用聚多巴胺(PDA)的超强粘附性和二次修饰功能, 接枝生物大分子酪蛋白磷酸肽(CPP), 制备PDA/CPP复合涂层。采用FESEM、EDS、XPS、水接触角分析仪、体外仿生矿化测试以及人骨髓间充质干细胞(hBMSCs)实验对样品表面进行表征分析和性能测试。结果表明, 两步浸泡法成功在SLA表面制得PDA/CPP复合涂层。该涂层在保留SLA多孔形貌的同时, 使水接触角从47.7°减小到25.5°, 明显提高了SLA表面亲水性; 在模拟体液浸泡1 d后的修饰表面即沉积了致密羟基磷灰石(HA), 说明PDA/CPP显著加快了SLA表面生物矿化的速度。同时, SLA/PDA/CPP表面细胞粘附、增殖以及矿化活性都得到了明显改善。     

Biomimetic approach to the formation of gold nanoparticle/silica core/shell structures and subsequent bioconjugation

The encapsulation of individual nanoparticles has gained great attention as a method for both stabilizing nanoparticles and tailoring their surface properties. In particular, the encapsulation of nanoparticles with silica shells is advantageous for bioconjugation and applications to (nano)biotechnology. Herein we report a method for constructing gold nanoparticle (AuNP)/silica core/shell hybrid structures by biomimetic silicification of silicic acids. The procedure consists of surface-initiated, atom transfer radical polymerization of 2-(dimethylamino)ethyl methacrylate (DMAEMA) from AuNPs and biomimetic polycondensation of silicic acids by using poly(DMAEMA) as a synthetic counterpart for silaffins that are found in diatoms. The resulting AuNP/silica hybrids were characterized by Fourier transform infrared spectroscopy, energy dispersive x-ray spectroscopy, UV-vis spectroscopy and transmission electron microscopy. In addition, the immobilization of biological ligands onto the hybrids was investigated for potential applications to biotechnology. As a model ligand, biotin was attached onto the AuNP/silica hybrids through substitution reaction and Michael addition reaction, and the attachment was confirmed by fluorescence microscopy after complexation with fluorescein-conjugated streptavidin.     

Enzyme-functionalized biomimetic apatites: concept and perspectives in view of innovative medical approaches

Biomimetic nanocrystalline calcium-deficient apatite compounds are particularly attractive for the setup of bioactive bone-repair scaffolds due to their high similarity to bone mineral in terms of chemical composition, structural and substructural features. As such, along with the increasingly appealing development of moderate temperature engineered routes for sample processing, they have widened the armamentarium of orthopedic and maxillofacial surgeons in the field of bone tissue engineering. This was made possible by exploiting the exceptional surface reactivity of biomimetic apatite nanocrystals, capable of easily exchanging ions or adsorbing (bio)molecules, thus leading to highly-versatile drug delivery systems. In this contribution we focus on the preparation of hybrid materials combining biomimetic nanocrystalline apatites and enzymes (lysozyme and subtilisin). This paper reports physico-chemical data as well as cytotoxicity evaluations towards Cal-72 osteoblast-like cells and finally antimicrobial assessments towards selected strains of interest in bone surgery. Biomimetic apatite/enzyme hybrids could be prepared in varying buffers. They were found to be non-cytotoxic toward osteoblastic cells and the enzymes retained their biological activity (e.g. bond cleavage or antibacterial properties) despite the immobilization and drying processes. Release properties were also examined. Beyond these illustrative examples, the concept of biomimetic apatites functionalized with enzymes is thus shown to be useable in practice, e.g. for antimicrobial purposes, thus widening possible therapeutic perspectives.     

Effect of expanded bone marrow-derived osteoprogenitor cells seeded into polycaprolactone/tricalcium phosphate scaffolds in new bone regeneration of rabbit mandibular defects

The purpose of this study was to assess and evaluate new bone formation in rabbit marginal mandibular defects using expanded bone marrow-derived osteoprogenitor cells seeded in three-dimensional scaffolds of polycaprolactone/tricalcium phosphate (PCL/TCP). Bone marrow was harvested from the rabbit ilium and rabbit bone marrow-derived osteoprogenitor cells were isolated and expanded in standard culture medium and osteogenic medium supplement. The cells were then seeded into the PCL/TCP scaffolds and the cell/scaffold constructions were implanted into prepared defects in rabbit mandibles. PCL/TCP scaffold alone and autogenous bone graft from the mandible were also implanted into the other prepared defects. The specimens were evaluated at 4 and 8 weeks after the implantation using clinical, radiographic, and histological techniques. The results of the experimental group demonstrated more newly formed bone on the surface and in the pores of the PCL/TCP scaffolds. In addition, the osteoblasts, osteocytes, and new bone trabeculae were identified throughout the defects that were implanted with the cell/scaffold constructions. The PCL/TCP alone group was filled mostly with fibrous cells particularly in the middle region with less bone formation. These results would suggest that the derived osteotoprogenitor cells have the potential to form bone tissue when seeded onto PCL/TCP scaffolds.     

Biological behavior of sol-gel coated dental implants

The biocompatibility of dental implants coated with titania/hydroxyapatite (HA) and titania/bioactive glass (BG) composites obtained via sol-gel process was investigated using an in vitro and in vivo model. A device for the in vitro testing of screw-shaped dental implants was developed, in order to well compare the two experimental models studying the behavior of human MG63 osteoblast-like cells seeded onto a particular geometry. The expression of some biochemical parameters of osteoblastic phenotype (alkaline phosphatase specific activity, collagen and osteocalcin production) and some indications on cells morphology obtained by scanning electron microscopy were evaluated. The in vitro and in vivo models were compared after implants insertion in rabbit tibia and femur. The removal torque and histomorphometric parameters (percentage of bone in contact with implant surface and the amount of bone inside the threaded area) were examined. A good agreement was found between the in vitro and in vivo models. These experiments showed better performances of HA and BG sol-gel coated dental implants with respect to uncoated titanium; in particular, it was found that in vitro the HA coating stimulates osteoblastic cells in producing higher level of ALP and collagen, whereas in vivo this surface modification resulted in a higher removal torque and a larger bone-implant contact area. This behavior could be ascribed to the morphology and the chemical composition of the implants with rough and bioactive surfaces.     

Preparation and tribological studies of self-assembled triple-layer films

A self-assembled triple-layer film was grafted onto a silicon surface with a simple three-step method. Firstly, 3-glycidoxypropyltrimethoxysilane molecules were self-assembled on silicon surfaces, then coupled to 3-aminopropyltriethoxysilane through a surface ring-opening reaction, and finally octadecyltrichlorosilane (OTS) molecules were attached to the resultant alkoxysilane-terminated surface via Si-O-Si bonds. The structure and morphology of this triple-layer film were characterized with various techniques, such as contact angle measurement, ellipsometry, X-ray photoelectron spectroscopy, and atomic force microscopy (AFM). The influence of different surface chemical groups on surface adhesion properties was identified using the AFM force-volume technique. The micro- and macro-tribological properties of the triple-layer film were evaluated by friction force microscopy and a ball-on-plate tribometer. The triple-layer film shows good adhesive resistance and can greatly reduce the micro- and macro-friction force. Moreover, compared to self-assembled monolayer of OTS, this triple-layer film exhibited much better wear-resistance. This improvement was mainly ascribed to the network structure of a lateral cross-linked polysiloxane layer formed within the film which can enhance the stability of the film.     

PLGA/TSF静电纺纳米纤维的仿生矿化及生物相容性

利用静电纺丝和模拟体液仿生矿化技术制备了聚乳酸-羟基乙酸共聚物/柞蚕丝素/羟基磷灰石((PLGA/TSF/HA)骨组织工程复合支架。通过扫描电子显微镜(SEM)、透射电子显微镜(TEM)、X射线衍射测试(XRD)和热重分析(TG)对复合纳米纤维的形貌结构进行了表征。此外,在复合纳米纤维支架材料上接种人骨髓间充质干细胞(hMSCs),通过四甲基偶氮噻唑蓝比色(Four methyl azo thiazole blue colorimetric,MTT)法,观察细胞在材料表面的生长情况评价纳米纤维的生物相容性。结果显示,PLGA/TSF纳米纤维毡具有精细的三维结构,纤维直径分布均匀,表面光滑。矿化后HA颗粒均匀地分布在PLGA/TSF纳米纤维表面,矿物含量约占63%。与PLGA/TSF纳米纤维支架相比,PLGA/TSF/HA纳米纤维支架的亲水性、生物相容性都得到显著提高。     

Biomimetic properties of an injectable chitosan/nano-hydroxyapatite/collagen composite

To meet the challenges of designing an injectable scaffold and regenerating bone with complex three-dimensional (3D) structures, a biomimetic and injectable hydrogel scaffold based on nano-hydroxyapatite (HA), collagen (Col) and chitosan (Chi) is synthesized. The chitosan/nano-hydroxyapatite/collagen (Chi/HA/Col) solution rapidly forms a stable gel at body temperature. It shows some features of natural bone both in main composition and microstructure. The Chi/HA/Col system can be expected as a candidate for workable systemic minimally invasive scaffolds with surface properties similar to physiological bone based on scanning electron microscopic (SEM), transmission electron microscopy (TEM), X-ray photoelectron spectroscopy (XPS), X-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FT-IR) results.     

Formation of bone-like apatite layer on chitosan fiber mesh scaffolds by a biomimetic spraying process

Bone-like apatite coating of polymeric substrates by means of biomimetic process is a possible way to enhance the bone bonding ability of the materials. The created apatite layer is believed to have an ability to provide a favorable environment for osteoblasts or osteoprogenitor cells. The purpose of this study is to obtain bone-like apatite layer onto chitosan fiber mesh tissue engineering scaffolds, by means of using a simple biomimetic coating process and to determine the influence of this coating on osteoblastic cell responses. Chitosan fiber mesh scaffolds produced by a previously described wet spinning methodology were initially wet with a Bioglass((R))-water suspension by means of a spraying methodology and then immersed in a simulated body fluid (SBF) mimicking physiological conditions for one week. The formation of apatite layer was observed morphologically by scanning electron microscopy (SEM). As a result of the use of the novel spraying methodology, a fine coating could also be observed penetrating into the pores, that is clearly within the bulk of the scaffolds. Fourier Transform Infrared spectroscopy (FTIR-ATR), Electron Dispersive Spectroscopy (EDS) and X-ray diffraction (XRD) analysis also confirmed the presence of apatite-like layer. A human osteoblast-like cell line (SaOs-2) was used for the direct cell contact assays. After 2 weeks of culture, samples were observed under the SEM. When compared to the control samples (unmodified chitosan fiber mesh scaffolds) the cell population was found to be higher in the Ca-P biomimetic coated scaffolds, which indicates that the levels of cell proliferation on this kind of scaffolds could be enhanced. Furthermore, it was also observed that the cells seeded in the Ca-P coated scaffolds have a more spread and flat morphology, which reveals an improvement on the cell adhesion patterns, phenomena that are always important in processes such as osteoconduction.     

Hyrdoxyapatite Coatings on Titanium

Some ceramics, including calcium phosphates and certain glasses and glass‐ceramics, form an important class of bioactive materials that are used extensively in repair and reconstruction of diseased or damaged parts of the musculoskeletal system. The similarity between synthetic hydroxyapatite (HA) (Ca₁₀(PO₄)₆(OH)₂) and the mineral phase of bone and tooth made HA one of the earliest materials to be used to impart osteoconductive (bone‐bonding) properties to the surface of biocompatible, metallic implants. HA and other calcium phosphate surfaces are used primarily to cause early stabilization of the implant in the surrounding bone through the formation of a direct bond and enhanced bone apposition. Although bioactive ceramics are used in several compositions and forms, the focus here is on the use of HA coatings in metallic implants. The coatings made by commercially available plasma‐spray and other techniques are compared to those made recently by surface‐mediated biomimetic routes.     

双键超支化聚硅氧烷在芳纶纤维表面的原位生长

以显著提升芳纶纤维与橡胶基体之间的界面结合强度为目标,本文开发了一种在芳纶纤维表面原位生长碳碳双键超支化聚硅氧烷的策略:首先借助多巴胺的仿生修饰建立初始层,接着采用迈克尔加成反应接枝氨基硅烷偶联剂创建中间层,最后通过脱水缩合反应实现原位生长。改性前后的芳纶纤维表面形貌和化学结构采用扫描电镜、傅里叶变换红外线光谱分析仪、X射线光电子能谱分析仪及热失重分析仪进行了测试表征。由芳纶纤维增强橡胶基复合材料的H-抽出力测试结果可得,本文采用的改性方法能够提升约66.4%的界面结合强度,其原因主要可以归结为表面粗糙度的提升及纤维束与橡胶共硫化的实现。对比分析发现,聚多巴胺初始层的采用使本改性方法具有较高的普适性和工业应用潜力。      

仿生矿化法制备可降解羟基磷灰石/氧化细菌纤维素复合材料

细菌纤维素是具有天然纳米网状结构的支架材料,对其进行氧化改性后可获得可调控的降解性能。通过仿生矿化氧化改性的细菌纤维素支架,制备了可降解羟基磷灰石/氧化细菌纤维素复合骨组织工程支架材料。观察并分析了仿生矿化过程氧化细菌纤维素的降解和羟基磷灰石的形成,并通过SEM、EDS、XRD对羟基磷灰石在可降解氧化细菌纤维素支架上沉积进行了表征,矿化7天的羟基磷灰石/氧化细菌纤维素复合材料表面和内部均有磷灰石形成,测得磷灰石的钙磷比为1.75,主要为羟基磷灰石,伴有少量碳羟磷灰石。结果表明,使用仿生矿化法成功获得了一种新型可降解羟基磷灰石/氧化纤维素复合材料支架。     

Improved mechanical properties of HIPS/hydroxyapatite composites by surface modification of hydroxyapatite via in-situ polymerization of styrene

High impact polystyrene (HIPS)/hydroxyapatite (HA) composites are potential biomaterials for bone replacements due to their good biocompatibility and adequate mechanical properties. At the present work, the surface of the micron-sized hydroxyapatite (HA) particles was modified by in situ polymerization of styrene (St), then compounded with HIPS. The effect of the modification of HA surface on morphology and mechanical properties of HIPS/HA composites were investigated. The results showed that the HA particles does not inhibit the polymerization of St. The PS segments coated on the HA surface by in situ polymerization of St enhances the compatibility between HA and HIPS, improves the dispersion of HA particles in HIPS matrix, and enhances the interfacial adhesion between HA and matrix. Thereby, the stiffness, tensile strength and notch impact strength of HIPS/HA composites are improved at the same time. And there is a critical coating thickness of PS on the HA surface for the optimum mechanical properties of HIPS/HA composites.     

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.