Fibrin: a versatile scaffold for tissue engineering applications

Tissue engineering combines cell and molecular biology with materials and mechanical engineering to replace damaged or diseased organs and tissues. Fibrin is a critical blood component responsible for hemostasis, which has been used extensively as a biopolymer scaffold in tissue engineering. In this review we summarize the latest developments in organ and tissue regeneration using fibrin as the scaffold material. Commercially available fibrinogen and thrombin are combined to form a fibrin hydrogel. The incorporation of bioactive peptides and growth factors via a heparin-binding delivery system improves the functionality of fibrin as a scaffold. New technologies such as inkjet printing and magnetically influenced self-assembly can alter the geometry of the fibrin structure into appropriate and predictable forms. Fibrin can be prepared from autologous plasma, and is available as glue or as engineered microbeads. Fibrin alone or in combination with other materials has been used as a biological scaffold for stem or primary cells to regenerate adipose tissue, bone, cardiac tissue, cartilage, liver, nervous tissue, ocular tissue, skin, tendons, and ligaments. Thus, fibrin is a versatile biopolymer, which shows a great potential in tissue regeneration and wound healing.     

Effects of structural properties of electrospun TiO2 nanofiber meshes on their osteogenic potential

Ideal outcomes in the field of tissue engineering and regenerative medicine involve biomaterials that can enhance cell differentiation and production of local factors for natural tissue regeneration without the use of systemic drugs. Biomaterials typically used in tissue engineering applications include polymeric scaffolds that mimic the three-dimensional structural environment of the native tissue, but these are often functionalized with proteins or small peptides to improve their biological performance. For bone applications, titanium implants, or more appropriately the TiO2 passive oxide layer formed on their surface, have been shown to enhance osteoblast differentiation in vitro and to promote osseointegration in vivo. In this study we evaluated the effect on osteoblast differentiation of pure TiO2 nanofiber meshes with different surface microroughness and nanofiber diameters, prepared by the electrospinning method. MG63 cells were seeded on TiO2 meshes, and cell number, differentiation markers and local factor production were analyzed. The results showed that cells grew throughout the entire surfaces and with similar morphology in all groups. Cell number was sensitive to surface microroughness, whereas cell differentiation and local factor production was regulated by both surface roughness and nanofiber diameter. These results indicate that scaffold structural cues alone can be used to drive cell differentiation and create an osteogenic environment without the use of exogenous factors.     

Apatite-coated poly(lactic-co-glycolic acid) microspheres as an injectable scaffold for bone tissue engineering

Biodegradable polymer/ceramic composite scaffold could overcome limitations of biodegradable polymers or ceramics for bone regeneration. Injectable scaffold has raised great interest for bone regeneration in vivo, since it allows one for easy filling of irregularly shaped bone defects and implantation of osteogenic cells through minimally invasive surgical procedures The purpose of this study was to determine whether apatite-coated poly(lactic-co-glycolic acid) (PLGA) microspheres could be used as an injectable scaffold to regenerate bone in vivo. Apatite-coated PLGA microspheres were fabricated by incubating PLGA microspheres in simulated body fluid. The apatite that coated the PLGA microsphere surfaces was similar to apatite in natural bone, as demonstrated by scanning electron microscopy, X-ray diffraction spectra, energy-dispersive spectroscopy, and Fourier transformed-infrared spectroscopy analyses. Rat osteoblasts were mixed with apatite-coated PLGA microspheres and injected immediately into subcutaneous sites of athymic mice. Osteoblast transplantation with plain PLGA microspheres served as a control. Histological analysis of the implants at 6 weeks with hematoxylin and eosin staining, Masson's trichrome staining, and von Kossa staining revealed much better regeneration of bone in the apatite-coated PLGA microsphere group than the plain PLGA microsphere group. The new bone formation area and the calcium content of the implants were significantly higher in the apatite-coated PLGA microsphere group than in the plain PLGA microsphere group. This study demonstrates the feasibility of using apatite-coated PLGA microspheres as an injectable scaffold for in vivo bone tissue engineering. This scaffold may be useful for bone regeneration through minimally invasive surgical procedures in orthopedic applications.     

骨组织工程支架材料的研究进展及临床应用

背景：骨组织工程研究的中心环节是研制能够作为细胞移植与引导新骨生长的支架材料,以作为细胞外基质的替代物。 目的：总结骨组织工程支架材料的研究进展,并展望其发展趋势。 方法：由第一作者检索1995-01-12/2011-01 PubMed数据、中国知网数据库及维普数据库有关人工合成材料及天然支架材料相关文献。英文检索词为“bone tissue engineering,scaffold material,DBM,chitosan,HA,β-TCP”;中文检索词为：“骨组织工程,支架材料,细胞外基质,脱钙骨基质,壳聚糖,羟基磷灰石,β-磷酸三钙”。根据纳入标准排除重复性研究,保留30篇文献进行归纳总结。 结果及结论：支架材料是构建组织工程骨的主体,行使细胞外基质的功能,包括为组织细胞的修复活动提供适宜的微环境,抵挡来自周围组织的压力,用于承重部位暂时行使部分功能等。目前人工合成材料和天然材料各有优缺点,人工合成材料现存问题主要在于材料的细胞黏附率低,难以完全降解,致畸性和致癌性有待明确;天然材料则需要更好地解决材料的抗原性和机械性能的关系,降解速度的调控问题。充分了解现有材料的特点为寻找新方法及新材料提供理论依据。     

Combined use of designed scaffolds and adenoviral gene therapy for skeletal tissue engineering

While tissue engineering remains the most researched alternative to conventional therapies for repair and regeneration, how to optimally combine two of the most promising techniques, designed solid scaffolds and localized gene therapy, is largely unknown. We have conducted a systematic screening of several variables that may affect generation of bone via adenoviral gene therapy vector delivery, on image-based designed and solid freeform-fabricated scaffolds. These variables included: gene therapy type (ex vivo or in vivo); scaffold base material (sintered hydroxyapatite or a polypropylene fumarate/ tricalcium phosphate (PPF/TCP) composite), secondary carrier used to attach the biofactor to the scaffold (fibrin gel or a poly-lactic acid sponge), and scaffold pores size (300 or 800 microm). The in vivo formation of bone following implantation of these scaffolds was then analyzed. Gene therapy method had the largest effect, with ex vivo gene therapy yielding significant amounts of bone on nearly all the implants and in vivo gene therapy failing to produce any bone on most implants. Secondary carrier was the next most important variable, with fibrin gel consistently producing bone encompassing the implants and producing 2-4 times as much bone as the polymer sponge, which triggered only isolated bone growth. Though both scaffold base materials allowed bone growth, hydoxyapatite scaffolds generated twice as much bone as PPF/TCP scaffolds. The pore sizes tested had no significant effect on tissue generation.     

Scaffolds in tissue engineering bone and cartilage

Musculoskeletal tissue, bone and cartilage are under extensive investigation in tissue engineering research. A number of biodegradable and bioresorbable materials, as well as scaffold designs, have been experimentally and/or clinically studied. Ideally, a scaffold should have the following characteristics: (i) three-dimensional and highly porous with an interconnected pore network for cell growth and flow transport of nutrients and metabolic waste; (ii) biocompatible and bioresorbable with a controllable degradation and resorption rate to match cell/tissue growth in vitro and/or in vivo; (iii) suitable surface chemistry for cell attachment, proliferation, and differentation and (iv) mechanical properties to match those of the tissues at the site of implantation. This paper reviews research on the tissue engineering of bone and cartilage from the polymeric scaffold point of view.     

The effects of PRGF on bone regeneration and on titanium implant osseointegration in goats: a histologic and histomorphometric study

The effect of local application of scaffold-like preparation rich in growth factors (PRGF) on bone regeneration in artificial defects and the potential effect of humidifying titanium dental implants with liquid PRGF on their osseointegration were investigated. The PRGF formulations were obtained from venous blood of three goats and applied either as a 3D fibrin scaffold (scaffold-like PRGF) in the regeneration of artificial defects or as liquid PRGF via humidifying the implants before their insertion. Initially, 12 defects were filled with scaffold-like PRGF and another 12 were used as controls. The histological analysis at 8 weeks revealed mature bone trabeculae when PRGF was used, whereas the control samples showed mainly connective tissue with incipient signs of bone formation. For the second set of experiments, 26 implants (13 humidified with liquid PRGF) were placed in the tibiae of goats. Histological and histomorphometric results demonstrated that application of liquid PRGF increased the percentage of bone-implant contact in 84.7%. The whole surface of the PRGF-treated implants was covered by newly formed bone, whereas only the upper half was surrounded in control implants. In summary, PRGF can accelerate bone regeneration in artificial defects and improve the osseointegration of titanium dental implants.     

组织工程支架材料在脊髓损伤修复中的应用

背景：脊髓损伤最初往往会导致细胞和组织的不断丢失,组织工程支架可以模拟细胞外基质的生理状态,从而有利于细胞的黏附、迁移、扩增和分化。 目的：总结近年来组织工程支架材料联合细胞和/或细胞因子修复脊髓损伤的新进展。 方法：应用计算机检索PubMed、Ovid Medline及CBM数据库中2000-10/2010-10 与组织工程支架材料修复脊髓损伤相关的文章。  结果与结论：组织工程材料治疗脊髓损伤需要3 因素：种子细胞、组织工程支架、细胞因子。组织工程支架对于损伤脊髓断端起到桥接作用,而种植于材料的种子细胞和/或细胞因子可以促进神经轴突的生长和迁移。可用于组织工程支架的材料可分为天然材料和人工合成材料,包括胶原、壳聚糖、琼脂糖/藻酸盐、聚乳酸、纤连蛋白、聚羟基乙酸/聚乳酸、聚β羟丁酸等,动物实验已经取得一些成果,显示组织工程支架材料联合细胞移植修复效果更好,但临床上目前尚无开展组织工程支架材料修复脊髓损伤的研究。     

Applications of gene therapy and adult stem cells in bone bioengineering

Bone tissue engineering is an emerging field, that could become a main therapeutic strategy in orthopedics in coming years. While bone has regenerative abilities that enable the self repair and regeneration of fractures, there are extreme situations in which the extent of bone loss is too large for complete regeneration to occur. In order to achieve bone regeneration, osteogenic genes (mainly from the bone morphogenetic protein family) can be delivered either directly into the target tissue, or by using adult stem cells, which are later implanted into the target site. Engineered adult stem cells combined with biodegradable polymeric scaffolds can be implanted into target sites, with or without ex vivo culture period. Several important factors influence the success of bone engineering approaches including: choice of cell and scaffold, the vector used in order to deliver the osteogenic gene, and the osteogenic gene itself. Cutting-edge imaging technologies, bioinformatics-based analysis of gene expression and exogenous regulation of transgene expression are among the tools that are being used to optimize and control bone formation in vivo. In this review we have attempted to provide an overview of the main factors that should be considered when utilizing adult stem cells and gene therapy strategies to regenerate bone defects or to promote new bone formation in vivo.     

Collagen for bone tissue regeneration

In the last decades, increased knowledge about the organization, structure and properties of collagen (particularly concerning interactions between cells and collagen-based materials) has inspired scientists and engineers to design innovative collagen-based biomaterials and to develop novel tissue-engineering products. The design of resorbable collagen-based medical implants requires understanding the tissue/organ anatomy and biological function as well as the role of collagen's physicochemical properties and structure in tissue/organ regeneration. Bone is a complex tissue that plays a critical role in diverse metabolic processes mediated by calcium delivery as well as in hematopoiesis whilst maintaining skeleton strength. A wide variety of collagen-based scaffolds have been proposed for different tissue engineering applications. These scaffolds are designed to promote a biological response, such as cell interaction, and to work as artificial biomimetic extracellular matrices that guide tissue regeneration. This paper critically reviews the current understanding of the complex hierarchical structure and properties of native collagen molecules, and describes the scientific challenge of manufacturing collagen-based materials with suitable properties and shapes for specific biomedical applications, with special emphasis on bone tissue engineering. The analysis of the state of the art in the field reveals the presence of innovative techniques for scaffold and material manufacturing that are currently opening the way to the preparation of biomimetic substrates that modulate cell interaction for improved substitution, restoration, retention or enhancement of bone tissue function.     

Sustained release of vascular endothelial growth factor from mineralized poly(lactide-co-glycolide) scaffolds for tissue engineering

Strategies to engineer bone tissue have focused on either: (1) the use of scaffolds for osteogenic cell transplantation or as conductive substrates for guided bone regeneration; or (2) release of inductive bioactive factors from these scaffold materials. This study describes an approach to add an inductive component to an osteoconductive scaffold for bone tissue engineering. We report the release of bioactive vascular endothelial growth factor (VEGF) from a mineralized, porous, degradable polymer scaffold. Three dimensional, porous scaffolds of the copolymer 85 : 15 poly(lactide-co-glycolide) were fabricated by including the growth factor into a gas foaming/particulate leaching process. The scaffold was then mineralized via incubation in a simulated body fluid. Growth of a bone-like mineral film on the inner pore surfaces of the porous scaffold is confirmed by mass increase measurements and quantification of phosphate content within scaffolds. Release of ¹²⁵I-labeled VEGF was tracked over a 15 day period to determine release kinetics from the mineralized scaffolds. Sustained release from the mineralized scaffolds was achieved, and growth of the mineral film had only a minor effect on the release kinetics from the scaffolds. The VEGF released from the mineralized and non-mineralized scaffolds was over 70% active for up to 12 days following mineralization treatment, and the growth of mineral had little effect on total scaffold porosity.     

Influence of scaffold design on host immune and stem cell responses

The combined culture of isolated stem cells in tissue engineering scaffolds represents a popular strategy for the regeneration of specialized tissues. Despite of improved outcomes in some tissues, this stem cell-seeded tissue engineering strategy has not led to significant tissue regeneration as expected. The lower-than-expected outcome may be caused by overwhelming immune responses to scaffold materials and poor survival of seeded stem cells following implantation. This review is aimed at summarizing the success and failure of this strategy and also shedding some light on new directions to design scaffolds for promoting regenerative responses via autologous stem cells. The first half of this review summarizes the influence of scaffold physical and chemical properties on immune cell responses to scaffold implants. The second half focuses on the influence of scaffold design to alter immune and stem cell responses for achieving desirable tissue regeneration.     

Tissue regeneration based on growth factor release

Tissue engineering is an emerging biomedical field intended to assist the regeneration of body tissue defects too large to self-repair as well as to substitute for the biological functions of damaged and injured organs by using cells with proliferative and differentiative potential. In addition to basic research on such cells, it is undoubtedly indispensable for successful tissue engineering to create an artificial environment enabling cells to induce tissue regeneration. Such an environment can be achieved by making use of a scaffold for cell proliferation and differentiation and for growth factors, as well as their combination. Growth factors are often required to promote tissue regeneration, as they can induce angiogenesis, which supplies oxygen and nutrients to cells transplanted for organ substitution to maintain their biological functions. However, the biological effects of growth factors cannot always be expected because of their poor in vivo stability, unless a drug delivery system is contrived. In this article, tissue regeneration based on the release of growth factors is reviewed to emphasize the significance of drug delivery systems in tissue engineering.     

In vitro localization of bone growth factors in constructs of biodegradable scaffolds seeded with marrow stromal cells and cultured in a flow perfusion bioreactor

Tissue engineering strategies aim at controlling the behavior of individual cells to stimulate tissue formation. This control is achieved by mimicking signals that manage natural tissue development or repair. Flow perfusion bioreactors that create culture environments with minimal diffusion constraints and provide cells with mechanical stimulation may closely resemble in vivo conditions for bone formation. Therefore, these culturing systems, in conjunction with an appropriate scaffold and cell type, may provide significant insight towards the development of in vitro tissue engineering models leading to improved strategies for the construction of bone tissue substitutes. The objective of this study was to investigate the in vitro localization of several bone growth factors that are usually associated with bone formation in vivo by culturing rat bone marrow stromal cells seeded onto starch-based biodegradable fiber meshes in a flow perfusion bioreactor. The localization of several bone-related growth factors-namely, transforming growth factor-beta1, platelet-derived growth factor- A, fibroblast growth factor-2, vascular endothelial growth factor, and bone morphogenetic protein- 2-was determined at two different time points in scaffolds cultured under perfusion conditions at two different flow rates using an immunohistochemistry technique. The results show the presence of regions positively stained for all the growth factors considered, except platelet-derived growth factor-A. Furthermore, the images obtained from the positively stained sections suggest an increase in the immunohistochemically stained area at the higher flow rate and culture time. These observations demonstrate that flow perfusion augments the functionality of scaffold/cell constructs grown in vitro as it combines both biological and mechanical factors to enhance cell differentiation and cell organization within the construct. This study also shows that flow perfusion bioreactor culture of marrow stromal cells, combined with the use of appropriate biodegradable fiber meshes, may constitute a useful model to study bone formation and assess bone tissue engineering strategies in vitro.     

Biomedical applications of polyurethanes: a review of past promises, present realities, and a vibrant future.

Polyurethanes, having extensive structure/property diversity, are one of the most bio- and blood-compatible materials known today. These materials played a major role in the development of many medical devices ranging from catheters to total artificial heart. Properties such as durability, elasticity, elastomer-like character, fatigue resistance, compliance, and acceptance or tolerance in the body during the healing, became often associated with polyurethanes. Furthermore, propensity for bulk and surface modification via hydrophilic/hydrophobic balance or by attachments of biologically active species such as anticoagulants or biorecognizable groups are possible via chemical groups typical for polyurethane structure. These modifications are designed to mediate and enhance the acceptance and healing of the device or implant. Many innovative processing technologies are used to fabricate functional devices, feeling and often behaving like natural tissue. The hydrolytically unstable polyester polyurethanes were replaced by more resistant but oxidation-sensitive polyether polyols based polyurethanes and their clones containing silicone and other modifying polymeric intermediates. Chronic in vivo instability, however, observed on prolonged implantation, became a major roadblock for many applications. Presently, utilization of more oxidation resistant polycarbonate polyols as soft segments, in combination with antioxidants such as Vitamin E, offer materials which can endure in the body for several years. The applications cover cardiovascular devices, artificial organs, tissue replacement and augmentation, performance enhancing coatings and many others. In situ polymerized, cross-linked systems could extend this biodurability even further. The future will expand this field by revisiting chemically-controlled biodegradation, in combination with a mini-version of RIM technology and minimally invasive surgical procedures, to form, in vivo, a scaffold, by delivery of reacting materials to the specific site in the body and polymerizing the mass in situ. This scaffold will provide anchor for tissue regeneration via cell attachment, proliferation, control of inflammation, and healing.     

Biomimetic nanofibrous scaffolds for bone tissue engineering

Bone tissue engineering is a highly interdisciplinary field that seeks to tackle the most challenging bone-related clinical issues. The major components of bone tissue engineering are the scaffold, cells, and growth factors. This review will focus on the scaffold and recent advancements in developing scaffolds that can mimic the natural extracellular matrix of bone. Specifically, these novel scaffolds mirror the nanofibrous collagen network that comprises the majority of the non-mineral portion of bone matrix. Using two main fabrication techniques, electrospinning and thermally-induced phase separation, and incorporating bone-like minerals, such as hydroxyapatite, composite nanofibrous scaffolds can improve cell adhesion, stem cell differentiation, and tissue formation. This review will cover the two main processing techniques and how they are being applied to fabricate scaffolds for bone tissue engineering. It will then cover how these scaffolds can enhance the osteogenic capabilities of a variety of cell types and survey the ability of the constructs to support the growth of clinically relevant bone tissue.     

骨组织工程支架材料的研究现状与应用前景

背景：目前组织工程骨修复骨缺损在临床应用中较为关键的问题是建立血管网,为新骨的形成提供氧气及营养物质,并为机体提供代谢途径。 目的：综述近年组织工程骨支架材料的特点,并着重介绍复合支架材料的研究现状。 方法：以“骨组织工程,血管化,支架材料,复合支架材料”为中文检索词,以“bone tissue engineering, vascularization,scaffold,composite scaffold”英文检索词,应用计算机在中国期刊全文数据库和PubMed数据库检索2001年1月至2014年1月的相关文章,将所有文章进行初步筛选后,对保留的文章进一步详细分析、归纳并总结。 结果与结论：按照组织工程骨支架材料的来源不同,可将其分为人工合成材料、天然衍生材料和复合支架材料,单一支架材料难以作为最理想的材料修复骨缺损,复合支架材料能在不同程度上弥补单一支架材料的缺陷,因此近年来组织工程支架材料的发展由单一材料发展为复合材料,并呈现人工合成材料与天然材料有机结合的趋势。但复合支架材料在临床应用中仍然有许多尚待解决的问题,主要有控制复合材料比例,使材料降解速率与组织细胞的生长速率相适应,保持复合材料的多孔隙和高机械强度。中国组织工程研究杂志出版内容重点：生物材料;骨生物材料; 口腔生物材料; 纳米材料; 缓释材料; 材料相容性;组织工程全文链接：     

A three-phase, fully resorbable, polyester/calcium phosphate scaffold for bone tissue engineering: Evolution of scaffold design

Bone tissue engineering strategies are fundamentally based upon porous scaffold materials that serve as a support for ingrowth of host cells and/or provide a substrate for exogenously delivered cells. Here we report the application of a surface calcium phosphate (CaP) mineral layer to a macroporous polymeric/CaP composite biomaterial, with a macroporous interconnectivity, and its subsequent in vivo evaluation in a rodent femoral defect. The application of the mineral layer eliminates the fibrous tissue encapsulation and foreign body giant cell response commonly seen at the interface of polymeric materials, yet retains the unique characteristics of the parent material as being macroporous, completely biodegradable and possessing a high degree of interconnectivity. This represents the third generation of this scaffold material, incorporating iterative changes to the scaffold design in response to both materials and biological design criteria to produce a material with enhanced in vitro and in vivo performance.     

Nonwoven membranes for tissue engineering: an overview of cartilage,epithelium, and bone regeneration

Scaffold-type biomaterials are crucial for application in tissue engineering. Among them, the use of a nonwoven scaffold has grown in recent years and has been widely investigated for the regeneration of different types of tissues. Several polymers, whether they are synthetic, biopolymers or both, have been used to produce a scaffold that can mimic the natural tissue to which it will be applied to. The scaffolds used in tissue engineering must be biocompatible and allow cell adhesion and proliferation to be applied in tissue engineering. In addition, the scaffolds should maintain the mechanical properties and architecture of the desired tissue. Nonwoven fabrics have produced good results and are more extensively applied for the regeneration of cartilage, epithelial and bone tissues. Recent advances in tissue engineering have shown promising results, however, no ideal material or standardization parameters and characteristics of the materials were obtained. The present review provides an overview of the application of nonwoven scaffolds, including the main results obtained regarding the properties of the biomaterials and their applications in vitro and in vivo, focusing on the cartilaginous, the epithelium, and bone tissue regeneration.