Composite scaffolds for cartilage tissue engineering

Tissue engineering remains a promising therapeutic strategy for the repair or regeneration of diseased or damaged tissues. Previous approaches have typically focused on combining cells and bioactive molecules (e.g., growth factors, cytokines and DNA fragments) with a biomaterial scaffold that functions as a template to control the geometry of the newly formed tissue, while facilitating the attachment, proliferation, and differentiation of embedded cells. Biomaterial scaffolds also play a crucial role in determining the functional properties of engineered tissues, including biomechanical characteristics such as inhomogeneity, anisotropy, nonlinearity or viscoelasticity. While single-phase, homogeneous materials have been used extensively to create numerous types of tissue constructs, there continue to be significant challenges in the development of scaffolds that can provide the functional properties of load-bearing tissues such as articular cartilage. In an attempt to create more complex scaffolds that promote the regeneration of functional engineered tissues, composite scaffolds comprising two or more distinct materials have been developed. This paper reviews various studies on the development and testing of composite scaffolds for the tissue engineering of articular cartilage, using techniques such as embedded fibers and textiles for reinforcement, embedded solid structures, multi-layered designs, or three-dimensionally woven composite materials. In many cases, the use of composite scaffolds can provide unique biomechanical and biological properties for the development of functional tissue engineering scaffolds.     

Electrospinning of highly porous scaffolds for cartilage regeneration

This study presents a new innovative method where electrospinning is used to coat single microfibers with nanofibers. The nanofiber-coated microfibers can be formed into scaffolds with the combined benefits of tailored porosity for cellular infiltration and nanostructured surface morphology for cell growth. The nanofiber coating is obtained by using a grounded collector rotating around the microfiber, to establish an electrical field yet allow collection of nanofibers on the microfiber. A Teflon tube surrounding the fibers and collector is used to force the nanofibers to the microfiber. Polycaprolactone nanofibers were electrospun onto polylactic acid microfibers and scaffolds of 95 and 97% porosities were made. Human chondrocytes were seeded on these scaffolds and on reference scaffolds of purely nanofibers and microfibers. Thereafter, cellular infiltration was investigated. The results indicated that scaffold porosity had great effects on cellular infiltration, with higher porosity resulting in increased infiltration, thereby confirming the advantage of the presented method.     

Hydrophilic gelatin and hyaluronic acid-treated PLGA scaffolds for cartilage tissue engineering

Tissue engineering provides a new strategy for repairing damaged cartilage. Surface and mechanical properties of scaffolds play important roles in inducing cell growth.?Aim: The aim of this study was to fabricate and characterize PLGA and gelatin/hyaluronic acid-treated PLGA (PLGA-GH) sponge scaffolds for articular cartilage tissue engineering. Methods: The PLGA-GH scaffolds were cross-linked with gelatin and hyaluronic acid. Primary chondrocytes isolated from porcine articular cartilages were used to assess cell compatibility. The characteristic PLGA-GH scaffold was higher in water uptake ratio and degradation rate within 42 days than the PLGA scaffold. Results: The mean compressive moduli of PLGA and PLGA-GH scaffolds were 1.72±0.50 MPa and 1.86±0.90 MPa, respectively. The cell attachment ratio, proliferation, and extracellular matrix secretion on PLGA-GH scaffolds are superior to those of PLGA scaffolds. Conclusions: In our study, PLGA-GH scaffolds exhibited improvements in cell biocompatibility, cell proliferation, extracellular matrix synthesis, and appropriate mechanical and structural properties for potential engineering cartilage applications.     

生物可降解材料构建组织工程软骨的研究进展

关节软骨修复困难,目前临床上治疗关节软骨损伤难以达到满意的效果。组织工程学的兴起为其提供了新的选择。本文介绍了组织工程软骨的发展历史,重点叙述了各种天然支架材料、人工合成材料、复合材料及纳米材料在软骨组织工程中的应用及其优势。目前应用的天然材料存在力学强度差及免疫源性的不足;人工合成材料降解速率快,降解产物具有细胞毒性,有待进一步完善。表面修饰等技术的应用在一定程度上克服了某些材料的不足;复合材料综合了数种材料的优点,是今后材料技术发展的方向;纳米技术的出现使新合成的材料成为纳米量级,具有了普通材料无可比拟的优势,这为组织工程材料的发展提供了新的思路。本文还对组织工程支架材料存在的问题、下一步的发展方向和前瞻性研究做了介绍。     

Processing and characterization of porous structures from chitosan and starch for tissue engineering scaffolds

Natural biodegradable polymers were processed by different techniques for the production of porous structures for tissue engineering scaffolds. Potato, corn, and sweet potato starches and chitosan, as well as blends of these, were characterized and used in the experiments. The techniques used to produce the porous structures included a novel solvent-exchange phase separation technique and the well-established thermally induced phase separation method. Characterization of the open pore structures was performed by measuring pore size distribution, density, and porosity of the samples. A wide range of pore structures ranging from 1 to 400 microm were obtained. The mechanisms of pore formation are discussed for starch and chitosan scaffolds. Pore morphology in starch scaffolds seemed to be determined by the initial freezing temperature/freezing rate, whereas in chitosan scaffolds the shape and size of pores may have been determined by the processing route used. The mechanical properties of the scaffolds were assessed by indentation tests, showing that the indentation collapse strength depends on the pore geometry and the material type. Bioactivity and degradation of the potential scaffolds were assessed by immersion in simulated body fluid.     

Fabrication of gelatin-hyaluronic acid hybrid scaffolds with tunable porous structures for soft tissue engineering

The development of three-dimensional (3-D) scaffolds with highly open porous structure is one of the most important issues in tissue engineering. In this study, 3-D macroporous gelatin/hyaluronic acid (GE/HA) hybrid scaffolds with varying porous morphology were prepared by freeze-drying their blending solutions and subsequent chemical crosslinking by using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC). The resulting scaffolds were characterized by scanning electron microscopy (SEM) and Fourier transform infrared spectroscopy (FTIR). Their swelling, in vitro degradation properties and compressive strength were also investigated. To evaluate in vitro cytocompatibility of scaffolds, mouse L929 fibroblasts were seeded onto the scaffolds for cell morphology and cell viability studies. It was found that the porous structure of scaffolds can be tailored by varying the ratios of gelatin to HA, both the swelling ratios and degradation rate increased with the increase of HA content in hybrid scaffolds, and crosslinking the scaffolds with EDC improved the degradation resistance of the scaffold in culture media and increased the mechanical strength of scaffolds. The in vitro results revealed that the prepared scaffolds do not induce cytotoxic effects and suitable for cell growth, especially in the case of scaffolds with higher gelatin content. The combined results of the physicochemical and biological studies suggested that the developed GE/HA hybrid scaffolds exhibit good potential and biocompatibility for soft tissue engineering applications.     

Tissue engineering of human cartilage in bioreactors using single and composite cell-seeded scaffolds

Chondrocytes isolated from human fetal epiphyseal cartilage were seeded under mixed conditions into 15-mm-diameter polyglycolic acid (PGA) scaffolds and cultured in recirculation column bioreactors to generate cartilage constructs. After seeding, the cell distributions in thick (4.75 mm) and thin (2.15 mm) PGA disks were nonuniform, with higher cell densities accumulating near the top surfaces. Composite scaffolds were developed by suturing together two thin PGA disks after seeding to manipulate the initial cell distribution before bioreactor culture. The effect of medium flow direction in the bioreactors, including periodic reversal of medium flow, was also investigated. The quality of the tissue-engineered cartilage was assessed after 5 weeks of culture in terms of the tissue wet weight, glycosaminoglycan (GAG), total collagen and collagen type II contents, histological analysis of cell, GAG and collagen distributions, and immunohistochemical analysis of collagen types I and II. Significant enhancement in construct quality was achieved using composite scaffolds compared with single PGA disks. Operation of the bioreactors with periodic medium flow reversal instead of unidirectional flow yielded further improvements in tissue weight and GAG and collagen contents with the composite scaffolds. At harvest, the constructs contained GAG concentrations similar to those measured in ex vivo human adult articular cartilage; however, total collagen and collagen type II levels were substantially lower than those in adult tissue. This study demonstrates that the location of regions of high cell density in the scaffold coupled with application of dynamic bioreactor operating conditions has a significant influence on the quality of tissue-engineered cartilage.     

Supercritical fluid-assisted controllable fabrication of open and highly interconnected porous scaffolds for bone tissue engineering

Recently tremendous progress has been evidenced by the advancements in developing innovative three-dimensional(3 D)scaffolds using various techniques for addressing the autogenous grafting of bone. In this work, we demonstrated the fabrication of porous polycaprolactone(PCL) scaffolds for osteogenic differentiation based on supercritical fluid-assisted hybrid processes of phase inversion and foaming. This eco-friendly process resulted in the highly porous biomimetic scaffolds with open and interconnected architectures. Initially, a 2~3 factorial experiment was designed for investigating the relative significance of various processing parameters and achieving better control over the porosity as well as the compressive mechanical properties of the scaffold. Then, single factor experiment was carried out to understand the effects of various processing parameters on the morphology of scaffolds. On the other hand, we encapsulated a growth factor, i.e., bone morphogenic protein-2(BMP-2), as a model protein in these porous scaffolds for evaluating their osteogenic differentiation. In vitro investigations of growth factor loaded PCL scaffolds using bone marrow stromal cells(BMSCs) have shown that these growth factor-encumbered scaffolds were capable of differentiating the cells over the control experiments. Furthermore, the osteogenic differentiation was confirmed by measuring the cell proliferation, and alkaline phosphatase(ALP) activity, which were significantly higher demonstrating the active bone growth. Together, these results have suggested that the fabrication of growth factor-loaded porous scaffolds prepared by the eco-friendly hybrid processing efficiently promoted the osteogenic differentiation and may have a significant potential in bone tissue engineering.     

几丁聚糖在组织工程中的应用

支架材料作为组织工程的生物学植入替代物,对细胞移植与引导新组织生长有重要的作用。几丁聚糖可制成无毒性,无刺激性,生物相容性和生物可降解性良好的生物医用材料,在人工皮肤,骨修复材料,手术缝线等方面已广泛应用。本文分析了纯几丁聚糖支架结构和它与其他天然或合成材料复合的支架结构的物理、化学性质及其独特的生物学功能,同时还进一步介绍了其应用的范例并探讨了发展前景。     

Collagen scaffolds for tissue engineering

There are two major approaches to tissue engineering for regeneration of tissues and organs. One involves cell-free materials and/or factors and one involves delivering cells to contribute to the regeneraion process. Of the many scaffold materials being investigated, collagen type I, with selective removal of its telopeptides, has been shown to have many advantageous features for both of these approaches. Highly porous collagen lattice sponges have been used to support in vitro growth of many types of tissues. Use of bioreactors to control in vitro perfusion of medium and to apply hydrostatic fluid pressure has been shown to enhance histogenesis in collagen scaffolds. Collagen sponges have also been developed to contain differentiating-inducing materials like demineralized bone to stimulate differentiation of cartilage tissue both in vitro and in vivo.     

Biodendrimer-based hydrogel scaffolds for cartilage tissue repair

Photo-crosslinkable dendritic macromolecules are attractive materials for the preparation of cartilage tissue engineering scaffolds that may be optimized for in situ formation of hydrated, mechanically stable, and well-integrated hydrogel scaffolds supporting chondrocytes and chondrogenesis. We designed and synthesized a novel hydrogel scaffold for cartilage repair, based on a multivalent and water-soluble tri-block copolymer consisting of a poly(ethylene glycol) core and methacrylated poly(glycerol succinic acid) dendrimer terminal blocks. The terminal methacrylates allow mild and biocompatible photo-crosslinking with a visible light, facilitating in vivo filling of irregularly shaped defects with the dendrimer-based scaffold. The multivalent dendrimer constituents allow high crosslink densities that inhibit swelling after crosslinking while simultaneously introducing biodegradation sites. The mechanical properties and water content of the hydrogel can easily be tuned by changing the biodendrimer concentration. In vitro chondrocyte encapsulation studies demonstrate significant synthesis of neocartilaginous material, containing proteoglycans and type II collagen.     

Smart materials as scaffolds for tissue engineering

In this review, we focused our attention on the more important natural extracellular matrix (ECM) molecules (collagen and fibrin), employed as cellular scaffolds for tissue engineering and on a class of semi-synthetic materials made from the fusion of specific oligopeptide sequences, showing biological activities, with synthetic materials. In particular, these new "intelligent" scaffolds may contain oligopeptide cleaving sequences specific for matrix metalloproteinases (MMPs), integrin binding domains, growth factors, anti-thrombin sequences, plasmin degradation sites, and morphogenetic proteins. The aim was to confer to these new "intelligent" semi-synthetic biomaterials, the advantages offered by both the synthetic materials (processability, mechanical strength) and by the natural materials (specific cell recognition, cellular invasion, and the ability to supply differentiation/proliferation signals). Due to their characteristics, these semi-synthetic biomaterials represent a new and versatile class of biomimetic hybrid materials that hold clinical promise in serving as implants to promote wound healing and tissue regeneration.     

Heparin-functionalized chitosan scaffolds for bone tissue engineering

The aim of this study is to investigate the effects of heparin-functionalized chitosan scaffolds on the activity of preosteoblasts. The chitosan scaffolds having the pore size of ∼100 μm were prepared by a freeze-drying method. Two different methods for immobilization of heparin to chitosan scaffolds were successfully performed. In the first method, functionalization of the scaffolds was achieved by means of electrostatic interactions between negatively charged heparin and positively charged chitosan. The covalent immobilization of heparin to chitosan scaffolds by 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDAC) and N-hydroxysuccinimide (NHS) was used as a second immobilization method. Morphology, proliferation, and differentiation of MC3T3-E1 preosteoblasts on heparin-functionalized chitosan scaffolds were investigated in vitro. The results indicate that covalently bound heparin containing chitosan scaffolds (CHC) stimulate osteoblast proliferation compared to other scaffolds, that is, unmodified chitosan scaffolds (CH), electrostatically bound heparin containing chitosan scaffolds (EHC), and CH+free heparin (CHF). SEM images also proved the stimulative effect of covalently bound heparin on the proliferation of preosteoblasts. Alkaline phosphatase (ALP) and osteocalcin (OCN) levels of cells proliferated on CHC and EHC were also higher than those for CH and CHF. In vitro studies have demonstrated that chitosan scaffolds increase viability and differentiation of MC3T3-E1 cells especially in the presence of immobilized heparin.     

Permeability analysis of scaffolds for bone tissue engineering

Porous artificial bone substitutes, especially bone scaffolds coupled with osteobiologics, have been developed as an alternative to the traditional bone grafts. The bone scaffold should have a set of properties to provide mechanical support and simultaneously promote tissue regeneration. Among these properties, scaffold permeability is a determinant factor as it plays a major role in the ability for cells to penetrate the porous media and for nutrients to diffuse. Thus, the aim of this work is to characterize the permeability of the scaffold microstructure, using both computational and experimental methods. Computationally, permeability was estimated by homogenization methods applied to the problem of a fluid flow through a porous media. These homogenized permeability properties are compared with those obtained experimentally. For this purpose a simple experimental setup was used to test scaffolds built using Solid Free Form techniques. The obtained results show a linear correlation between the computational and the experimental permeability. Also, this study showed that permeability encompasses the influence of both porosity and pore size on mass transport, thus indicating its importance as a design parameter. This work indicates that the mathematical approach used to determine permeability may be useful as a scaffold design tool.     

Biocompatibility of hydrogel-based scaffolds for tissue engineering applications

Recently, understanding of the extracellular matrix (ECM) has expanded rapidly due to the accessibility of cellular and molecular techniques and the growing potential and value for hydrogels in tissue engineering. The fabrication of hydrogel-based cellular scaffolds for the generation of bioengineered tissues has been based on knowledge of the composition and structure of ECM. Attempts at recreating ECM have used either naturally-derived ECM components or synthetic polymers with structural integrity derived from hydrogels. Due to their increasing use, their biocompatibility has been questioned since the use of these biomaterials needs to be effective and safe. It is not surprising then that the evaluation of biocompatibility of these types of biomaterials for regenerative and tissue engineering applications has been expanded from being primarily investigated in a laboratory setting to being applied in the multi-billion dollar medicinal industry. This review will aid in the improvement of design of non-invasive, smart hydrogels that can be utilized for tissue engineering and other biomedical applications. In this review, the biocompatibility of hydrogels and design criteria for fabricating effective scaffolds are examined. Examples of natural and synthetic hydrogels, their biocompatibility and use in tissue engineering are discussed. The merits and clinical complications of hydrogel scaffold use are also reviewed. The article concludes with a future outlook of the field of biocompatibility within the context of hydrogel-based scaffolds.     

Mechanical and biological performances of new scaffolds made of collagen hydrogels and fibroin microfibers for vascular tissue engineering

A microstructured composite material made of collagen hydrogel (matrix) and silk fibroin microfibers (randomly oriented reinforcing fibers) is investigated in order to conjugate the mechanical resistance of fibroin with the suitable biological performance of collagen to design new scaffolds for vascular tissue engineering. Results show that fibroin microfibers and collagen fibrils have suitable interfacial adhesion, and the scaffold exhibits improved mechanical properties if compared with a pure collagen hydrogel. Furthermore, the overall biological performance is improved.     

Competent processing techniques for scaffolds in tissue engineering

Engineering a functional tissue ex vivo requires a synchronized effort towards developing technologies for ECM mimicking scaffold and cultivating tissue-specific cells in an integrated and controlled manner. Cell-interactive scaffolds in three dimensions (3D), designed and processed appropriately with an apt biomaterial to yield optimal porosity and mechanical strength is the key in tissue engineering (TE). In order to accomplish these facets in a 3D scaffold, multiple techniques and processes have been explored by researchers all over the world. New techniques offering reasonable flexibility to use blends of different materials for integrated tissue-specific mechanical strength and biocompatibility have an edge over conventional methods. They may allow a combinatorial approach with a mix of materials while incorporating multiple processing techniques for successful creation of tissue-specific ECM mimics. In this review, we analyze the material requirement from different TE perspectives, while discussing pros and cons of advanced fabrication techniques for scale-up manufacturing.