Isotropic and Anisotropic Scaffolds for Tissue Engineering: Collagen, Conventional, and Textile Fabrication Technologies and Properties.

In this review article, tissue engineering and regenerative medicine are briefly explained and the importance of scaffolds is highlighted. Furthermore, the requirements of scaffolds and how they can be fulfilled by using specific biomaterials and fabrication methods are presented. Detailed insight is given into the two biopolymers chitosan and collagen. The fabrication methods are divided into two categories: isotropic and anisotropic scaffold fabrication methods. Processable biomaterials and achievable pore sizes are assigned to each method. In addition, fiber spinning methods and textile fabrication methods used to produce anisotropic scaffolds are described in detail and the advantages of anisotropic scaffolds for tissue engineering and regenerative medicine are highlighted.

Anisotropic Chitosan Scaffolds Generated by Electrostatic Flocking Combined with Alginate Hydrogel Support Chondrogenic Differentiation.

The replacement of damaged or degenerated articular cartilage tissue remains a challenge, as this non-vascularized tissue has a very limited self-healing capacity. Therefore, tissue engineering (TE) of cartilage is a promising treatment option. Although significant progress has been made in recent years, there is still a lack of scaffolds that ensure the formation of functional cartilage tissue while meeting the mechanical requirements for chondrogenic TE. In this article, we report the application of flock technology, a common process in the modern textile industry, to produce flock scaffolds made of chitosan (a biodegradable and biocompatible biopolymer) for chondrogenic TE. By combining an alginate hydrogel with a chitosan flock scaffold (CFS+ALG), a fiber-reinforced hydrogel with anisotropic properties was developed to support chondrogenic differentiation of embedded human chondrocytes. Pure alginate hydrogels (ALG) and pure chitosan flock scaffolds (CFS) were studied as controls. Morphology of primary human chondrocytes analyzed by cLSM and SEM showed a round, chondrogenic phenotype in CFS+ALG and ALG after 21 days of differentiation, whereas chondrocytes on CFS formed spheroids. The compressive strength of CFS+ALG was higher than the compressive strength of ALG and CFS alone. Chondrocytes embedded in CFS+ALG showed gene expression of chondrogenic markers (COL II, COMP, ACAN), the highest collagen II/I ratio, and production of the typical extracellular matrix such as sGAG and collagen II. The combination of alginate hydrogel with chitosan flock scaffolds resulted in a scaffold with anisotropic structure, good mechanical properties, elasticity, and porosity that supported chondrogenic differentiation of inserted human chondrocytes and expression of chondrogenic markers and typical extracellular matrix.

Silk Fiber-Reinforced Hyaluronic Acid-Based Hydrogel for Cartilage Tissue Engineering.

A continuing challenge in cartilage tissue engineering for cartilage regeneration is the creation of a suitable synthetic microenvironment for chondrocytes and tissue regeneration. The aim of this study was to develop a highly tunable hybrid scaffold based on a silk fibroin matrix (SM) and a hyaluronic acid (HA) hydrogel. Human articular chondrocytes were embedded in a porous 3-dimensional SM, before infiltration with tyramine modified HA hydrogel. Scaffolds were cultured in chondropermissive medium with and without TGF-ß1. Cell viability and cell distribution were assessed using CellTiter-Blue assay and Live/Dead staining. Chondrogenic marker expression was detected using qPCR. Biosynthesis of matrix compounds was analyzed by dimethylmethylene blue assay and immuno-histology. Differences in biomaterial stiffness and stress relaxation were characterized using a one-step unconfined compression test. Cell morphology was investigated by scanning electron microscopy. Hybrid scaffold revealed superior chondro-inductive and biomechanical properties compared to sole SM. The presence of HA and TGF-ß1 increased chondrogenic marker gene expression and matrix deposition. Hybrid scaffolds offer cytocompatible and highly tunable properties as cell-carrier systems, as well as favorable biomechanical properties.

Effects of (Oxy-)Fluorination on Various High-Performance Yarns.

In this work, typical high-performance yarns are oxy-fluorinated, such as carbon fibers, ultra-high-molecular-weight polyethylene, poly(p-phenylene sulfide) and poly(p-phenylene terephthalamide). The focus is on the property changes of the fiber surface, especially the wetting behavior, structure and chemical composition. Therefore, contact angle, XPS and tensile strength measurements are performed on treated and untreated fibers, while SEM is utilized to evaluate the surface structure. Different results for the fiber materials are observed. While polyethylene exhibits a relevant impact on both surface and bulk properties, polyphenylene terephthalamide and polyphenylene sulfide are only affected slightly by (oxy-)fluorination. The wetting of carbon fiber needs higher treatment intensities, but in contrast to the organic fibers, even its textile-physical properties are enhanced by the treatment. Based on these findings, the capability of (oxy-)fluorination to improve the adhesion of textiles in fiber-reinforced composite materials can be derived.

Collagen multifilament spinning.

The benefits of fiber based implants and scaffolds for tissue engineering applications are their anisotropic, highly porous, and controllable macro-, micro-, and nanostructure. Collagen is one of the most commonly used material for the fabrication of scaffolds, as this biopolymer is present in the natural extracellular matrix. For textile processing and textile scaffold fabrication methods, multifilament yarns are required, however, only monofilaments can be generated by state-of-the-art collagen spinning. Hence, the research presented in here aimed at the development of a collagen multifilament wet-spinning process in reproducible quality as well as the characterization of non-crosslinked and crosslinked wet-spun multifilament yarns. Wet spun collagen yarns were comprised of 6 single filaments each having a fineness of 5 tex and a diameter of 80 µm. The tensile strength of the glutaraldehyde crosslinked yarns was 169 MPa (Young's modulus 3534 MPa) in the dry state and 40 MPa (Young's modulus 281 MPa) in the wet state. Furthermore, wet spun collagen filaments showed a characteristic fibrillar structure, which was similar the morphological structure of natural collagen fibers. The textile processing of collagen multifilament yarn was demonstrated by means of knitting technology.

Functionalized silk fibers from transgenic silkworms for wound healing applications: Surface presentation of bioactive epidermal growth factor.

Growth factors play a crucial role in wound healing in general and are promising tools for the treatment of chronic wounds as they can restore the physiological wound healing process. In growth factor-loaded wound dressings, human epidermal growth factor (EGF) is released in a burst and washed out quickly. The developed matrix consists of recombinant EGF produced in transgenic silkworms as a fusion protein with the fibroin light chain. The covalent linkage prevents EGF from draining into the surrounding tissue while presenting the growth factor on the surface. EGF-functionalized silk membranes and nonwovens lead to a 2.5-fold increase in the cell number of fibroblasts, while retaining full bioactivity even after e-beam sterilization. EGF is long-term presented without burst release and significantly reduces the wound area by 15% in an in vitro wound model. Hence, the cost-effective production of a biomaterial using transgenic silkworm larvae in combination with a growth factor paves the way for a promising new multifactorial wound cover for chronic wound healing. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 2643-2652, 2018.

Wet spinning and riboflavin crosslinking of collagen type I/III filaments.

Reconstituted fibrillary collagen is one of the most advantageous biomaterials for biomedical applications. The objective of the research project described in this paper was to evaluate whether riboflavin-induced photo-crosslinking could be used as a non-toxic alternative to glutaraldehyde (GA)-crosslinking for the preparation of wet spun collagen filaments. Collagen filaments were produced on a laboratory wet spinning line and crosslinked with GA or riboflavin with and without UV exposure. Based on mechanical and thermal analyses, it was concluded that the combination of riboflavin and UV light leads to crosslinked collagen filaments having improved mechanical and thermal properties. Furthermore, riboflavin-crosslinked filaments exhibited a higher cytocompatibility for human mesenchymal stem cells compared to GA-crosslinked filaments.

In silico modeling of structural and porosity properties of additive manufactured implants for regenerative medicine.

Additive manufacturing technologies are a promising technology towards patient-specific implants for applications in regenerative medicine. The Net-Shape-Nonwoven technology is used to manufacture structures from short fibers with interconnected pores and large functional surfaces that are predestined for cell adhesion and growth. The present study reports on a modeling approach with a particular focus on the specific structural properties. The overall porosities and mean pore-sizes of the digital models are simulated according to liquid-displacement porosity in a tool implemented in the modeling software. This allows adjusting the process parameters fiber length and fiber diameter to generate biomimetic structures with pore-sizes adapted to the requirements of the tissue that is to be replaced. Modeling the structural and porosity properties of scaffolds and implants leads to an efficient use of the processed biomaterials as the trial-and-error method is avoided.

Electrostatic flocking of chitosan fibres leads to highly porous, elastic and fully biodegradable anisotropic scaffolds.

UNLABELLED: Electrostatic flocking - a common textile technology which has been applied in industry for decades - is based on the deposition of short polymer fibres in a parallel aligned fashion on flat or curved substrates, covered with a layer of a suitable adhesive. Due to their highly anisotropic properties the resulting velvet-like structures can be utilised as scaffolds for tissue engineering applications in which the space between the fibres can be defined as pores. In the present study we have developed a fully resorbable compression elastic flock scaffold from a single material system based on chitosan. The fibres and the resulting scaffolds were analysed concerning their structural and mechanical properties and the biocompatibility was tested in vitro. The tensile strength and Young's modulus of the chitosan fibres were analysed as a function of the applied sterilisation technique (ethanol, supercritical carbon dioxide, γ-irradiation and autoclaving). All sterilisation methods decreased the Young's modulus (from 14GPa to 6-12GPa). The tensile strength was decreased after all treatments - except after the autoclaving of chitosan fibres submerged in water. Compressive strength of the highly porous flock scaffolds was 18±6kPa with a elastic modulus in the range of 50-100kPa. The flocked scaffolds did not show any cytotoxic effect during indirect or direct culture of human mesenchymal stem cells or the sarcoma osteogenic cell line Saos-2. Furthermore cell adhesion and proliferation of both cell types could be observed. This is the first demonstration of a fully biodegradable scaffold manufactured by electrostatic flocking. STATEMENT OF SIGNIFICANCE: Most tissues possess anisotropic fibrous structures. In contrast, most of the commonly used scaffolds have an isotropic morphology. By utilising the textile technology of electrostatic flocking, highly porous and clearly anisotropic scaffolds can be manufactured. Flocking leads to parallel aligned short fibres, glued on the surface of a substrate. Such structures are characterised by a high and adjustable porosity, accompanied by distinct stiffness in fibre direction. The present article describes for the first time a fully biodegradable flock scaffold, solely made of chitosan. Utilisation of only one material for manufacturing of flock substrate, adhesive and fibres allow a uniform degradation of the whole construct. Such a new type of scaffold can be of great interest for a variety of biomedical applications.

Chitosan(PEO)/silica hybrid nanofibers as a potential biomaterial for bone regeneration.

New hybrid nanofibers prepared with chitosan (CTS), containing a total amount of polyethylene oxide (PEO) down to 3.6wt.%, and silica precursors were produced by electrospinning. The solution of modified sol-gel particles contained tetraethoxysilane (TEOS) and the organosilane 3-glycidyloxypropyltriethoxysilane (GPTEOS). This is rending stable solution toward gelation and contributing in covalent bonding with chitosan. The fibers encompass advantages of biocompatible polymer template silicate components to form self-assembled core-shell structure of the polymer CTS/PEO encapsulated by the silica. Potential applicability of this hybrid material to bone tissue engineering was studied examining its cellular compatibility and bioactivity. The nanofiber matrices were proved cytocompatible when seeded with bone-forming 7F2-cells, promoting attachment and proliferation over 7 days. These found to enhance a fast apatite formation by incorporation of Ca(2+) ions and subsequent immersion in modified simulated body fluid (m-SBF). The tunable properties of these hybrid nanofibers can find applications as active biomaterials in bone repair and regeneration.

Ulvan and ulvan/chitosan polyelectrolyte nanofibrous membranes as a potential substrate material for the cultivation of osteoblasts.

A new generation of biomaterials composed of the natural polysaccharides, ulvans extracted from the green seaweed Ulva rigida and chitosan have been investigated. Ulvan, chitosan alone and ulvan/chitosan polyelectrolyte membranes have been synthesised and characterised. The structure of the membranes was altered by the weight ratio of the polyion components. Fibrous and nanofibrous morphology was created, in accordance with a supramolecular self assembly. ATR-FTIR measurements suggested the presence of both polycationic chitosan and polyanionic ulvan in the polyelectrolyte membranes. The cytocompatibility of these new materials was examined by fluorescence microscopy. The results show that ulvan as well as ulvan/chitosan membranes promoted the attachment and proliferation of 7F2 osteoblasts and maintained the cell morphology and viability. Thus, ulvan and chitosan which possess unique properties might have high impact in biomedical applications as potential scaffold materials.

Inorganic/organic (SiO2)/PEO hybrid electrospun nanofibers produced from a modified sol and their surface modification possibilities.

Ceramic silica (SiO(2)) hybrid nanofibers were prepared by electrospinning of solutions containing biocompatible polymer and modified silica precursors. The new hybrid nanofibers are based on polyethylene oxide (PEO) and a new solution of modified sol-gel particles of mixture containing tetraethoxysilane (TEOS) and 3-glycidyloxypropyltriethoxysilane (GPTEOS) in a weight ratio of 3:1. Adding high-molecular-weight PEO into the silica sol is found to enhance the formation of the silica nanofibers and leads to reduce the water-soluble carrying polymer down to 1.2%wt. Transmission electron microscopy (TEM) and attenuated total reflection fourier transformation infrared ATR-FTIR measurements are suggested that PEO is encapsulated by the silica component. This excellent formulation renders electrospinning of SiO(2) a robust process for an easy production of controllable silicate nanofibrous matrices. For instance, nanofibers with average diameter down to 400 nm are accessible by varying the weight ratio between the polymer and the inorganic precursor. These are reduced to 120 nm after the pyrolysis process. Moreover, the surface of the nanofibers could be easily modified, either by Al(3+) leading to aluminium silicate coatings, or by incorporation of Ca(2+) ions and subsequent bioactive hydroxyl carbonate apatite (HAP) formation. These hybrid silica nanofibers are possess a unique collective properties can have a great impact either in high-temperature reinforced materials and filtration or in biomedical applications such as in dentistry and bone tissue engineering.