Scaffolds For Use In Tissue Engineering Research Articles

A novel, microfluidic high-throughput single-cell encapsulation of human bone marrow mesenchymal stromal cells

The efficacy of stem-cell therapy depends on the ability of the transplanted cells to escape early immunological reactions and to be retained at the site of transplantation. The use of tissue engineering scaffolds or injectable biomaterials as carriers has been proposed, but they still present limitations linked to a reliable manufacturing process, surgical practice and clinical outcomes. Alginate microbeads are potential candidates for the encapsulation of mesenchymal stromal cells with the aim of providing a delivery carrier suitable for minimally-invasive and scaffold-free transplantation, tissue-adhesive properties and protection from the immune response. However, the formation of stable microbeads relies on the cross-linking of alginate with divalent calcium ions at concentrations that are toxic for the cells, making control over the beads’ size and a single-cell encapsulation unreliable. The present work demonstrates the efficiency of an innovative, high throughput, and reproducible microfluidic system to produce single-cell, calcium-free alginate coatings of human mesenchymal stromal cells. Among the various conditions tested, visible light and confocal microscopy following staining of the cell nuclei by DAPI showed that the microfluidic system yielded an optimal single-cell encapsulation of 2000 cells/min in 2% w/v alginate microcapsules of reproducible morphology and an average size of 28.2 ± 3.7 µm. The adhesive properties of the alginate microcapsules, the viability of the encapsulated cells and their ability to escape the alginate microcapsule were demonstrated by the relatively rapid adherence of the beads onto tissue culture plastic and the cells’ ability to gradually disrupt the microcapsule shell after 24 h and proliferate. To mimic the early inflammatory response upon transplantation, the encapsulated cells were exposed to proliferating macrophages at different cell seeding densities for up to 2 days and the protection effect of the microcapsule on the cells assessed by time-lapse microscopy showing a shielding effect for up to 48 h. This work underscores the potential of microfluidic systems to precisely encapsulate cells by good manufacturing practice standards while favouring cell retention on substrates, viability and proliferation upon transplantation.Graphical

Applications of Polymers for Tissue Regeneration in Medical Treatment

Multiple types of damage to human organs can result in dysfunction and incapacity. The importance of tissue repair and regeneration in the medical industry results in high demand for tissue regeneration materials. Since the scaffolds considerably speed up the healing process, the use of tissue engineering scaffolds in the restoration of injured tissues has shown amazing potential. Due to their superior processability and biocompatibility, polymers are frequently used to build three-dimensional scaffolds. However, the main issues that restrict their widespread clinical application are their inadequate mechanical strength and unsuitable degradation rate in the bone regeneration process. Because of their superior mechanical strength, synthetic biodegradable polymer composites are used. In this research, the most frequently used polymers including polycaprolactone (PCL) and gelatine methacrylate (GelMA) were employed to create polymer composite scaffolds. And this research categorized polymer materials for tissue regeneration based on their uses in different human organs and described their physical or chemical properties. The reader is given evaluations of each material as well as prospects.

Preparation and characterization of photocured poly (ε-caprolactone) diacrylate/poly (ethylene glycol) diacrylate/chitosan for photopolymerization-type 3D printing tissue engineering scaffold application

Because of its biocompatible, biodegradable and antimicrobial properties, chitosan is an attractive biomaterial for use in tissue engineering scaffolds. This work builds on previous research by incorporating 95% DD chitosan into a visible-light curable resin which is compatible with a digital light processing (DLP™) projection additive manufacturing (3D printing) system. Different concentrations of chitosan were added to a poly (ε-caprolactone)-diacrylate/poly (ethylene glycol)-diacrylate baseline resin and the samples were extensively characterized. Thermal and mechanical analysis conformed to established scaffold requirements. L929 cells were cultured on the photo-crosslinked films and MTT assays were performed at 1, 3, and 5days to assess cytocompatibility of the resins. Data and SEM images verified a correlation between the concentration of chitosan in the photocurable resin and the adhesion, proliferation, and viability of cell cultures. Finally, the processability of the resins with the dynamic masking DLP system was demonstrated by constructing multi-layer scaffolds with actual measurements that were consistent with the CAD models. These findings encourage the use of chitosan as an additive in visible-light curable resins to improve desired properties in tissue engineering scaffolds.

Magnetic nanocomposites for tissue regeneration therapies

Event Abstract Back to Event Magnetic nanocomposites for tissue regeneration therapies Rebecca Zhiyu Yuan1, Kaveh Memarzadeh1, Robert A. Brown2 and Jie Huang1 1 University College London, Mechanical Engineering, United Kingdom 2 University College London, Faculty of Medical Sciences, United Kingdom Introduction: The use of tissue engineering scaffolds to repair and promote bone formation has received much clinical attention over the past decades. However, to guide and stimulate cells towards the target site remains challenging. There are increasing efforts to promote current cell therapies by developing new methodologies to target, monitor, and control cells. Recently, magnetic nanoparticles (MNPs) have been identified as being capable of promoting the proliferation and differentiation of osteoblast under static magnetic field (SMF)[1]. Iron oxide nanoparticles (IONPs) are clinically approved and can be injected intravenously, however, the majority of the IONPs are rapidly cleared from the bloodstream by the reticuloendothelial system. IONPs have been encapsulated in a gel matrix; there are concerns of toxic side-effects of organic solvents. Encapsulated electrospun nanofibre web has offered an attractive alternative solution[2]. In this study, IONPs were synthesised and their nanofiber mesh were produced by eletrospinning (ES). A device to support cell culture under SMF was prepared by 3D printing, and used for in vitro study. Materials and Methods: IONPs were prepared by precipitation of iron (III) chloride hexahydrate and Iron (II) chloride tetrahydrate with an addition of ammonia[3]. The microstructure of IONPs was examined by Transmission Electron Microscopy (TEM) and phase purity was determined by X-ray diffraction (XRD). The biocompatibility of IONPs was assessed by in vitro culture using a human osteoblast MG63 cell model. IONPs / polyvinylpyrrolidone (PVP) / ethanol / water suspensions were formulated for ES, and various parameters were employed to control the size of fibres. The microstructure of the nanofibre mesh was examined by Scanning Electron Microscopy (SEM) and Scanning Probe Microscopy (SPM). Results and Discussion: By using TEM examination, the image analysis showed that the average diameter of the IONPs was 30∓ 20 nm. Phase purity was further confirmed by XRD; IONP nanofibre meshes were produced with size of 100 nm to 2 μm. An uniform distribution of IONPs in the nanocomposites was revealed form SPM (Fig. 1). Figure 1. (A) TEM micrograph of IONPs (B) SEM and (C) SPM micrographs of IONPs nanofibre mesh. Alamar Blue assay showed that MG63 cells were able to proliferate when in contact with IONPs coated substrates during 7 days culture. The cell activity was increased significantly under SMF (Fig. 2) indicating a stimulating effect. Figure 2. A comparison of the proliferation of MG63 cells under SMF and non-static magnetic field (NSMF) from alamar blue assay (n=3). Conclusion: Biocompatible iron oxide nanoparticles were successfully prepared and tested. The size and morphology of electrospun IONP nanofibres can be controlled by varying solution properties and processing parameters. With the aid of external static magnetic field, the three-dimensional magnetic nanocomposites scaffolds can offer great potential in magnetic cell guiding applications.

A multi-scale controlled tissue engineering scaffold prepared by 3D printing and NFES technology

The current focus in the field of life science is the use of tissue engineering scaffolds to repair human organs, which has shown great potential in clinical applications. Extracellular matrix morphology and the performance and internal structure of natural organs are required to meet certain requirements. Therefore, integrating multiple processes can effectively overcome the limitations of the individual processes and can take into account the needs of scaffolds for the material, structure, mechanical properties and many other aspects. This study combined the biological 3D printing technology and the near-field electro-spinning (NFES) process to prepare a multi-scale controlled tissue engineering scaffold. While using 3D printing technology to directly prepare the macro-scaffold, the compositing NFES process to build tissue micro-morphology ultimately formed a tissue engineering scaffold which has the specific extracellular matrix structure. This scaffold not only takes into account the material, structure, performance and many other requirements, but also focuses on resolving the controllability problems in macro- and micro-forming which further aim to induce cell directed differentiation, reproduction and, ultimately, the formation of target tissue organs. It has in-depth immeasurable significance to build ideal scaffolds and further promote the application of tissue engineering.

Special feature on stem cells: current research and future prospects

Stem cell-based therapies are viewed as among the most promising new strategies against developmental, traumatic, oncogenic and degenerative diseases. Indeed, it was the ability to identify, isolate, culture, interrogate, expand and transplant stem cells and/or their derivatives that gave rise to the new field of ‘regenerative medicine'. While the hope for stem cell-based regenerative medicine grows rapidly, it is becoming clear from extant clinical trials that we still need a greater understanding of the fundamentals of stem cell biology as well as of the disease process in order to make an impact that is efficient, efficacious, safe, reliable, scalable, cost-effective, practical, accessible and affordable. During the past 20 years, the breadth of what is regarded as ‘stem cell research' has expanded immensely, now embracing, for example, tissue-specific, embryonic and induced/reprogrammed stem cells as well as cancer stem cells. Moreover, in the past 3–5 years, clinical trials using or relying on exogenous or endogenous stem cells (including cancer stem cells) have been launched with many more on the drawing board. With such rapid advances in the field, we felt it necessary to concisely take an overview of current thinking in stem cell biology, with a particular eye to potential applications. In this special issue of Experimental & Molecular Medicine, we offer concise coverage of the state-of-the art in a series of representative areas in stem cell research provided by thought leaders in each topic. First, the developmental changes in the characteristics of hematopoietic stem cells (HSCs), the one cobble stone model of tissue-specific stem cells, is discussed by Dr C Eaves. HSCs undergo extensive changes during development in their proliferation, self-renewal and growth factor responsiveness as well as their regenerative capacity. While the mechanisms underlying the shift in HSC properties remain largely unclear, this paper provides a recent view on the miRNA-mediated regulatory mechanisms that can define the developmental changes in HSCs. Another type of stem cell, one that is sometimes shrouded in controversy, ‘very small embryonic-like stem cells (VSELs)', is discussed by Dr M Ratajczak. Dr Ratajczak addresses this controversy by describing their isolation, in-vivo identity and evidence for pluripotency based on transcriptomics and multi-lineage differentiation potential. Dr E Snyder's group provides a recent view on the use of human-induced pluripotent stem cells (hiPSCs) for understanding the mechanisms underlying a complex disease (for example, bipolar disorder) based on responsiveness to a known clinically efficacious drug (for example, lithium). Based on a better understanding of lithium's heretofore unknown mechanism-of-action, then the same hiPSCs may be used to help screen and identify drugs that work better and with less toxicity than lithium against these same targets. In other words, in addition to being potentially useful for autologous cell therapy, hiPSCs can also serve as powerful tools for drug discovery based on their ability to be obtained from patients that harbor a disease and yield cells in a particular lineage that presumably also ‘have that disease'. Moving to more applied stem cell biology, Dr A Caplan provides a concise review of mesenchymal stromal cell (MSC)-based therapy, describing basic concept as well as a mode of therapeutic action based on the secretion of factors that facilitate establishment of a regenerative microenvironment. Dr Caplan offers a spectrum of clinical studies ranging from immunological disorders to organ regeneration that have employed MSCs. Dr J Yoo provides insight on the use of tissue-engineering scaffolds to facilitate a regenerative microenvironment in injured organs by stimulating the recruitment and proliferation of endogenous stem cells. In addition, the current edition provides representative original researches covering each area of stem cell biology and therapeutic applications. The feasibility of using stem cell therapy in the treatment of neuronal diseases using neuronal stem cells and embryonic stem cell-derived neuronal progenitor cells as model systems is explored. Strategies for recruiting endogenous stem cells during wound healing and the possibility of salivary gland regeneration was explored. Finally, the clinical efficacy of stem cell therapy for femoral head necrosis was evaluated for 128 patients after 5 years of follow-up is discussed. Collectively, these articles not only provide an overview of the state-of-the-art and future prospects for stem cell biology, but also describe the pipeline required to enable stem cell research to impact regenerative medicine. We are particularly thankful to Dr Dae-Myung Jue, the Editor-in-Chief of Experimental & Molecular Medicine and to the Nature Publishing Group for their support on this special series ‘Special Feature on Stem Cells: Current Research and Future Prospects'.

Strengthening mechanism of PDLLA coated titania foam

This paper addresses the numerical analysis of the mechanical properties of titania foam intended for use in tissue engineering scaffolds. Special focus is given to a PDLLA coating that has been shown to distinctly increase the mechanical strength of the scaffold. Micro-computed tomography (μCT) data of a real scaffold are obtained and converted into numerical calculation models. Finite element simulations are performed alternately with and without the PDLLA coating. In addition, a strut model containing a single micro-crack is analysed. Numerical results indicate that filling the crack with PDLLA significantly increases the strength of the strut by attenuating stress concentrations.

Bio-inspired materials for biosensing and tissue engineering

This review gives an overview of the research presented in the lecture by Professor Molly Stevens given on the occasion of the 2010 Polymer International-IUPAC award for creativity in polymer science. Herein we describe some highlights of our biomaterials-based approaches in the fields of regenerative medicine and biosensing. We have developed a range of polymeric and inorganic materials for use in tissue engineering scaffolds as well as for elucidating the impact of various materials-based cues on cell behaviour and tissue formation. Specific examples in the field of bone repair are outlined. Furthermore, recent achievements in the creation of sensitive, simple and robust assays for enzyme detection are presented. Our approaches in this field are based on the combination of various nanoparticles with designer polypeptides to provide quantitative colorimetric read-outs where a specific active enzyme is present. These enzyme assays are likely to be useful for portable point-of-care diagnoses of a range of diseases with significant global impact. Copyright © 2012 Society of Chemical Industry

Effects of lactic acid and glycolic acid on human osteoblasts: A way to understand PLGA involvement in PLGA/calcium phosphate composite failure

The use of degradable composite materials in orthopedics remains a field of intense research due to their ability to support new bone formation and degrade in a controlled manner, broadening their use for orthopedic applications. Poly (lactide-co-glycolide) acid (PLGA), a degradable biopolymer, is now a popular material for different orthopedic applications and is proposed for use in tissue engineering scaffolds either alone or combined with bioactive ceramics. Interference screws composed of calcium phosphates and PLGA are readily available in the market. However, some reports highlight problems of screw migration or aseptic cyst formation following screw degradation. In order to understand these phenomena and to help to improve implant formulation, we have evaluated the effects of PLGA degradation products: lactic acid and glycolic acid on human osteoblasts in vitro. Cell proliferation, differentiation, and matrix mineralization, important for bone healing were studied. It was found that the toxicity of polymer degradation products under buffering conditions was limited to high concentrations. However, non-toxic concentrations led to a decrease in cell proliferation, rapid cell differentiation, and mineralization failure. Calcium, whilst stimulating cell proliferation was not able to overcome the negative effects of high concentrations of lactic and glycolic acids on osteoblasts. These effects help to explain recently reported clinical failures of calcium phosphate/PLGA composites, but further in vitro analyses are needed to mimic the dynamic situation which occurs in the body by, for example, culture of osteoblasts with materials that have been pre-degraded to different extents and thus be able to relate these findings to the degradation studies that have been performed previously.

Self-Crimping, Biodegradable, Electrospun Polymer Microfibers

Semicrystalline poly(l-lactide-co-ε-caprolactone) (P(LLA-CL)) was used to produce electrospun fibers with diameters on the subcellular scale. P(LLA-CL) was chosen because it is biocompatible and its chemical and physical properties are easily tunable. The use of a rotating wire mandrel as a collection device in the electrospinning process, along with high collection speeds, was used to align electrospun fibers. Upon removal of the fibers from the mandrel, the fibers shrunk in length, producing a crimp pattern characteristic of collagen fibrils in soft connective tissues. The crimping effect was determined to be a result of the residual stresses resident in the fibers due to the fiber alignment process and the difference between the operating temperature (T(op)) and the glass-transition temperature (T(g)) of the polymer. The electrospun fibers could be induced to crimp by adjusting the operating temperature to be greater than that of the polymer glass-transition temperature. Moreover, the crimped fibers exhibited a toe region in their stress-strain profile that is characteristic of collagen present in tendons and ligaments. The crimp pattern was retained during in vitro degradation over 4 weeks. Primary bovine fibroblasts seeded onto these crimped fibers attached, proliferated, and deposited extracellular matrix (ECM) molecules on the surface of the fiber mats. These self-crimping fibers hold great promise for use in tissue engineering scaffolds for connective tissues that require fibers similar in structure to that of crimped collagen fibrils.

Development of collagen condensation method to improve mechanical strength of tissue engineering scaffolds

A major drawback of collagen sponges regarding their use in tissue engineering scaffolds is their weak mechanical properties under wet conditions. To overcome this problem without the use of other skeletal materials, the exhaustive condensation technique of reconstituted collagen fibrils was developed to fabricate high-density collagen sponges using freeze drying. The density linearly increased with an increase in the concentration of collagen fibrils. The compression tests under wet conditions demonstrated that the toughness and stiffness of the collagen sponges increased with an increase in the density. The collagen sponge with a density of 129 mg/cm 3 showed a compressive strength (to a strain of 30%) of 8.88 kPa and a modulus of 332 kPa. These results suggested the possibility that the mechanical properties of collagen sponges can increase significantly while retaining the sponges' inherent bioresorbability.

Preparation and characterization of 2,3-dialdehyde bacterial cellulose for potential biodegradable tissue engineering scaffolds

Bacterial cellulose (BC) is suitable for applications as scaffolds in tissue engineering due to its unique properties. However, BC is not enzymatically degradable in vivo and this has become an essential limiting factor in its potential applications. In this work, BC was modified by periodate oxidation to give rise to a biodegradable 2,3-dialdehyde bacterial cellulose (DABC). After characterization by scanning electron microscopy (SEM), energy dispersive spectroscopy (EDS), attenuated total reflectance–Fourier transform infrared (ATR–FTIR) spectroscopy, thin-film X-ray diffractometry (XRD) and X-ray photoelectron spectroscopy (XPS), we demonstrated that the modified DABC nano-network was able to degrade into porous scaffold with micro-sized pores in water, phosphate buffered saline (PBS) and the simulated body fluid (SBF). The degradation process began from the oxidized amorphous part of the network and concurrently hydroxyapatite formed on the scaffold surface during the process in SBF. Our data also demonstrated that the tensile mechanical properties of the DABC nano-network were suitable for its use in tissue engineering scaffolds.

The design of scaffolds for use in tissue engineering. Part II. Rapid prototyping techniques.

Tissue engineering (TE) is an important emerging area in biomedical engineering for creating biological alternatives for harvested tissues, implants, and prostheses. In TE, a highly porous artificial extracellular matrix or scaffold is required to accommodate mammalian cells and guide their growth and tissue regeneration in three-dimension (3D). However, existing 3D scaffolds for TE proved less than ideal for actual applications because they lack mechanical strength, interconnected channels, and controlled porosity or pores distribution. In this paper, the authors review the application and advancement of rapid prototyping (RP) techniques in the design and creation of synthetic scaffolds for use in TE. We also review the advantages and benefits, and limitations and shortcomings of current RP techniques as well as the future direction of RP development in TE scaffold fabrication.