Biomaterials For Tissue Engineering Scaffolds Research Articles

Quantitative nanomechanical mapping on poly(lactic acid)/poly(ε-caprolactone)/carbon nanotubes bionanocomposites using atomic force microscopy

Abstract Poly(lactic acid)/poly(e-caprolactone) (PLA/PCL) blends are among the most widely investigated biomaterials for tissue engineering scaffolds. Incorporating carboxylic multi-walled carbon nanotubes (MWCNTs) into the blend can further tune the microphase distribution and interface adhesion which conversely dominates the micromechanical properties. In this work, atomic force microscope (AFM) based quantitative nanomechanical measurements were applied on PLA/PCL/MWCNTs bionanocomposites for gaining knowledge at the nanoscale. Results show that MWCNTs are mainly dispersed in the PCL phase which is adjacent to the PLA phase. The microphase structures and the onsite microscopic Young's moduli distributions have significant changes with the introduction of MWCNTs. No obvious intermediate layer in the PLA/MWCNTs and PCL/MWCNTs interfaces is observed in the Young's moduli map, indicating weak interactions in the polymer/MWCNTs interface. Moreover, microscopic structure-property relationships of PLA/PCL/MWCNTs nanocomposites were discussed and correlated to the macroscopic mechanical properties. A better understanding of these properties would be helpful in tailoring the PLA/PCL based bionanomaterials for applications in tissue engineering scaffolds, where the nanomechanical information is critical.

Interpenetrating polymer network hydrogels composed of chitosan and photocrosslinkable gelatin with enhanced mechanical properties for tissue engineering

Gelatin and chitosan (CS) are widely used natural biomaterials for tissue engineering scaffolds, but the poor mechanical properties of pure gelatin or CS hydrogels become a big obstacle that limits their use as scaffolds, especially in load-bearing tissues. This study provided a novel mechanism of forming interpenetrating network (IPN) of gelatin methacryloyl (GelMA) and CS hydrogels by covalent bonds and hydrophobic interactions through photocrosslinking and basification, respectively. By characterization of the compressive and tensile moduli, ultimate tensile stress and strain, it was found that semi-IPN and IPN structure can greatly enhance the mechanical properties of GelMA-CS hydrogels compared to the single network CS or GelMA. Moreover, the increase of either GelMA or CS concentration can strengthen the hydrogel network. Then, the swelling, enzymatic degradation, and morphology of GelMA-CS hydrogels were also systematically investigated. The excellent biocompatibility of GelMA-CS hydrogels was demonstrated by large spreading area of bone mesenchymal stem cells on hydrogel surfaces when CS concentration was <2% (w/v). According to this study, the multiple requirements of properties can be fulfilled by carefully selecting the GelMA and CS compositions for IPN hydrogels.

Evaluation of Extracted β-chitosan From Loligo duvauceli for the Preparation of Tissue Engineering Scaffolds

Over the years, there has been a growing need for producing cost effective, porous scaffold biomaterials for tissue engineering. Therefore, this study focused on fabrication of β-chitosan based scaffold from the waste of the pen of locally available squid variety Loligo duvauceli (Indian Squid). The scaffold was prepared by the solvent casting method which requires less time and minimum energy. To the best of our knowledge,there are no reports on scaffold preparation from the extracted β-chitosan. The biological properties of extracted chitosan scaffolds C(L) were evaluated and compared with those of commercial chitosan scaffold C(S). The prepared scaffolds were characterized by SEM, swelling property, in vitro enzymatic degradation, and hemo, biocompatibility properties. C(L) scaffolds had an average pore size of 60 µm and the swelling ratio was 16.2 ± 2.2% with mass loss percentage of 20.8 ± 4.5% after 4 weeks of degradation. It has exhibited good hemocompatible properties and thermal stability. Mouse fibroblast 3T3 cells were able to adhere and proliferate well in the extracted β-chitosan C(L) scaffold. All these results indicated that extracted β-chitosan C(L) is a suitable alternative for biomaterials used for tissue engineering purposes.

Star-shaped polyhedral oligomeric silsesquioxane-polycaprolactone-polyurethane as biomaterials for tissue engineering application

Polyhedral oligomeric silsesquioxane (POSS) is a unique molecule that is composed of an inorganic silica core with eight organic functional arms, which can be used as a nucleus for covalent bonding to create star-shaped block copolymers with improved mechanical and biological properties. In this work, highly porous star-shaped POSS-polycaprolactone-polyurethane (POSS-PCL-PU) films were synthesized as scaffold biomaterials for tissue engineering. These films have an interesting morphology consisting of rough spherulites with filamentous structures spreading out from their centers; the unique nanotopography was shown to be suitable for cell growth. In vitro degradation was monitored for 52 weeks in terms of the weight loss and morphology changes of the films. The degradation profile exhibited a slow initial weight loss of <1% during the first 24 weeks, followed by a rapid weight loss of ~18% in the following 28 weeks. The films demonstrated excellent biocompatibility and cell-substrate affinity, with a high cell viability of >95% and rapid cell proliferation. The high porosity, unique surface nanotopography and excellent biocompatibility of the star-shaped POSS-PCL-PU film make it a great candidate as a tissue engineering scaffold biomaterial. Researchers in Singapore have prepared highly porous films that are very promising as scaffold biomaterials for tissue engineering. They synthesized the films by using star-shaped polyhedral oligomeric silsesquioxane as the starting inorganic core and incorporating polycaprolactone (PCL) and polyurethane (PU). The researchers, who are from the Institute of Materials Research and Engineering and the National University of Singapore, demonstrate that the inorganic-organic nanocomposite films satisfy all the key criteria for a biodegradable scaffold biomaterial — they have an excellent biocompatibility, a high porosity and a controlled and slow degradability. In addition, the scientists show that the material's unique nanotopography, which is produced by filamentous structures branching out from the centres of rough spherulites, promotes cell growth. The combination of organic and inorganic functionalities makes these films superior to conventional PCL-PU films. Highly porous star-shaped polyhedral oligomeric silsesquioxane (POSS) hybrid films were synthesized using POSS as the starting core followed by polycaprolactone (PCL) extension and polyurethane (PU) cross-linking. The unique three-dimensional nanotopography of these films is conducive for cell growth. The combination of favorable factors such as high porosity, reactive surface topography, excellent biocompatibility and biphasic degradation makes the star-shaped POSS-PCL-PU film a great candidate as a tissue engineering scaffold biomaterial.

Purification and biocompatibility of fermented hyaluronic acid for its applications to biomaterials.

BackgroundHyaluronic acid (HA) is of importance due to its diverse applications in pharmaceuticals and medical devices such as dermal filler, adhesion barriers, carrier for cells and bioactive molecules as well as scaffold biomaterials for tissue engineering. Evaluations of purification and biocompatibility of HA are required for its applications to biomaterials.ResultsAfter synthesizing HA by fermentation of streptococcus zooepidemicus for 25 hr, extensively purification of the fermented broth was performed to remove impurities using a filtration process for insoluble components and cells, and diverse adsorbents for soluble impurities. Its in vitro biocompatibility has been evaluated by measurement of cell counting and assay of cell live and dead. 60% yield of white HA powder was obtained, having 15–17 dL/g intrinsic viscosity with a molecular weight of approximately 1,000 kDa. While low molecular weight impurities and insoluble impurities were successfully removed using a ultrafiltration membrane with 50 KDa molecular weight cut, endotoxins, high molecular weight proteins and nucleic acids were removed from the broth by employing adsorbents such as alumina and activated carbons. Alumina showed the best results for the removal of endotoxins, all of the activated carbons were very effective in the removal of high molecular weight proteins and nucleic acids. The purified HA solution showed excellent cell compatibility with no cell damages as observed by both measurement of cell proliferation and observation of cell viability.ConclusionsWe obtained high molecular weight HA with excellent biocompatibility as judged by both measurement of cell proliferation and viability, indicating high possibility of its applications to biomaterials.

Customized biomimetic scaffolds created by indirect three-dimensional printing for tissue engineering

Three-dimensional printing (3DP) is a rapid prototyping technique that can create complex 3D structures by inkjet printing of a liquid binder onto powder biomaterials for tissue engineering scaffolds. Direct fabrication of scaffolds from 3DP, however, imposes a limitation on material choices by manufacturing processes. In this study, we report an indirect 3DP approach wherein a positive replica of desired shapes was printed using gelatin particles, and the final scaffold was directly produced from the printed mold. To create patient-specific scaffolds that match precisely to a patient's external contours, we integrated our indirect 3DP technique with imaging technologies and successfully created custom scaffolds mimicking human mandibular condyle using polycaprolactone and chitosan for potential osteochondral tissue engineering. To test the ability of the technique to precisely control the internal morphology of the scaffolds, we created orthogonal interconnected channels within the scaffolds using computer-aided-design models. Because very few biomaterials are truly osteoinductive, we modified inert 3D printed materials with bioactive apatite coating. The feasibility of these scaffolds to support cell growth was investigated using bone marrow stromal cells (BMSC). The BMSCs showed good viability in the scaffolds, and the apatite coating further enhanced cellular spreading and proliferation. This technique may be valuable for complex scaffold fabrication.

Recombinant production of self-assembling β-structured peptides using SUMO as a fusion partner

BackgroundSelf-assembling peptides that form nanostructured hydrogels are important biomaterials for tissue engineering scaffolds. The P11-family of peptides includes, P11-4 (QQRFEWEFEQQ) and the complementary peptides P11-13 (EQEFEWEFEQE) and P11-14 (QQOrnFOrnWOrnFOrnQQ). These form self-supporting hydrogels under physiological conditions (pH 7.4, 140 mM NaCl) either alone (P11-4) or when mixed (P11-13 and P11-14). We report a SUMO-peptide expression strategy suitable for allowing release of native sequence peptide by SUMO protease cleavage.ResultsWe have expressed SUMO-peptide fusion proteins from pET vectors by using autoinduction methods. Immobilised metal affinity chromatography was used to purify the fusion protein, followed by SUMO protease cleavage in water to release the peptides, which were recovered by reverse phase HPLC. The peptide samples were analysed by electrospray mass spectrometry and self-assembly was followed by circular dichroism and transmission electron microscopy.ConclusionsThe fusion proteins were produced in high yields and the β-structured peptides were efficiently released by SUMO protease resulting in peptides with no additional amino acid residues and with recoveries of 46% to 99%. The peptides behaved essentially the same as chemically synthesised and previously characterised recombinant peptides in self-assembly and biophysical assays.

Leveraging "raw materials" as building blocks and bioactive signals in regenerative medicine.

Components found within the extracellular matrix (ECM) have emerged as an essential subset of biomaterials for tissue engineering scaffolds. Collagen, glycosaminoglycans, bioceramics, and ECM-based matrices are the main categories of "raw materials" used in a wide variety of tissue engineering strategies. The advantages of raw materials include their inherent ability to create a microenvironment that contains physical, chemical, and mechanical cues similar to native tissue, which prove unmatched by synthetic biomaterials alone. Moreover, these raw materials provide a head start in the regeneration of tissues by providing building blocks to be bioresorbed and incorporated into the tissue as opposed to being biodegraded into waste products and removed. This article reviews the strategies and applications of employing raw materials as components of tissue engineering constructs. Utilizing raw materials holds the potential to provide both a scaffold and a signal, perhaps even without the addition of exogenous growth factors or cytokines. Raw materials contain endogenous proteins that may also help to improve the translational success of tissue engineering solutions to progress from laboratory bench to clinical therapies. Traditionally, the tissue engineering triad has included cells, signals, and materials. Whether raw materials represent their own new paradigm or are categorized as a bridge between signals and materials, it is clear that they have emerged as a leading strategy in regenerative medicine. The common use of raw materials in commercial products as well as their growing presence in the research community speak to their potential. However, there has heretofore not been a coordinated or organized effort to classify these approaches, and as such we recommend that the use of raw materials be introduced into the collective consciousness of our field as a recognized classification of regenerative medicine strategies.

The effect of molecular design on the physical and biological properties of complementary self-assembling peptides

Self-assembling peptides are important biomaterials for tissue engineering scaffolds. Single peptides such as P 11-4 (QQRFEWEFEQQ) can self-assemble under physiological conditions (pH 7.4, 140 m m NaCl) to form a hydrogel that supports 3D cell proliferation over 28 days. By contrast, P 11-13 (EQEFEWEFEQE) and P 11-14 (QQOFOWOFOQQ) remain monomeric but upon equimolar mixing of P 11-13 and P 11-14, under physiological conditions, an instantaneous self-supporting hydrogel is formed. Analysis of this hydrogel by spectroscopy showed a β-sheet conformation while solutions of P 11-13 and P 11-14 alone adopted a random coil conformation. Transmission electron microscopy identified the formation of fibrils as seen previously with P 11-4. Preliminary in vitro amyloidogenicity studies found that P 11-13/P 11-14 fibrils were more amyloidogenic than P 11-4 fibrils, however, both peptide fibrils were found to be significantly less amyloidogenic than a naturally occurring amyloidogenic protein, β 2-microglobulin. Hydrogels of P 11-13/P 11-14 and of P 11-4 were cytocompatible with primary human dermal fibroblasts. However, when seeded in P 11-13/P 11-14 the fibroblasts declined in cell number over 28 days. By contrast P 11-4 was able to support cell proliferation over 28 days. Histological studies revealed differences between these cell-seeded hydrogel systems. The P 11-13/P 11-14 hydrogel showed a brick-like network of large fibres and cell remnants were observed. By contrast, the P 11-4 hydrogel was much less dense and the cells elaborated neomatrix and ECM components at 14 days. While the properties of P 11-13 and P 11-14 are not ideally suited for supporting cell proliferation they are likely to be of value in other applications.

Computer-aided design and finite-element modelling of biomaterial scaffolds for bone tissue engineering

Scaffold biomaterials for tissue engineering can be produced in many different ways depending on the applications and the materials used. Most research into new biomaterials is based on an experimental trial-and-error approach that limits the possibility of making many variations to a single material and studying its interaction with its surroundings. Instead, computer simulation applied to tissue engineering can offer a more exhaustive approach to test and screen out biomaterials. In this paper, a review of the current approach in biomaterials designed through computer-aided design (CAD) and through finite-element modelling is given. First we review the approach used in tissue engineering in the development of scaffolds and the interactions existing between biomaterials, cells and mechanical stimuli. Then, scaffold fabrication through CAD is presented and characterization of existing scaffolds through computed images is reviewed. Several case studies of finite-element studies in tissue engineering show the usefulness of computer simulations in determining the mechanical environment of cells when seeded into a scaffold and the proper design of the geometry and stiffness of the scaffold. This creates a need for more advanced studies that include aspects of mechanobiology in tissue engineering in order to be able to predict over time the growth and differentiation of tissues within scaffolds. Finally, current perspectives indicate that more efforts need to be put into the development of such advanced studies, with the removal of technical limitations such as computer power and the inclusion of more accurate biological and genetic processes into the developed algorithms.

Novel bioassay to evaluate biocompatibility of bioactive glass scaffolds for tissue engineering

The aim of the present study was to investigate a novel ex ovo bioassay for the first time using the chick embryo chorioallantoic membrane (CAM) for testing tissue engineering bioceramic scaffolds. Bioglass based scaffolds with porosity in the range of 90–95% were fabricated using the foam replica technique and sintering at 1100°C for 1 h. Scaffolds (5 × 5 × 2 mm3) were placed on the CAM at 10 days of total incubation. The embryos were killed 5 days after implantation. The scaffolds and CAM were explanted, fixed in formalin solution and processed for embedding in methyl methacrylate. Histological analysis using ground sections showed that the scaffolds were surrounded by CAM. There was no occurrence of macrophages or related inflammatory cells. The results described in this paper indicate that the developed bioassay is an appropriate approach as an alternative to conventional animal models to evaluate the biocompatibility of scaffold biomaterials for tissue engineering and regenerative medicine.

Composite hydrogels filled with inclusion complexes made from β-cyclodextrins with poly(propylene glycol) bisamine

A novel kind of composite hydrogel filled with inclusion complexes (ICs) made from β-cyclodextrins (β-CDs) with amine-terminated poly(propylene glycol) (PPG-BA) was prepared by copolymerization of methacryloyl end-capped poly( l -lactide)-poly(ethylene glycol)-poly-( l -lactide) macromer (DMPEGL) with acrylic acid (AA) in water under UV irradiation. Owing to the hydrogen bonding interaction of –COOH in AA on the network backbones with –NH 2 and –OH in ICs embedded in the crosslinked network, the composite hydrogels are different from those without ICs in their swelling and degradation behaviors. Furthermore, adding ICs into the network results in the mechanical properties being significantly improved. The mass loss test conducted in water indicates that they degrade by following a dual degradation mechanism, that is, not only β-CDs slipping from the polymer axles and then escaping out of the hydrogel network, but also the bulk hydrogel network hydrolyzing itself to lose the weight. These composite hydrogels show potential to be used as biomaterials for tissue engineering scaffolds and carriers of drugs with controlled release.

Biomaterial films of Bombyx mori silk fibroin with poly(ethylene oxide).

Phase separation into controllable patterned microstructures was observed for Bombyx mori silkworm silk and poly(ethylene oxide) (PEO) (900000 g/mol) blends cast from solution. The evolution of the microstructures with increasing PEO volume fraction is strikingly similar to the progression of phases and microstructures observed with surfactants. The chemically patterned materials obtained provide engineerable biomaterial surfaces with predictable microscale features which can be used to create topographically patterned or chemically functionalized biomaterials. Solution blending was used to incorporate water-soluble PEO into silk to enhance elasticity and hydrophilicity. The sizes of the globule fibroin phase ranged from 2.1 +/- 0.5 to 18.2 +/- 2.1 microm depending on the ratio of silk/PEO. Optical microscopy and SEM analysis confirmed the micro-phase separation between PEO and silk. Surface properties were determined by XPS and contact angle. Methanol can be used to control the conformational transition of silk fibroin to the insoluble beta-sheet state. Subsequentially, the PEO can be easily extracted from the films with water to generate silk matrixes with definable porosity and enhanced surface roughness. These blend films formed from two biocompatible polymers provide potential new biomaterials for tissue engineering scaffolds.