Effect of feeder free poly(lactide-co-glycolide) scaffolds on morphology, proliferation, and pluripotency of mouse embryonic stem cells.

The aim of the study has been to evaluate the morphology, proliferation, and pluripotency maintenance of mouse embryonic stem cells (mESCs) cultivated on poly(lactic-co-glycolic acid) scaffolds. The scaffolds were hydrolyzed with NaOH (treated) and nonhydrolyzed (untreated). Morphological and mechanical characterization of the scaffolds was performed. mESC were evaluated for cell viability, cytotoxicity, expression of pluripotency markers, colony morphology, and overall distribution. The treatment generated a reduction in the hydrophobic characteristics of the scaffolds, leading to a higher wettability compared to the untreated group. The viability, cytotoxicity, number of colonies, and the thickness of the cell layer presented similar results between the scaffold groups. The viability test showed that it was possible to cultivate the mESCs on the scaffolds. The cytotoxicity analysis showed that the PLGA scaffolds were not harmful for the cells. The cells maintained the expression of the pluripotency markers Oct4 and Sox2. The number of colonies and the thickness of the cell layer on the scaffold showed that they were not able to colonize the entire volume of the scaffolds. The area occupied by the mESCs was the same between the treated and untreated groups after 14 days in culture. It is possible to conclude that both conditions are equally suitable for maintaining mESC culture. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 424-432, 2017.

Advantages and challenges offered by biofunctional core-shell fiber systems for tissue engineering and drug delivery.

Whereas highly porous scaffolds composed of electrospun nanofibers can mimick major features of the extracellular matrix in tissue engineering, they lack the ability to incorporate and release biocompounds (drugs, growth factors) safely in a controlled way. Here, electrospun core-shell fibers (core made from water and aqueous solutions of hydrophilic polymers and the shell from materials with well-defined release mechanisms) offer unique advantages in comparison with those that have helped make porous nanofibrillar scaffolds highly successful in tissue engineering. This review considers the preparation and biofunctionalization of such core-shell fibers as well as applications in various areas, including neural, vascular, cardiac, cartilage and bone tissue engineering, and touches on the topic of clinical trials.

Biofunctionalized nanofibers using Arthrospira (Spirulina) biomass and biopolymer.

Electrospun nanofibers composed of polymers have been extensively researched because of their scientific and technical applications. Commercially available polyhydroxybutyrate (PHB) and polyhydroxybutyrate-co-valerate (PHB-HV) copolymers are good choices for such nanofibers. We used a highly integrated method, by adjusting the properties of the spinning solutions, where the cyanophyte Arthrospira (formally Spirulina) was the single source for nanofiber biofunctionalization. We investigated nanofibers using PHB extracted from Spirulina and the bacteria Cupriavidus necator and compared the nanofibers to those made from commercially available PHB and PHB-HV. Our study assessed nanofiber formation and their selected thermal, mechanical, and optical properties. We found that nanofibers produced from Spirulina PHB and biofunctionalized with Spirulina biomass exhibited properties which were equal to or better than nanofibers made with commercially available PHB or PHB-HV. Our methodology is highly promising for nanofiber production and biofunctionalization and can be used in many industrial and life science applications.

Mesenchymal stem cell adherence on poly(D, L-lactide-co-glycolide) nanofibers scaffold is integrin-beta 1 receptor dependent.

Tissue engineering is a potential approach to regenerate damaged tissue by the combination and synergism among the scaffolding material, cell source and signaling factors. In the present study, mesenchymal stem cells (MSCs) were isolated from C57BL/6 mice, cultured on poly(D, L-lactide-co-glycolide) (PLGA) scaffold produced by electrospinning technique and differentiated into chondrogenic lineage. After seeding, MSCs were responsive and became flattened with fibroblast-like morphology demonstrated by the presence of actin stress fibers. Integrin-beta1 receptor blockage reduced significantly cell adhesion with loss of actin stress fibers, demonstrating the ability of PLGA nanofiber to trigger integrin receptor-mediated cell adhesion. Present data contribute to the understanding of MSCs' behavior on these biodegradable and biocompatible scaffolds that can be used as carriers in treatments involving cell transplantation.

A novel globular protein electrospun fiber mat with the addition of polysilsesquioxane.

The aim of this work has been to elaborate well defined gliadin nanofibers with incorporation of inorganic molecules, such as polyhedral oligomeric silsesquioxane (POSS). Nanofibers were obtained by electrospinning processing, controlling the relevant parameters such as tip-to-collector distance, voltage and feed rate. The fiber mats were characterized by SEM, confocal images, DSC, viscosity, FTIR and conductivimetry analysis. FTIR spectra showed characteristic absorption bands related to the presence of POSS-NH(2) within the matrices. SEM micrographs showed that gliadin fibers decreased their dimensions as the amount of POSS-NH(2) increased in the spinning solution. The electrical conductivity of gliadin solutions diminished as the concentration of POSS-NH(2) was increased. Besides, confocal micrographs revealed that POSS-NH(2) might be dispersed as nanocrystals into gliadin and gluten fibers. The dimension of gluten nanofibers was also affected by the POSS-NH(2) concentration, but conversely, this dependence was not proportional to the POSS-NH(2) amount. Somehow, the interaction between gliadin and POSS-NH(2) in aqueous TFE affected the solution viscosity and, as a consequence, higher jet instabilities and thinner fiber dimensions were obtained.

Polymer-assisted preparation of nanoscale films of thermoelectric PbSe and PbTe and of lead chalcogenide-polymer composite films.

Polymer films of polyethyleneoxide (PEO) or poly(L-lactide) (PLLA) containing a single-source precursor for either PbSe or PbTe were used to produce films of nanoparticles of these thermoelectric materials. The monomeric homoleptic chalcogenolates lead(II) bis-(2,4,6-trifluoromethylphenylselenolate) Pb[SeC(6)H(2)(CF(3))(3)](2) and lead(II) bis-[tris(trimethylsilyl)silyl-tellurolate] Pb[TeSi(SiMe(3))(3)](2) were used as single-source precursors for the thermolytic formation of the lead chalcogenides. The thickness and the quality of as-obtained thin films depended decisively on the spin-coating conditions, on the polymer, on the precursor concentration in the composite film before thermolysis and on the annealing time. Thin layers of particles of 30-50 nm size and high crystallinity were obtained. They were characterized by X-ray diffraction, thermal analysis and electron microscopy.

Preparation of nanofibers containing the microalga Spirulina (Arthrospira).

Spirulina is a microalga which offers biological functions highly favorable for tissue engineering. Highly porous scaffolds can be produced by electrospinning containing biomass of Spirulina. The goal of this contribution was therefore to establish spinning conditions allowing to produce well defined nanofibers with diameters down to about 100 nm and to produce nanofibers with various concentration of the biomass for subsequent studies in tissue engineering applications. The experimental results reveal that the blend system PEO/biomass is behaved surprisingly well in electrospinning. Very thin bead-free nanofibers with diameters of about 110 nm can be produced for different biomass contents of up to 67 wt.% of the nanofibers and for PEO concentrations in the spinning solution well below 4 wt.%. These results suggest to us the use of the biomass containing nanofibers as extracellular matrices for stem cell culture and future treatment of spinal chord injury.

Chemistry on electrospun polymeric nanofibers: merely routine chemistry or a real challenge?

Nanofiber-based non-wovens can be prepared by electrospinning. The chemical modification of such nanofibers or chemistry using nanofibers opens a multitude of application areas and challenges. A wealth of chemistry has been elaborated in recent years on and with electrospun nanofibers. Known methods as well as new methods have been applied to modify the electrospun nanofibers and thereby generate new materials and new functionalities. This Review summarizes and sorts the chemistry that has been reported in conjunction with electrospun nanofibers. The major focus is on catalysis and nanofibers, enzymes and nanofibers, surface modification for biomedical and specialty applications, coatings of fibers, crosslinking, and bulk modifications. A critical focus is on the question: what could make chemistry on or with nanofibers different from bulk chemistry?

RGD-functionalisation of PLLA nanofibers by surface coupling using plasma treatment: influence on stem cell differentiation.

The aim of this study was to functionalize the surface of synthetic poly-(l-lactic) (PLLA) nanofibers with RGD peptide, in order to promote growth and osteogenic differentiation of human mesenchymal stem cells (hMSC) in vitro. The cRGD was coupled onto PLLA nanofibers using oxygen plasma combined with EDC/sulfo-NHS activation. Matrices were seeded with hMSC and cultivated over a period of 22 days under growth conditions and analyzed during the course of cultivation. The plasma activation of PLLA nanofibers resulted in a reduction of hydrophobicity as well as a formation of carboxyl groups on the surface of the fibers. Furthermore, maximum load, but not young's modulus was influenced by the treatment with oxygen plasma. When hMSC were cultured onto the cRGD functionalized scaffolds, cells showed no increased proliferation or cell density but an induction of genes associated with the osteoblast lineage. In brief, this study indicates that functional peptides of the extracellular matrix can be coupled onto PLLA nanofibers using plasma treatment in combination with EDC/sulfo-NHS treatment. These groups are accessible for the growing cell and mediate probably some osteoinductive properties of collagen nanofibers.

Influence of poly-(L-lactic acid) nanofiber functionalization on maximum load, Young's modulus, and strain of nanofiber scaffolds before and after cultivation of osteoblasts: an in vitro study.

The aim of this study was to characterize the influence of functionalization of synthetic poly-(L-lactic acid) (PLLA) nanofibers on mechanical properties such as maximum load, elongation, and Young's modulus. Furthermore, the impact of osteoblast growth on the various nanofiber scaffolds stability was determined. Nanofiber matrices composed of PLLA, PLLA-collagen, or BMP-2-incorporated PLLA were produced from different solvents by electrospinning. Standardized test samples of each nanofiber scaffold were subjected to failure protocol before or after incubation in the presence of osteoblasts over a period of 22 days under osteoinductive conditions. PLLA nanofibers electrospun from hexafluoroisopropanol (HFIP) showed a higher strain and tended to have increased maximum loads and Young s modulus compared to PLLA fibers spun from dichloromethane. In addition, they had a higher resistance during incubation in the presence of cells. Functionalization by incorporation of growth factors increased Young's modulus, independent of the solvent used. However, the incorporation of growth factors using the HFIP system resulted in a loss of strain. Similar results were observed when PLLA was blended with different ratios of collagen. Summarizing the results, this study indicates that different functionalization strategies influence the mechanical stability of PLLA nanofibers. Therefore, an optimization of nanofibers should not only account for the optimization of biological effects on cells, but also has to consider the stability of the scaffold.

Collagen matrices from sponge to nano: new perspectives for tissue engineering of skeletal muscle.

BACKGROUND: Tissue engineering of vascularised skeletal muscle is a promising method for the treatment of soft tissue defects in reconstructive surgery. In this study we explored the characteristics of novel collagen and fibrin matrices for skeletal muscle tissue engineering. We analyzed the characteristics of newly developed hybrid collagen-I-fibrin-gels and collagen nanofibers as well as collagen sponges and OPLA-scaffolds. Collagen-fibrin gels were also tested with genipin as stabilizing substitute for aprotinin. RESULTS: Whereas rapid lysis and contraction of pure collagen I- or fibrin-matrices have been great problems in the past, the latter could be overcome by combining both materials. Significant proliferation of cultivated myoblasts was detected in collagen-I-fibrin matrices and collagen nanofibers. Seeding cells on parallel orientated nanofibers resulted in strongly aligned myoblasts. In contrast, common collagen sponges and OPLA-scaffolds showed less cell proliferation and in collagen sponges an increased apoptosis rate was evident. The application of genipin caused deleterious effects on primary myoblasts. CONCLUSION: Collagen I-fibrin mixtures as well as collagen nanofibers yield good proliferation rates and myogenic differentiation of primary rat myoblasts in vitro In addition, parallel orientated nanofibers enable the generation of aligned cell layers and therefore represent the most promising step towards successful engineering of skeletal muscle tissue.

Effect of direct RGD incorporation in PLLA nanofibers on growth and osteogenic differentiation of human mesenchymal stem cells.

The aim of this study was to functionalize synthetic poly-(L-lactic) (PLLA) nanofibers by direct incorporation of cRGD, in order to promote adhesion, growth and osteogenic differentiation of human mesenchymal stem cells (hMSC) in vitro. The cRGD was incorporated into PLLA nanofibers either by emulsion [PLLA-cRGD (d)] or suspension [PLLA-cRGD (s)]. Matrices were seeded with hMSC and cultivated over a period of 28 days under growth conditions and analyzed during the course. Although the mode of incorporation resulted in different distributions of the RGD peptide, it had no impact on the fiber characteristics when compared to corresponding unblended PLLA control fibers. However, hMSC showed better adherence on PLLA-cRGD (d). Nevertheless, this advantage was not reflected during the course of cultivation. Furthermore, the PLLA-cRGD (s) fibers mediated the osteogenic potential of collagen (determined as the expression and deposition of collagen and osteocalcin) to some extent. Further studies are needed in order to optimize the RGD distribution and concentration.

Characterization of a PLLA-collagen I blend nanofiber scaffold with respect to growth and osteogenic differentiation of human mesenchymal stem cells.

We developed a nanofiber scaffold by blending PLLA with collagen I, suitable for bone regeneration. Among several PLLA-COLI ratios tested, cell growth was better enhanced when blends with a ratio of PLLA-COLI 4:1 were used. Here, growth as well as osteoblast differentiation of hMSC was improved when compared to PLLA nanofibers alone. Therefore, blending is a suitable tool to enhance PLLA nanofibers with respect to bone tissue engineering.

Influence of nanofibers on the growth and osteogenic differentiation of stem cells: a comparison of biological collagen nanofibers and synthetic PLLA fibers.

The aim of this study was to compare biological collagen I (ColI) and synthetic poly-(L: -lactide) (PLLA) nanofibers concerning their stability and ability to promote growth and osteogenic differentiation of human mesenchymal stem cells in vitro. Matrices were seeded with human stem cells and cultivated over a period of 28 days under growth and osteoinductive conditions and analyzed during the course. During this time the PLLA nanofibers remained stable while the presence of cells resulted in an attenuation of the ColI nanofiber mesh. Although there was a tendency for better growth and osteoprotegerin production of stem cells when cultured on collagen nanofibers, there was no significant difference compared to PLLA nanofibers or controls. The gene expression of alkaline phosphate, osteocalcin and collagen I diminished in the initial phase of cultivation independent of the polymer used. In the case of PLLA fibers, this gene expression normalized during the course of cultivation, whereas the presence of collagen nanofibers resulted in an increased gene expression of osteocalcin and collagen during the course of the experiment. Taken together the PLLA fibers were easier to produce, more stable and did not compromise growth and differentiation of stem cells over the course of experiment. On the other hand, collagen nanofibers supported the differentiation process to some extent. Nevertheless, the need for fixation as well as the missing stability during cell culture requires further work.

Progress in the field of electrospinning for tissue engineering applications.

Electrospinning is an extremely promising method for the preparation of tissue engineering (TE) scaffolds. This technique provides nonwovens resembling in their fibrillar structures those of the extracellular matrix (ECM), and offering large surface areas, ease of functionalization for various purposes, and controllable mechanical properties. The recent developments toward large-scale productions combined with the simplicity of the process render this technique very attractive. Progress concerning the use of electrospinning for TE applications has advanced impressively. Different groups have tackled the problem of electrospinning for TE applications from different angles. Nowadays, electrospinning of the majority of biodegradable and biocompatible polymers, either synthetic or natural, for TE applications is straightforward. Different issues, such as cell penetration, incorporation of growth and differentiating factors, toxicity of solvents used, productivity, functional gradient, etc. are main points of current considerations. The progress in the use of electrospinning for TE applications is highlighted in this article with focus on major problems encountered and on various solutions available until now.

Influence of poly(L-lactic acid) nanofibers and BMP-2-containing poly(L-lactic acid) nanofibers on growth and osteogenic differentiation of human mesenchymal stem cells.

The aim of this study was to characterize synthetic poly-(L-lactic acid) (PLLA) nanofibers concerning their ability to promote growth and osteogenic differentiation of stem cells in vitro, as well as to test their suitability as a carrier system for growth factors. Fiber matrices composed of PLLA or BMP-2-incorporated PLLA were seeded with human mesenchymal stem cells and cultivated over a period of 22 days under growth and osteoinductive conditions, and analyzed during the course of culture, with respect to gene expression of alkaline phosphatase (ALP), osteocalcin (OC), and collagen I (COL-I). Furthermore, COL-I and OC deposition, as well as cell densities and proliferation, were analyzed using fluorescence microscopy. Although the presence of nanofibers diminished the dexamethasone-induced proliferation, there were no differences in cell densities or deposition of either COL-I or OC after 22 days of culture. The gene expression of ALP, OC, and COL-I decreased in the initial phase of cell cultivation on PLLA nanofibers as compared to cover slip control, but normalized during the course of cultivation. The initial down-regulation was not observed when BMP-2 was directly incorporated into PLLA nanofibers by electrospinning, indicating that growth factors like BMP-2 might survive the spinning process in a bioactive form.

Electrospinning: a fascinating method for the preparation of ultrathin fibers.

Electrospinning is a highly versatile method to process solutions or melts, mainly of polymers, into continuous fibers with diameters ranging from a few micrometers to a few nanometers. This technique is applicable to virtually every soluble or fusible polymer. The polymers can be chemically modified and can also be tailored with additives ranging from simple carbon-black particles to complex species such as enzymes, viruses, and bacteria. Electrospinning appears to be straightforward, but is a rather intricate process that depends on a multitude of molecular, process, and technical parameters. The method provides access to entirely new materials, which may have complex chemical structures. Electrospinning is not only a focus of intense academic investigation; the technique is already being applied in many technological areas.

Polymer fibers as carriers for homogeneous catalysts.

This paper describes a polymer fiber-based approach for the immobilization of homogeneous catalysts. The goal is to generate products that are free of catalysts which would be of great importance for the development of optoelectronic or pharmaceutical compounds. Electrospinning was employed to prepare the non-woven fiber assembly composed of polystyrene. The homogeneous catalyst scandium triflate was immobilized on the polystyrene fibers during electrospinning and on corresponding core shell fibers using a fiber template approach. An imino aldol and an aza-Diels-Alder model reaction were carried out with each fibrous catalytic system. This resulted in the immobilization of homogeneous catalysts in a polymer environment without loss of their catalytic activity and may even be enhanced when compared with reactions carried out in homogeneous solutions.

Nondestructive mechanical release of ordered polymer microfiber arrays from porous templates.

The fabrication of one-dimensional (1D) nanostructures and microstructures inside the pores of porous templates is intensively investigated. The release of these structures is commonly accomplished by etching and destroying the templates. The 1D nanostructures and microstructures tend to condense because of the occurrence of capillary forces during drying of the specimens. It is shown that highly ordered arrays of polymer microfibers can be easily detached from silanized porous templates by mechanical lift-off. This procedure leaves the templates intact, thus allowing their recycling, and does not involve the use of solutions or solvents, thus circumventing condensation. Therefore, mechanical lift-off may enable the up-scaling of template-based approaches to the fabrication of highly ordered assemblies of 1D nanostructures and microstructures.

Electrospinning approaches toward scaffold engineering--a brief overview.

Tissue engineering involves the in vitro seeding of cells onto scaffolds which assume the role of supporting cell adhesion, migration, proliferation, and differentiation, and which define the three-dimensional shape of the tissue to be engineered. Among the various types of scaffold architectures available, scaffolds based on nanofibers mimicking to a certain extent the structure of the extracellular matrix offer great advantages. Electrospinning is the technique of choice for the preparation of such scaffolds. Investigations have revealed that the nanofibrous structure promotes cell adhesion, proliferation, and differentiation. Parameters relevant for these processes such as fiber diameters, surface topology, porosity, mechanical properties, and the fibrous architecture of the scaffold can be controlled by electrospinning in a broad range.