Synthetic vs natural scaffolds for human limbal stem cells.

AIM: To investigate the impact of synthetic electrospun polyurethane (PU) and polycaprolactone (PCL) nanoscaffolds, before and after hydrolytic surface modification, on viability and differentiation of cultured human eye epithelial cells, in comparison with natural scaffolds: fibrin and human amniotic membrane. METHODS: Human placenta was taken at elective cesarean delivery. Fibrin scaffolds were prepared from commercial fibrin glue kits. Nanoscaffolds were fabricated by electrospinning. Limbal cells were isolated from surpluses of human cadaveric cornea and seeded on feeder 3T3 cells. The scaffolds used for viability testing and immunofluorescence analysis were amniotic membrane, fibrin, PU, and PCL nanoscaffolds, with or without prior NaOH treatment. RESULTS: Scanning electron microscope photographs of all tested scaffolds showed good colony spreading of seeded limbal cells. There was a significant difference in viability performance between cells with highest viability cultured on tissue culture plastic and cells cultured on all other scaffolds. On the other hand, electrospun PU, PCL, and electrospun PCL treated with NaOH had more than 80% of limbal cells positive for stem cell marker p63 compared to only 27%of p63 positive cells on fibrin. CONCLUSION: Natural scaffolds, fibrin and amniotic membrane, showed better cell viability than electrospun scaffolds. On the contrary, high percentages of p63 positive cells obtained on these scaffolds still makes them good candidates for efficient delivery systems for therapeutic purposes.

Development of hybrid braided composite rods for reinforcement and health monitoring of structures.

In the present study, core-reinforced braided composite rods (BCRs) were developed and characterized for strain sensing capability. A mixture of carbon and glass fibre was used in the core, which was surrounded by a braided cover of polyester fibres. Three compositions of core with different carbon fibre/glass fibre weight ratios (23/77, 47/53, and 100/0) were studied to find out the optimum composition for both strain sensitivity and mechanical performance. The influence of carbon fibre positioning in BCR cross-section on the strain sensing behaviour was also investigated. Strain sensing property of BCRs was characterized by measuring the change in electrical resistance with flexural strain. It was observed that BCRs exhibited increase (positive response) or decrease (negative response) in electrical resistance depending on carbon fibre positioning. The BCR with lowest amount of carbon fibre was found to give the best strain sensitivity as well as the highest tensile strength and breaking extension. The developed BCRs showed reversible strain sensing behaviour under cyclic flexural loading with a maximum gauge factor of 23.4 at very low strain level (0.55%). Concrete beams reinforced with the optimum BCR (23/77) also exhibited strain sensing under cyclic flexural strain, although the piezoresistive behaviour in this case was irreversible.

Detection of Limbal Stem Cells Adhered to Melt Electrospun Silk Fibroin and Gelatin-Modified Polylactic Acid Scaffolds.

Limbal stem cells (LSCs) are of paramount importance in corneal epithelial tissue repair. The cornea becomes opaque in case of limbal stem cell deficiency (LSCD), which may cause serious damage to the ocular visual function. There are many techniques to restore damaged epithelium, one of which is the transplantation of healthy cultured LSCs, usually onto a human amniotic membrane or onto bio-based engineered scaffolds in recent years. In this study, melt electrospun polylactic acid (PLA) was modified by silk fibroin or gelatin and further cultured with LSCs originating from three different donors. In terms of physicochemical properties, both modifications slightly increased PLA scaffold porosity (with a significantly larger pore area for the PLA/gelatin) and improved the scaffolds' swelling percentage, as well as their biodegradation rate. In terms of the scaffold application function, the aim was to detect/visualize whether LSCs adhered to the scaffolds and to further determine cell viability (total number), as well as to observe p63 and CK3 expressions in the LSCs. LSCs were attached to the surface of microfibers, showing flattened conformations or 3D spheres in the formation of colonies or agglomerations, respectively. All scaffolds showed the ability to bind the cells onto the surface of individual microfibers (PLA and PLA/gelatin), or in between the microfibers (PLA/silk fibroin), with the latter showing the most intense red fluorescence of the stained cells. All scaffolds proved to be biocompatible, while the PLA/silk fibroin scaffolds showed the highest 98% viability of 2.9 × 106 LSCs, with more than 98% of p63 and less than 20% of CK3 expressions in the LSCs, thus confirming the support of their growth, proliferation and corneal epithelial differentiation. The results show the potential of these bio-engineered scaffolds to be used as an alternative clinical approach.

Agro-Industrial Plant Proteins in Electrospun Materials for Biomedical Application.

Plant proteins are receiving a lot of attention due to their abundance in nature, customizable properties, biodegradability, biocompatibility, and bioactivity. As a result of global sustainability concerns, the availability of novel plant protein sources is rapidly growing, while the extensively studied ones are derived from byproducts of major agro-industrial crops. Owing to their beneficial properties, a significant effort is being made to investigate plant proteins' application in biomedicine, such as making fibrous materials for wound healing, controlled drug release, and tissue regeneration. Electrospinning technology is a versatile platform for creating nanofibrous materials fabricated from biopolymers that can be modified and functionalized for various purposes. This review focuses on recent advancements and promising directions for further research of an electrospun plant protein-based system. The article highlights examples of zein, soy, and wheat proteins to illustrate their electrospinning feasibility and biomedical potential. Similar assessments with proteins from less-represented plant sources, such as canola, pea, taro, and amaranth, are also described.

The Reliability of PCL/Anti-VEGF Electrospun Scaffolds to Support Limbal Stem Cells for Corneal Repair.

Since only few reported studies propose anti-vascular endothelial growth factor (anti-VEGF) delivery through electrospun scaffolds, this study greatly contributes to the potential prevention of patient's vision loss, as it explores electrospun polycaprolactone (PCL) coated with anti-VEGF for the blockage of abnormal cornea vascularization. In terms of physicochemical properties, the biological component increased the PCL scaffold fiber diameter (by ~24%) and pore area (by ~82%), while ut slightly reduced its total porosity as the anti-VEGF solution filled the voids of the microfibrous structure. The addition of the anti-VEGF increased the scaffold stiffness almost three-fold at both strains of 5 and 10%, as well as its biodegradation rate (~36% after 60 days) with a sustained release profile after Day 4 of phosphate buffered saline incubation. In terms of scaffold application function, the PCL/Anti-VEGF scaffold proved to be more favorable for the adhesion of cultured limbal stem cells (LSCs); this was confirmed by the SEM images, where the cells showed flat and elongated conformations. Further support of the LSC growth and proliferation was confirmed by the identified p63 and CK3 markers after cell staining. These results demonstrate the advantageous effect of the surface-adsorbed anti-VEGF to stop vision loss and help damaged corneal tissue repair.

Bioactivity Comparison of Electrospun PCL Mats and Liver Extracellular Matrix as Scaffolds for HepG2 Cells.

Cells grown on bioactive matrices have immensely advanced many aspects of biomedical research related to drug delivery and tissue engineering. Our main objective was to perform simple evaluation of the structural and biotic qualities of cell scaffolds made of affordable biomaterials for liver cell line (HepG2) cultivation in vitro. In this work the electrospun matrix made of synthetic polyester poly(ε-caprolactone) (PCL) was compared with the natural protein-based extracellular matrix isolated from porcine liver (ECM). Mechanical and structural analysis showed that ECM was about 12 times less resistant to tensile stress while it had significantly larger pore size and twice smaller water contact angle than PCL. Bioactivity assessment included comparison of cell growth and transfection efficiency on cell-seeded scaffolds. Despite the differences in composition and structure between the two respective matrices, the rate of cell spreading and the percentage of transfected cells on both scaffolds were fairly comparable. These results suggest that in an attempt to produce simple, cell carrying structures that adequately simulate the natural scaffold, one can rely on PCL electrospun mats.

Poly(ε-caprolactone) Titanium Dioxide and Cefuroxime Antimicrobial Scaffolds for Cultivation of Human Limbal Stem Cells.

Limbal Stem Cell Deficiency (LSCD) is a very serious and painful disease that often results in impaired vision. Cultivation of limbal stem cells for clinical application is usually performed on carriers such as amniotic membrane or surgical fibrin gel. Transplantation of these grafts is associated with the risk of local postoperative infection that can destroy the graft and devoid therapeutic benefit. For this reason, electrospun scaffolds are good alternatives, as proven to mimic the natural cells surroundings, while their fabrication technique is versatile with regard to polymer functionalization and scaffolds architecture. This study considers the development of poly(ε-caprolactone) (PCL) immune-compatible and biodegradable electrospun scaffolds, comprising cefuroxime (CF) or titanium dioxide (TiO2) active components, that provide both bactericidal activity against eye infections and support of limbal stem cells growth in vitro. The PCL/CF scaffolds were prepared by blend electrospinning, while functionalization with the TiO2 particles was performed by ultrasonic post-processing treatment. The fabricated scaffolds were evaluated in regard to their physical structure, wetting ability, static and dynamic mechanical behaviour, antimicrobial efficiency and drug release, through scanning electron microscopy, water contact angle measurement, tensile testing and dynamic mechanical analysis, antimicrobial tests and UV-Vis spectroscopy, respectively. Human limbal stem cells, isolated from surgical remains of human cadaveric cornea, were cultured on the PCL/CF and PCL/TiO2 scaffolds and further identified through immunocytochemistry in terms of cell type thus were stained against p63 marker for limbal stem cells, a nuclear transcription factor and cytokeratin 3 (CK3), a corneal epithelial differentiation marker. The electrospun PCL/CF and PCL/TiO2 successfully supported the adhesion, proliferation and differentiation of the cultivated limbal cells and provided the antimicrobial effect against Pseudomonas aeruginosa, Staphylococcus aureus and Candida albicans.