Controlling integrin-based adhesion to a degradable electrospun fibre scaffold via SI-ATRP (original) (raw)
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Role of electrospun fibre diameter and corresponding specific surface area (SSA) on cell attachment
Journal of Tissue Engineering and Regenerative Medicine, 2009
In order to develop scaffolds for tissue regeneration applications, it is important to develop an understanding of the kinetics of cell attachment as a function of scaffold geometry. In the present study, we investigated how the specific surface area of electrospun scaffolds affected cell attachment and spreading. Number of cells attached to the scaffold was measured by the relative intensity of a metabolic dye (MTS) and cell spreading was analysed for individual cells by measuring the area of projected F-actin cytoskeleton. We varied the fibre diameter to obtain a specific surface area distribution in the range 2.24-18.79 µm −1. In addition, we had one case where the scaffolds had beads in them and therefore had non-uniform fibres. For each of these different geometries, we varied the cell-seeding density (0.5-1 × 10 5) and the serum concentration (0-12%) over the first 8 h in an electrospun polycaprolactone NIH 3T3 fibroblast system. Cells on beaded scaffolds showed the lowest attachment and almost no F-actin spreading in all experiments indicating uniform fibre diameter is essential for electrospun scaffolds. For the uniform fibre scaffolds, cell attachment was a function of scaffold specific surface area (SSA) (18.79-2.24 µm −1) and followed two distinct trends: when scaffold SSA was <7.13 µm −1 , cell adhesion rate remained largely unchanged; however, for SSA >7.13 µm −1 there was a significant increase in cellular attachment rate with increasing SSA. This indicated that nanofibrous scaffolds increased cellular adhesion compared to microfibrous scaffolds. This phenomenon is true for serum concentrations of 7.5% and higher. For 5% and lower serum concentration, cell attachment is low and higher SSA fails to make a significant improvement in cell attachment. When cell attachment was investigated at a single-cell level by measuring the projected actin area, a similar trend was noted where the effect of higher SSA led to higher projected area for cells at 8 h. These results indicate that uniform electrospun scaffolds with SSA provide a faster cell attachment compared to lower SSA and beaded scaffolds. These results indicate that continuous electrospun nanofibrous scaffolds may be a good substrate for rapid tissue regeneration.
Low fouling electrospun scaffolds with clicked bioactive peptides for specific cell attachment
Biomacromolecules, 2015
While electrospun fibers are of interest as scaffolds for tissue engineering applications, nonspecific surface interactions such as protein adsorption often prevent researchers from controlling the exact interactions between cells and the underlying material. In this study we prepared electrospun fibers from a polystyrene-based macroinitiator, which were then grafted with polymer brushes using surfaceinitiated atom transfer radical polymerization (SI-ATRP). These brush coatings incorporated a trimethylsilyl-protected PEG-alkyne monomer, allowing azide functional molecules to be covalently attached, while simultaneously reducing nonspecific protein adsorption on the fibers. Cells were able to attach and spread on fibrous substrates functionalized with a pendant RGD-containing peptide, while spreading was significantly reduced on nonfunctionalized fibers and those with the equivalent RGE control peptide. This effect was observed both in the presence and absence of serum in the culture media, indicating that protein adsorption on the fibers was minimal and cell adhesion within the fibrous scaffold was mediated almost entirely through the cell-adhesive RGD-containing peptide.
Biointerphases, 2013
Background: The ability to present signalling molecules within a low fouling 3D environment that mimics the extracellular matrix is an important goal for a range of biomedical applications, both in vitro and in vivo. Cell responses can be triggered by non-specific protein interactions occurring on the surface of a biomaterial, which is an undesirable process when studying specific receptor-ligand interactions. It is therefore useful to present specific ligands of interest to cell surface receptors in a 3D environment that minimizes non-specific interactions with biomolecules, such as proteins. Method: In this study, surface-initiated atom transfer radical polymerization (SI-ATRP) of poly(ethylene glycol)-based monomers was carried out from the surface of electrospun fibers composed of a styrene/vinylbenzyl chloride copolymer. Surface initiated radical addition-fragmentation chain transfer (SI-RAFT) polymerisation was also carried out to generate bottle brush copolymer coatings consisting of poly(acrylic acid) and poly(acrylamide). These were grown from surface trithiocarbonate groups generated from the chloromethyl styrene moieties existing in the original synthesised polymer. XPS was used to characterise the surface composition of the fibers after grafting and after coupling with fluorine functional XPS labels.
Nanocellulose and Elastin Act as Plasticizers of Electrospun Bioinspired Scaffolds
In this work, we produced cross-linked electrospun hybrid scaffolds composed of gelatin/poly-D,L-lactide (PDLLA), gelatin/PDLLA/ nanocellulose, and gelatin/PDLLA/cellulose nanocrystals/elastin. Fouriertransform infrared spectroscopy, X-ray diffraction, and high-performance liquid chromatography demonstrated the complete embedding of each component in the hybrid scaffolds. The degree of cross-linking was quantified by the 2,4,6-trinitrobenzenesulfonic acid assay, and attenuated total reflectance spectroscopy revealed the effectiveness of the cross-linking reaction. Notably, the interconnected porous structure revealed in uncrosslinked scaffolds persisted even after cross-linking. Scaffolds were characterized in water through their contact angle showing total wettability. We investigated their mechanical properties by uniaxial tensile testing, which showed that even in the dry state, nanocellulose-and elastin-containing scaffolds exhibit higher elongation at rupture compared to those with pure gelatin/PDLLA. Therefore, we succeeded in tuning the toughness of the scaffolds by modulating the composition. In order to use scaffolds as medical devices, we assayed fibroblasts on scaffold extraction media, indicating that they were noncytotoxic. Finally, the attachment and proliferation of fibroblasts on the surface of different scaffolds were evaluated.
Tissue Engineering, 2007
The goal of the current study was to find the quantitative relationship between electrospun polycaprolactone scaffold fiber diameter and NIH 3T3 fibroblast adhesion and growth kinetics. By varying 3 important process parameters-solution concentration, voltage, and collector screen distance-different average fiber diameters ranging from 117 to 1,647 nm were obtained. Although 117 nm represented the lowest possible fiber diameter obtainable, these fibers had beads in them. An increase in fiber diameter to 428 nm led to uniform fibers without any beads. Fiber distribution pattern was a single mode for all the scaffolds except at the largest-diameter end. The diameter distribution changed from single to bimodal at 1,647 nm, suggesting some instability in the process. It was found that cell adhesion and growth kinetics are significantly affected as a function of fiber diameter. Beaded scaffolds offered the lowest cell adhesion and minimal growth kinetics despite having the lowest average fiber diameter. When uniform fibers were formed and the average fiber was in the nanofiber range (428 -1051 nm), cell adhesion and growth kinetics decreased as a function of increasing fiber diameter. Cell adhesion kinetics remained invariant when the average fiber diameter was in the micron range (1,647 nm), whereas cell-growth kinetics were slightly greater than with 900 nm scaffolds. We propose that the uniformness of fibers and the average fiber diameter may play an important role in modulating cellular attachment and proliferation in electrospun tissue engineering scaffolds.
Biomacromolecules, 2018
Electrospun ultrafine fibres prepared using a blend of poly(lactide-co-glycolide) (PLGA) and bromine terminated poly(ʟ-lactide) (PLA-Br), were surface modified using surface-initiated (SI) Cu(0) mediated polymerization. Copolymers based on N-acryloxysuccinimide (NAS) and a low fouling monomer (either N,N-dimethylacrylamide (DMA), N-(2-hydroxypropyl)acrylamide (HPA) or N-acryloylmorpholine (NAM)) were grafted from the fibre surface to impart surface functionality and to reduce non-specific protein adsorption. Inclusion of the functional NAS monomer facilitated the conjugation of a non-bioactive cyclic RAD peptide and a bioactive cyclic RGD peptide, the latter expected to facilitate cell adhesion through its affinity for the α v β 3 integrin receptor. A detailed analysis of the surface of the electrospun fibre scaffolds in non-grafted form compared to the surface functionalized state is presented. Characteristic amino acid peaks are observed for both conjugated RGD and RAD peptides. Cell culture experiments confirmed cell specific attachment mediated through the presence of the bioactive RGD peptide mainly at high surface density.
Biomaterials, 2008
Aligned electrospun scaffolds are a promising tool for engineering fibrous musculoskeletal tissues as they reproduce the mechanical anisotropy of these tissues and can direct ordered neo-tissue formation. However, these scaffolds suffer from a slow cellular infiltration rate, likely due in part to their dense fiber packing. We hypothesized that cell ingress could be expedited in scaffolds by increasing porosity, while at the same time preserving overall scaffold anisotropy. To test this hypothesis, poly(ε-caprolactone) (a slow-degrading polyester) and poly(ethylene oxide) (a watersoluble polymer) were co-electrospun from two separate spinnerets to form dual-polymer composite fiber-aligned scaffolds. Adjusting fabrication parameters produced aligned scaffolds with a full range of sacrificial (PEO) fiber contents. Tensile properties of scaffolds were a function of the ratio of PCL to PEO in the composite scaffolds, and were altered in a predictable fashion with removal of the PEO component. When seeded with mesenchymal stem cells (MSCs), increases in the starting sacrificial fraction (and porosity) improved cell infiltration and distribution after three weeks in culture. In pure PCL scaffolds, cells lined the scaffold periphery, while scaffolds containing >50% sacrificial PEO content had cells present throughout the scaffold. These findings indicate that cell infiltration can be expedited in dense fibrous assemblies with the removal of sacrificial fibers. This strategy may enhance in vitro and in vivo formation and maturation of a functional constructs for fibrous tissue engineering.
Cell adhesion on artificial materials for tissue engineering
Physiological research / Academia Scientiarum Bohemoslovaca, 2004
Advanced interdisciplinary scientific field of tissue engineering has been developed to meet increasing demand for safe, functional and easy available substitutes of irreversibly damaged tissues and organs. First biomaterials were constructed as "two-dimensional" (allowing cell adhesion only on their surface), and durable (non-biodegradable). In contrast, biomaterials of new generation are characterized by so-called three dimensional porous or scaffold-like architecture promoting attachment, growth and differentiation of cells inside the material, accompanied by its gradual removal and replacement with regenerated fully functional tissue. In order to control these processes, these materials are endowed with a defined spectrum of bioactive molecules, such as ligands for adhesion receptors on cells, functional parts of natural growth factors, hormones and enzymes or synthetic regulators of cell behavior, incorporated in defined concentrations and spatial distribution against...
Biological Performance of Electrospun Polymer Fibres
Materials, 2019
The evaluation of biological responses to polymeric scaffolds are important, given that the ideal scaffold should be biocompatible, biodegradable, promote cell adhesion and aid cell proliferation. The primary goal of this research was to measure the biological responses of cells against various polymeric and collagen electrospun scaffolds (polycaprolactone (PCL) and polylactic acid (PLA) polymers: PCL–drug, PCL–collagen–drug, PLA–drug and PLA–collagen–drug); cell proliferation was measured with a cell adhesion assay and cell viability using 5-bromo-2′-deoxyuridine (BrdU) and resazurin assays. The results demonstrated that there is a distinct lack of growth of cells against any irgasan (IRG) loaded scaffolds and far greater adhesion of cells against levofloxacin (LEVO) loaded scaffolds. Fourteen-day studies revealed a significant increase in cell growth after a 7-day period. The addition of collagen in the formulations did not promote greater cell adhesion. Cell viability studies rev...
A review of key challenges of electrospun scaffolds for tissue-engineering applications
Journal of Tissue Engineering and Regenerative Medicine, 2015
Tissue engineering holds great promise to develop functional constructs resembling the structural organization of native tissues to improve or replace biological functions, with the ultimate goal of avoiding organ transplantation. In tissue engineering, cells are often seeded into artificial structures capable of supporting three-dimensional (3D) tissue formation. An optimal scaffold for tissueengineering applications should mimic the mechanical and functional properties of the extracellular matrix (ECM) of those tissues to be regenerated. Amongst the various scaffolding techniques, electrospinning is an outstanding one which is capable of producing non-woven fibrous structures with dimensional constituents similar to those of ECM fibres. In recent years, electrospinning has gained widespread interest as a potential tissue-engineering scaffolding technique and has been discussed in detail in many studies. So why this review? Apart from their clear advantages and extensive use, electrospun scaffolds encounter some practical limitations, such as scarce cell infiltration and inadequate mechanical strength for load-bearing applications. A number of solutions have been offered by different research groups to overcome the above-mentioned limitations. In this review, we provide an overview of the limitations of electrospinning as a tissue-engineered scaffolding technique, with emphasis on possible resolutions of those issues. /term grounded points for fibre attachment and structural packing density. (d) Electrospinning in wet media; using a metal bath filled with liquid medium as a grounded collector induces large spaces between the forming fibres and lessens packing density. (e) Applying a postprocessing modification; laser irradiation over an electrospun scaffold can create a desirable pore structure Challenges regarding electrospun scaffolds: a review Figure 4. Schematic illustration indicating various approaches for improving cell infiltration into the electrospun scaffold. (a) Biological factors inclusion within an electrospun scaffold; association of biochemical cues, such as chemokine gradient, cell-friendly natural polymers, etc., with an electrospun scaffold encourages cell migration into the interior part. (b) Cell electrospraying and cell layering; direct incorporation of cells among fibres or fibrous layers creates a fully cell-laden electrospun structure. (c) Creating dynamic cell culture condition; agitation of the culture medium via dynamic culture enhances nutrient-waste exchange and drives cells to the interior parts. (d) Combining electrospinning with other scaffold fabrication methods; associating electrospun fibres with hydrogel creates a highly ECM-mimicking composite with the desirable cell infiltration Challenges regarding electrospun scaffolds: a review