Elisabete D. Pinho

Surface modification of electrospun polycaprolactone nanofiber meshes by plasma treatment to enhance biological performance

A critical aspect in the development of biomaterials is the optimization of their surface properties to achieve an adequate cell response. In the present work, electrospun polycaprolactone nanofiber meshes (NFMs) are treated by radio-frequency (RF) plasma using different gases (Ar or O(2)), power (20 or 30 W), and exposure time (5 or 10 min). Morphological and roughness analysis show topographical changes on the plasma-treated NFMs. X-ray photoelectron spectroscopy (XPS) results indicate an increment of the oxygen-containing groups, mainly --OH and --C==O, at the plasma-treated surfaces. Accordingly, the glycerol contact angle results demonstrate a decrease in the hydrophobicity of plasma-treated meshes, particularly in the O(2)-treated ones. Three model cell lines (fibroblasts, chondrocytes, and osteoblasts) are used to study the effect of plasma treatments over the morphology, cell adhesion, and proliferation. A plasma treatment with O(2) and one with Ar are found to be the most successful for all the studied cell types. The influence of hydrophilicity and roughness of those NFMs on their biological performance is discussed. Despite the often claimed morphological similarity of NFMs to natural extracellular matrixes, their surface properties contribute substantially to the cellular performance and therefore those should be optimized.

Melt-based compression-molded scaffolds from chitosan-polyester blends and composites: Morphology and mechanical properties

Blends of chitosan and synthetic aliphatic polyesters (polybutylene succinate, polybutylene succinate adipate, polycaprolactone, and polybutylene terepthalate adipate) were compounded with and without hydroxyapatite, a bioactive mineral filler known to enhance osteoconduction. The blends and composites were compression molded with two different granulometric salt sizes (63-125 microm and 250-500 microm) having different levels of salt content (60, 70, and 80%) by weight. By leaching the salt particles, it was possible to produce porous scaffolds with distinct morphologies. The relationship between scaffold morphology and mechanical properties was evaluated using scanning electron microscopy, microcomputed tomography, compression testing, differential scanning calorimetry, small-angle X-ray scattering (SAXS), and wide-angle X-ray scattering. The produced scaffolds are characterized by having different morphologies depending on the average particle size and the amount of NaCl used. Specimens with higher porosity level have a less organized pore structure but increased interconnectivity of the pores. The stress-strain curve under compression displayed a linear elasticity followed by a plateau whose characteristics depend on the scaffold polymer composition. A decrease in the salt particle size used to create the porosity caused in general a decrease in the mechanical properties of the foams. Composites with hydroxyapatite had a sharp reduction in yield stress, modulus, and strain at break. The melting temperature decreased with increased chitosan content. SAXS results indicate no preferential crystalline orientation in the scaffolds. Cytotoxicity evaluation were carried out using standard tests (accordingly to ISO/EN 10993 part 5 guidelines), namely MTS test with a 24-h extraction period, revealing that L929 cells had comparable metabolic activities to that obtained for the negative control.

Surface controlled biomimetic coating of polycaprolactone nanofiber meshes to be used as bone extracellular matrix analogues

The aim of this work was to develop novel electrospun nanofiber meshes coated with a biomimetic calcium phosphate (BCP) layer that mimics the extracellular microenvironment found in the human bone structure. Poly (ε-caprolactone) (PCL) was selected because of its well-known medical applications, its biodegradability, biocompatibility and its susceptibility to partial hydrolysis by a straightforward alkaline treatment. The deposition of a calcium phosphate layer, similar to the inorganic phase of bone, on PCL nanofiber meshes was achieved by means of a surface modification. This initial surface modification was followed by treatment with solutions containing calcium and phosphate ions. The process was finished by a posterior immersion in a simulated body fluid (SBF) with nearly 1.5 × the inorganic concentration of the human blood plasma ions. After some optimization work, the best conditions were chosen to perform the biological assays. The influence of the bone-like BCP layer on the viability and adhesion, as well as on the proliferation of human osteoblast-like cells, was assessed. It was shown that PCL nanofiber meshes coated with a BCP layer support and enhance the proliferation of osteoblasts for long culture periods. The attractive properties of the coated structures produced in the present work demonstrated that those materials have potential to be used for applications in bone tissue engineering. This is the first time that nanofiber meshes could be coated with a biomimetic bone-like calcium phosphate layer produced in a way that the original mesh architecture can be fully maintained.

Solving cell infiltration limitations of electrospun nanofiber meshes for tissue engineering applications

AIM Utilize the dual composition strategy to increase the pore size and solve the low cell infiltration capacity on random nanofiber meshes, an intrinsic limitation of electrospun scaffolds for tissue engineering applications. MATERIALS & METHODS Polycaprolactone and poly(ethylene oxide) solutions were electrospun simultaneously to obtain a dual composition nanofiber mesh. Selective dissolution of the poly(ethylene oxide) nanofiber fraction was performed. The biologic performance of these enhanced pore size nanofibrous structures was assessed with human osteoblastic cells. RESULTS The electrospun nanofiber meshes, after the poly(ethylene oxide) dissolution, showed statistically significant larger pore sizes when compared with polycaprolactone nanofiber meshes with a similar polycaprolactone volume fraction. This was also confirmed by interferometric optical profilometry. Using scanning electron microscopy and laser scanning confocal microscopy, it was observed that osteoblastic cells could penetrate into the nanofibrous structure and migrate into the opposite and unseeded side of the mesh. CONCLUSION An electrospun mesh was created with sufficient pore size to allow cell infiltration into its structure, thus resulting in a fully populated construct appropriate for 3D tissue engineering applications.

Processing ulvan into 2D structures: Cross-linked ulvan membranes as new biomaterials for drug delivery applications

The polysaccharide ulvan, composed of sulphated rhamnose, glucoronic and iduronic acids was used to produce polymeric membranes by solvent casting. As ulvan is soluble in water, a cross-linking step was necessary to render the membrane insoluble in water and stable at physiological conditions. Cross-linked ulvan membranes were characterized by FTIR, SEM, swelling behaviour was investigated and the mechanical performance assessed by quasi-static tensile testing. Furthermore, the ability and mechanism of sustained release of a model drug from ulvan membranes was investigated. Produced membranes revealed remarkable ability to uptake water (up to ∼1800% of its initial dry weight) and increased mechanical performance (1.76 MPa) related with cross-linking. On the other hand, medicated ulvan dressings demonstrate the potential as drug delivery devices. Using a model drug we have observed an initial steady release of the drug - of nearly 49% - followed by slower and sustained release up to 14 days. The properties of ulvan membranes herein revealed suggest a great potential of this natural sulphated polysaccharide as a wound dressing.

Degradable particulate composite reinforced with nanofibres for biomedical applications

Nanofibre-based structures and their composites are increasingly being studied for many biomedical applications, including tissue engineering scaffolds. These materials enable architectures resembling the extracellular matrix to be obtained. The search for optimized supports and carriers of cells is still a major challenge for the tissue engineering field. The main purpose of this work is to develop a novel composite structure that combines microparticles and nanofibres in reinforced polymeric microfibres. This innovative combination of materials is obtained by melting extrusion of a particulate composite reinforced with chitosan nanofibre meshes (0.05 wt.%) produced by the electrospinning technique. The reinforced microfibres were analysed by scanning electron microscopy and showed a considerable alignment of the chitosan nanofibres along the longitudinal main axis of the microfibre composite structure. The tensile mechanical properties revealed that the introduction of the nanofibre reinforcement in the particulate microfibre composite increased the tensile modulus by up to 70%. The various structures were subjected to swelling and degradation tests immersed in an isotonic saline solution at 37 degrees C. The presence of chitosan nanofibres in the particulate microfibres enhances the water uptake by up to 24%. The combination of good mechanical properties and enhanced degradability of the developed structures is believed to have great potential for various biomedical applications, including three-dimensional fibre mesh scaffolds to be applied in the field of bone tissue engineering.

Performance of biodegradable microcapsules of poly(butylene succinate), poly(butylene succinate-co-adipate) and poly(butylene terephthalate-co-adipate) as drug encapsulation systems.

Poly(butylene succinate) (PBSu), poly(butylene succinate-co-adipate) (PBSA) and poly(butylene terephthalate-co-adipate) (PBTA) microcapsules were prepared by the double emulsion/solvent evaporation method. The effect of polymer and poly(vinyl alcohol) (PVA) concentration on the microcapsule morphologies, drug encapsulation efficiency (EE) and drug loading (DL) of bovine serum albumin (BSA) and all-trans retinoic acid (atRA) were all investigated. As a result, the sizes of PBSu, PBSA and PBTA microcapsules were increased significantly by varying polymer concentrations from 6 to 9%. atRA was encapsulated into the microcapsules with an high level of approximately 95% EE. The highest EE and DL of BSA were observed at 1% polymer concentration in values of 60 and 37%, respectively. 4% PVA was found as the optimum concentration and resulted in 75% EE and 14% DL of BSA. The BSA release from the capsules of PBSA was the longest, with 10% release in the first day and a steady release of 17% until the end of day 28. The release of atRA from PBSu microcapsules showed a zero-order profile for 2 weeks, keeping a steady release rate during 4 weeks with a 9% cumulative release. Similarly, the PBSA microcapsules showed a prolonged and a steady release of atRA during 6 weeks with 12% release. In the case of PBTA microcapsules, after a burst release of 10% in the first day, showed a parabolic release profile of atRA during 42 days, releasing 36% of atRA.

Surface modification of a biodegradable composite by UV laser ablation: in vitro biological performance

Melt blends of chitosan and biodegradable aliphatic polyester have been physically and biologically studied, presenting great potential for biomedical applications. Structurally, poly(butylene succinate)–chitosan (PBS/Cht) composite scaffolds are covered by a thin PBS layer, preventing the desired interaction of cells/tissues with the chitosan particules. In the present work, a selective and controlled ablation of this skin layer was induced by UV laser processing. X‐ray photoelectron spectroscopy (XPS) and time‐of‐flight secondary ion mass spectrometry (ToF–SIMS) data demonstrated an increment of chitosan components and others resulting from the laser ablation process. The biological activity (i.e. cell viability and proliferation) on the inner regions of the composite scaffolds is not significantly different from those of the external layer, despite the observed differences in surface roughness (determined by interferometric optical profilometry) and wettability (water contact angle). However, the morphology of human osteoblastic cells was found to be considerably different in the case of laser‐processed samples, since the cells tend to aggregate in multilayer columnar structures, preferring the PBS surface and avoiding the chitosan‐rich areas. Thus, UV laser ablation can be considered a model technique for the physical surface modification of biomaterials without detrimental effects on cellular activity. Copyright