Aligned nanofibres made of poly(3-hydroxybutyrate) grafted to hyaluronan for potential healthcare applications.

In this work, a hybrid copolymer consisting of poly(3-hydroxybutyrate) grafted to hyaluronic acid (HA) was synthesised and characterised. Once formed, the P(3HB)-g-HA copolymer was soluble in water allowing a green electrospinning process. The diameters of nanofibres can be tailored by simply varying the Mw of polymer. The optimization of the process allowed to produce fibres of average diameter in the range of 100-150 nm and low polydispersity. The hydrophobic modification has not only increased the fibre diameter, but also the obtained layers were homogenous. At the nanoscale, the hybrid copolymer exhibited an unusual hairy topography. Moreover, the hardness and tensile properties of the hybrid were found to be superior compared to fibres made of unmodified HA. Particularly, this reinforcement was achieved at the longitudinal direction. Additionally, this work reports the use in the composition of a water-soluble copolymer containing photo cross-linkable moieties to produce insoluble materials post-electrospinning. The derivatives as well as their nanofibrous mats retain the biocompatibility of the natural polymers used for the fabrication.

Water-insoluble fibres, threads, and fabrics from lauroyl derivatives of hyaluronan.

Hyaluronan (HA) is a naturally occurring polysaccharide widely used in medicine and cosmetics. To further broaden its potential, various HA derivatives have been developed with the aim of reducing solubility, slowing degradation, or providing other beneficial properties. However, for most medical applications, these derivatives must be processed into suitable forms. Here we present water-insoluble fibres prepared from lauroyl-modified HA using a wet spinning process. Important properties of the fibres, such as swelling or the degradation rate, can be fine-tuned by adjusting the degree of HA modification. Due to their mechanical properties, the lauroyl HA fibres can be easily processed into threads and subsequently into fabrics of various sizes, shapes, and degrees of porosity. In addition, in vitro cytotoxicity testing of the fibres showed that they were non-cytotoxic. Overall, our results suggest that lauroyl HA fibres are a promising material that could be used to develop a variety of medical devices.

Effects of emulsion, dispersion, and blend electrospinning on hyaluronic acid nanofibers with incorporated antiseptics.

Nanofibrous materials are used in drug delivery as carriers of active ingredients. These can be incorporated into the materials with various electrospinning methods that differ mainly in the way spinning solutions are prepared. Each method affects primarily the encapsulation efficiency and distribution of active ingredients in the materials. This study focuses on the incorporation of octenidine dihydrochloride (OCT) and triclosan (TRI) into nanofibrous materials electrospun from native hyaluronic acid emulsions, dispersions, and blends. OCT had no substantial effect on fiber morphology, which is affected by the solvent system. All OCT encapsulation efficiencies were comparable (approximately 90%). TRI encapsulation efficiencies varied greatly depending on the method used. Merely 3% of TRI was encapsulated when it was spun from a dispersion. Encapsulation efficiency was higher, and TRI was incorporated in clusters when an emulsion was used. The best result was achieved with a blend, in which case 96% of TRI was encapsulated.

Nanofibrous material from hyaluronan derivatives preserving fibrous structure in aqueous environment.

Nanofibrous materials produced from natural polymers have wide range of potential uses in regenerative medicine. This paper focuses on preparation of nanofibrous layers produced from intentionally hydrophobized derivatives of hyaluronan, which is known for its ability to promote wound healing. This structural modification of hyaluronan expands the range of potential uses of this promising material, which is otherwise limited due to the hydrophilic nature of hyaluronic acid. The aim of this research was preparation of nanofibrous material that would retain its fibrous structure and dimensional stability even after getting into contact with an aqueous medium, which is impossible to achieve with layers composed solely of native hyaluronan. As a result, such material would be able to retain its breathability and good mechanical properties when both dry and wet. Furthermore, all prepared materials were proved non-toxic for cells. This self-supporting nanofibrous matrix can be used as a scaffold, or porous wound dressing.

Antimicrobial nanofibrous mats with controllable drug release produced from hydrophobized hyaluronan.

Due to their large active surface, high loading efficiency, and tunable dissolution profiles, nanofibrous mats are often cited as promising drug carriers or antimicrobial membranes. Hyaluronic acid has outstanding biocompatibility, but it is hydrophilic. Nanofibrous structures made from hyaluronan dissolve immediately, making them unsuitable for controlled drug release and longer applications. We aimed to prepare a hyaluronan-based antimicrobial nanofibrous material, which would retain its integrity in aqueous environments. Self-supporting nanofibrous mats containing octenidine dihydrochloride or triclosan were produced by electrospinning from hydrophobized hyaluronan modified with a symmetric lauric acid anhydride. The nanofibrous mats required no cross-linking to be stable in PBS for 7 days. The encapsulation efficiency of antiseptics was nearly 100%. Minimal release of octenidine was observed, while up to 30% of triclosan was gradually released in 72 h. The nanofibrous materials exhibited antimicrobial activity, the fibroblast viability was directly dependent on the antiseptic content and its release.

Electrospinning of Fibrous Layers Containing an Antibacterial Chlorhexidine/Kaolinite Composite.

The aim of this study was to prepare self-supporting homogeneous nano/microfibrous layers with a content of the clay mineral kaolinite and kaolinite modified with the antibacterial agent chlorhexidine (CH). Fibers were made of hydrophobic polymers-polyurethane and polycaprolactone. Polymer suspensions for electrospinning contained 2, 5, and 8 wt % (relative to the total weight of the suspension) of kaolinite or CH/kaolinite and were electrospun using 4SPIN LAB. The morphology of prepared fibrous layers was characterized using scanning electron microscopy; energy-dispersive X-ray spectroscopy mapping and Raman spectroscopy were used to confirm the presence and distribution of kaolinite in the layers. Fiber diameters decreased after adding kaolinite or CH/kaolinite and ranged from 600 nm to 5 µm. Antibacterial CH was found in kaolinite itself as well as separately in the fibers (result of imperfect bonding of CH onto the surface of kaolinite). The encapsulation efficiency of all samples exceeded 64%, and the highest efficiency was observed in samples with 2 wt % CH/kaolinite. Samples containing CH exhibited good antibacterial activity against Staphylococcus aureus, and the effectiveness of which was affected by the concentration of the antibacterial agent. The release of CH was very slow, and there was no initial burst release. Overall, no more than 5% of the CH was released over a course of 168 h. The Korsmeyer-Peppas model revealed that CH is released by a diffusion mechanism.

Effects of wound dressings containing silver on skin and immune cells.

Wound dressings with silver have been shown to be cytotoxic in vitro. However, the extrapolation of this cytotoxicity to clinical settings is unclear. We applied dressings with various forms of silver on porcine skin ex vivo and investigated silver penetration and DNA damage. We assessed antimicrobial efficacy, cytotoxicity to skin cells, and immune response induced by the dressings. All dressings elevated the DNA damage marker γ-H2AX and the expression of stress-related genes in explanted skin relative to control. This corresponded with the amount of silver in the skin. The dressings reduced viability, induced oxidative stress and DNA damage in skin cells, and induced the production of pro-inflammatory IL-6 by monocytes. The oxidative burst and viability of activated neutrophils decreased. The amount of silver released into the culture medium varied among the dressings and correlated with in vitro toxicity. However, antimicrobial efficiencies did not correlate strongly with the amount of silver released from the dressings. Antimicrobial efficiency and toxicity are driven by the form of silver and the construction of dressings and not only by the silver concentration. The damaging effects of silver dressings in ex vivo skin highlight the importance of thorough in vivo investigation of silver dressing toxicity.

Synthesis and Physicochemical Characterization of Undecylenic Acid Grafted to Hyaluronan for Encapsulation of Antioxidants and Chemical Crosslinking.

In this work, a new amphiphilic derivative made of 10-undecylenic acid grafted to hyaluronan was prepared by mixed anhydrides. The reaction conditions were optimized, and the effect of the molecular weight (Mw), reaction time, and the molar ratio of reagents was explored. Using this methodology, a degree of substitution up to 50% can be obtained. The viscosity of the conjugate can be controlled by varying the substitution degree. The physicochemical characterization of the modified hyaluronan was performed by infrared spectroscopy, Nuclear Magnetic Resonance, Size-Exclusion Chromatography combined with Multiangle Laser Light Scattering (SEC-MALLS), and rheology. The low proton motility and self-aggregation of the amphiphilic conjugate produced overestimation of the degree of substitution. Thus, a novel method using proton NMR was developed. Encapsulation of model hydrophobic guest molecules, coenzyme Q10, curcumin, and α-tocopherol into the micellar core was also investigated by solvent evaporation. HA-UDA amphiphiles were also shown to self-assemble into spherical nanostructures (about 300 nm) in water as established by dynamic light scattering. Furthermore, HA-UDA was crosslinked via radical polymerization mediated by ammonium persulphate (APS/TEMED). The cross-linking was also tested by photo-polymerization catalyzed by Irgacure 2959. The presence of the hydrophobic moiety decreases the swelling degree of the prepared hydrogels compared to methacrylated-HA. Here, we report a novel hybrid hyaluronan (HA) hydrogel system of physically encapsulated active compounds and chemical crosslinking for potential applications in drug delivery.

A novel photopolymerizable derivative of hyaluronan for designed hydrogel formation.

A new photopolymerizable derivative of hyaluronan (methacrylhydrazide-HA, MAHA) was prepared by carbodiimide chemistry. The reaction conditions were optimized for molecular weight (Mw), reaction time and amount of reagents with a degree of methacrylation (DM) ranging from 2% to 58%. Methacrylhydrazide-HA was hydrolytically stable (PBS, 7days, 37°C) in contrast to commonly used methacrylester analoque (23% hydrolyzed). MAHA readily photopolymerized into densely crosslinked hydrogels under physiological conditions. The varied DM, Mw, irradiation time (texp) and macromer concentration in photocrosslinking afforded hydrogels with different physical (swelling ratio, degradation rate) and mechanical properties (stiffness, toughness). Three-dimensional fabrication and surface patterning of MAHA hydrogels were demonstrated by photolithography and light mediated micromolding. A live-dead assay with skin fibroblasts showed convenient biocompatibility of MAHA (16%, 116kDa) for potential scaffolding applications in tissue engineering and regenerative medicine.

One-pot synthesis of α,ß-unsaturated polyaldehyde of chondroitin sulfate.

Chondroitin sulfate (CS) was chemoselectively oxidized by Tempo/NaClO to C-6 aldehyde of a D-galactosamine subunit (GalNAc). The subsequent, spontaneous desulfatation of oxidized CS gave rise to α,ß-unsaturated aldehyde. A new derivative of CS was fully characterized and a degree of oxidation was determined by spectroscopic analysis. The optimization of reaction conditions showed a proportional degree of oxidation to an amount of sodium hypochlorite. The utility of α,ß-unsaturated aldehyde for crosslinking and conjugation was demonstrated by a seamless condensation with various N-nucleophiles. We also demonstrated pH-dependent release of biologically active agents from oxidized CS. A live-dead assay in the presence of α,ß-unsaturated aldehyde revealed unaffected viability of NIH 3T3 fibroblasts, which made this precursor promising for several biomedical applications including drug delivery and tissue engineering.

Solid-state photocrosslinking of hyaluronan microfibres.

Hyaluronan (HA) was chemically modified to a photocurable derivative by the acylation with a mixed anhydride of trans-cinnamic acid and characterized by UV, IR and 2D NMR spectroscopy. Photocurable HA was processed to a microfibrous structure by wet-spinning technology. Water solubility of otherwise water-soluble HA microfibres was reduced significantly by the solid-state photocrosslinking. We also investigated that the nature of polymer structure had a great impact to a final crosslink ratio. The analysis of the mechanical properties showed higher ultimate tensile strength and increased rigidity of the photocrosslinked fibres in comparison to the untreated ones. Moreover all tested materials are regarded as biocompatible according to the tests of cell viability, phototoxicity and enzymatic degradability, which make them suitable candidates for numerous biomedical applications.