Novel bioactive scaffolds with fibronectin recognition nanosites based on molecular imprinting technology

Biomimetic materials for application in the field of tissue engineering are usually obtained through covalent bonding between the polymer backbone and the bioactive molecules. A totally new approach, proposed for the first time by our research group, for the creation of advanced synthetic support structures for cell adhesion and proliferation is represented by molecular imprinting (MI) technology. In this article, we describe the synthesis and characterization of molecularly imprinted polymers with recognition properties toward a fibronectin peptide sequence and their application as functionalization structures. Polymers, in the form of densely fused microgel particles, were obtained by precipitation polymerization. The imprinted particles showed good performance in terms of recognition capacity and quantitative rebinding; moreover, the epitope effect was observed, with the particles able to recognize and rebind not only the specific peptide sequence but also a larger fibronectin fragment. The cytotoxicity tests showed normal vitality in C2C12 myoblasts cultured in a medium that was put in contact with the imprinted particles. Therefore, imprinted particles were used to functionalize synthetic polymeric films by deposition on their surface. The deposition of the imprinted particles did not alter their specific recognition and rebinding behavior. The most remarkable result was obtained by the biological characterization: in fact, the functionalized materials appeared able to promote cell adhesion and proliferation. These results are very promising and suggest that MI can be used as an innovative functionalization technique to prepare bioactive scaffolds with an effective capacity for improving tissue regeneration. © 2010 Wiley Periodicals, Inc. J Appl Polym Sci, 2010     

Characterisation of blends between poly(ε-caprolactone) and polysaccharides for tissue engineering applications

In this work, bioartificial binary blends between poly(ε-caprolactone) (PCL) and a polysaccharide (chitosan (CS) or starch (S)) with different contents of the natural polymer (5–30 wt.%) were produced. Melt-mixing and double-precipitation were the methods used for the obtainment of PCL/S and PCL/CS blends, respectively. Tubular scaffolds were produced from bioartificial blends by melt-extrusion. Physico-chemical characterisation was performed by differential scanning calorimetry analysis (DSC), thermogravimetry (TGA), scanning electron microscopy (SEM), infrared analysis (FTIR-ATR and micro-ATR mapping), atomic force microscopy (AFM) and stress–strain tests. Blends were not miscible, phase-separated systems, showing a homogeneous composition and morphology only at low polysaccharide content (≤ 10 wt.%). The biocompatibility of bioartificial guides was investigated by culturing NIH-3T3 mouse fibroblasts. Cells response showed the following order: PCL/S > PCL > PCL/CS. For each blend type, biocompatibility increased with decreasing the polysaccharide content. In vitro cell tests using S5Y5 neuroblastoma cells, carried out on the most biocompatible blends, assessed their absence of cytotoxicity towards these model cells of the nervous tissue. Results showed that blends with a low chitosan or starch content (≤ 10 wt.%) are promising for the regeneration of tissues requiring tubular scaffolds, such as the peripheral nerves.     

Grafting of polypropylene and its potential use as metal ion adsorption resin

Polypropylene (PP) was modified by radical grafting of acrylic acid (AA) in the melt by using dicumyl peroxide (DCP) as initiator. To reach a high grafting degree (GD) without substantially modifying PP‐chain structure, a few runs were carried out by employing butyl 3‐(2‐furanyl)propenoate (BFA), which is a coagent able to preserve the molecular weight (MW) of PP macromolecules in the presence of radical grafting reagents. All the samples were extracted with selective solvents to remove unreacted chemicals and free poly(acrylic acid) (PAA), and the GD was accurately determined by using the FTIR methodology. The GD ranged from 1.51 to 4.67 mol %. High‐temperature size permeation chromatography analysis was used to evaluate MW behavior, confirming the control exerted by the presence of BFA on the degradation reactions. DSC, TGA, and SEM analyses were performed to analyze the chemical modification effects on the polymer products' thermal and morphological properties. Finally, selected samples, with a particle size distribution in the range 100–850 μm, were tested as metal ion adsorption resins. Al(III), Cr(III), Zn(II), Cd(II), Pb(II), and Hg(II) were studied, and the highest adsorption efficiency values (in percentage) were obtained for Al(III) (15–20%) and Hg(II) (25–30%). © 2009 Wiley Periodicals, Inc. J Appl Polym Sci, 2009     

Synthesis and structure–property relationship of polyester-urethanes and their evaluation for the regeneration of contractile tissues

The structure–property relationship of degradable polyurethanes from non toxic building blocks was studied by synthesising four different biodegradable poly(ester urethanes) from poly(ε-caprolactone) (PCL) diol, 1,4-diisocyanatobutane and different chain extenders. For instance, the chain extenders were an amino acid derivative diamine, an amino acid derivative diol, a cyclic diol and a custom made diamine, containing an enzymatically degradable peptide (Ala–Ala sequence). Physicochemical and morphological characterisation (SEC, DSC, DMA, AFM) was performed, showing the influence of the chain extender on the polyurethane properties. A correlation between surface domain morphologies and thermal properties was highlighted and a relationship between the biological response and surface morphologies was observed. Collecting mechanical characterisation and myoblast cell culture results together, the polyurethane synthesised with the amino acid derivative diamine resulted the most promising candidate for fabricating scaffolds supporting the regeneration of muscle tissues.     

Radical functionalization of poly(butylene succinate-co-adipate): Effect of cinnamic co-agents on maleic anhydride grafting

Maleic anhydride (MAH), trans cinnamic acid (AcCin) and ethyl cinnamate (EtCin) were radically graft onto poly(butylene succinate-co-adipate) (PBSA). Samples were prepared in Brabender at 175 °C by addition of increasing amounts of MAH, AcCin, EtCin and their combinations, i.e. MAH/AcCin and MAH/EtCin, setting DCP content in the 0.2-0.6 wt% range. Monomer grafting was quantitatively determined by FT-IR. MAH grafting degrees (FD_M) resulted up to 1 mol%. Conversely, AcCin grafting degrees (FD_A) were found almost negligible in all cases. EtCin was found grafted in the 0.3-1.0 mol% range when the binary system MAH/EtCin was applied. Same MAH feeds returned almost doubled MAH grafting degrees (FD_M) when AcCin was the stoichiometric co-agent, and even three times higher FD_M when EtCin was the stoichiometric co-agent. On accounts of all the collected results, a kinetic model of the investigated systems is proposed.     

Defined Chitosan-based networks by C-6-Azide–alkyne “click” reaction

Chitosan was converted to its –NH₂-protected derivative by reacting with phthalic anhydride and then selectively functionalized with an azido group in C-6 position. Two different procedures were employed. The first was a “one-pot” procedure and the second was a two steps reaction through the tosylated intermediate. Both methods resulted in an effective functionalization of chitosan with azido groups. For the two before mentioned procedures, the functionalization degree was estimated by elemental analysis as number of azido groups per pyranoside ring and was 0.28 mol/mol and 0.26 mol/mol, respectively. The azido-functionalized derivatives underwent further modification by Cu(I) catalyzed dipolar cycloaddition with mono- or di-alkynes. The reaction with phenylacetylene produced soluble derivatives that were fully characterized at molecular level by FT-IR and NMR spectroscopies, elemental analysis and size exclusion chromatography (SEC). Crosslinked derivatives were obtained by reactions with 1,7-octadiyne or 1,4-diethynylbenzene and subsequently deprotected to restore the free amino groups. These last systems showed selective swellability in acid medium.     

Innovative tissue engineering structures through advanced manufacturing technologies

Awide range of rapid prototyping (RP) techniques for the construction of three-dimensional (3-D) scaffolds for tissue engineering has been recently developed. In this study, we report and compare two methods for the fabrication of poly-(epsilon-caprolactone) and poly-(epsilon-caprolactone)-poly-(oxyethylene)-poly-(epsilon-caprolactone) copolymer scaffolds. The first technique is based on the use of a microsyringe and a computer-controlled three-axis micropositioner, which regulates motor speed and position. Polymer solutions are extruded through the needle of the microsyringe by the application of a constant pressure of 10-300 mm Hg, resulting in controlled polymer deposition of 5-600 microm lateral dimensions. The second method utilises the heating energy of a laser beam to sinter polymer microparticles according to computer-guided geometries. Materials may be fed either as dry powder or slurry of microparticles. Both powder granulometry and laser working parameters influence resolution (generally 300 microm x 700 microm), accuracy of sintering and surface and bulk properties of the final structures. The two RP methods allow the fabrication of 3-D scaffolds with a controlled architecture, providing a powerful means to study cell response to an environment similar to that found     

Reactive Blending of Polyamides with Different Carbonyl Containing Olefin Polymers

In this paper the blending of polyamides nylon 6 and nylon 12, with a perfectly alternating ethylene/CO copolymer containing 50 mol‐% carbonyl groups (polyketone) is investigated in comparison to blends of the same polyamides with polyolefins containing varying degrees of carbonyl group incorporation. These include a poly[ethylene‐co‐(methyl acrylate)] copolymer containing 1.9 mol‐% methyl ester groups and poly[ethylene‐co‐(ethyl undecylenate)] copolymers with between 0.20 and 1.25 mol‐% ester incorporation. Blends were obtained of polyamides and the polyolefins in compositions between 20/80 and 80/20 in solution and in a Brabender mixer. SEM studies together with TGA, DSC and FTIR measurements show excellent compatibilization for both polyketone and poly[ethylene‐co‐(methyl acrylate)] copolymers with the nylons. The poly[ethylene‐co‐(ethyl undecylenate)] polymers displayed much less compatibilization although they still performed significantly better compared to pure polyethylene. The difference in compatibilization is discussed with respect to the importance of both the number of interactive groups present in the polyolefin and the steric requirements of hydrogen bond formation.

SEM micrograph of the fracture surface of the blend nylon 6/polyethylene 70:30.     

Genipin-crosslinked chitosan/gelatin blends for biomedical applications

Blends between chitosan (CS) and gelatin (G) with various compositions (CS/G 0/100 20/80, 40/60, 60/40 100/0 w/w) were produced, as candidate materials for biomedical applications. Different amounts of genipin (0.5 wt.% and 2.5 wt.%) were used to crosslink CS/G blends, promoting the formation of amide and tertiary amine bonds between the macromolecules and the crosslinker. The effects of composition and crosslinking on the physico-chemical properties of samples were evaluated by infrared analysis, thermogravimetry, contact angle measurements, dissolution and swelling tests. Mechanical properties of crosslinked samples were also determined through stress–strain and creep tests: samples stiffness increased with increasing the crosslinker amount and the CS content. Blend composition affected mouse fibroblasts adhesion and proliferation on substrates, depending on the crosslinker amount. Finally, crosslinked CS/G blends containing 80 wt.% G were found to support neuroblastoma cells adhesion and proliferation which made them promising candidates for uses in the field of nerve regeneration.