Gianluca Ciardelli

Bioartificial polymeric materials based on polysaccharides

Bioartificial polymeric materials, based on blends of polysaccharides with synthetic polymers such as poly(vinyl alcohol) (PVA) and poly(acrylic acid) (PAA), were prepared as films or hydrogels. The physico-chemical, mechanical, and biological properties of these materials were investigated by different techniques such as differential scanning calorimetry, dynamic mechanical thermal analysis, scanning electron microscopy, and in vitro release tests, with the aim of evaluating the miscibility of the polymer blends and to establish their potential applications. The results indicate that while dextran is perfectly miscible with PAA, dextran/PVA, chitosan/PVA, starch/PVA, and gellan/PVAblends behave mainly as two-phase systems, although interactions can occur between the components. Cross-linked starch/PVAfilms could be employed as dialysis membranes: they showed transport properties comparable to, and in some cases better than, those of currently used commercial membranes. Hydrogels based on dextran/PVA and chitosan/PVA blends could find applications as delivery systems. They appeared able to release physiological amounts of human growth hormone, offering the possibility to modulate the release of the drug by varying the content of the biological component.

Enzymatically crosslinked porous composite matrices for bone tissue regeneration

Three-dimensional porous hydroxyapatite/collagen (HA/Coll) composites with a random pore structure were obtained by freeze-drying and crosslinked by an enzymatic treatment using microbial transglutaminase (mTGase). The procedure resulted in improved mechanical strength and thermal stability of the scaffolds. The scaffolds were characterized in terms of their stability (Coll release, swelling, collagenase-mediated degradation), thermal properties (thermogravimetric analysis, differential scanning calorimetry), mechanical behavior under compression and cell compatibility. Enzymatic treatment stabilized the sponges to water vapors, with measurable swelling ratio between 100% for HA/Coll/mTGase 0/100 to 5% for HA/Coll/mTGase 80/20. Weight loss in water due to Coll release was between 2 and 10% in mTGase-crosslinked samples and decreased with increasing HA content. Cultures of MG63 osteoblast-like cells and human umbilical vein endothelial cells (HUVEC) showed good adhesion and proliferation on the scaffolds, good viability (through MTT test, 100-150% of control), and good differentiation (alkaline phosphatase, up to 40 UI/L with respect to 35 UI/L for control).

Kinetics and reaction mechanism of template polymerization investigated by conductimetric measurements. Part 3. Radical polymerization of acrylic acid in the presence of poly(N-vinylpyrrolidone)

Conductimetry was used for monitoring the radical polymerization of a weak electrolyte (acrylic acid) in aqueous solution and for determination of the kinetic parameters of the reaction (reaction order with respect to monomer, activation energy). The results obtained were consistent with those determined by other techniques (such as dilatometry) or expected from theory. Dilatometric and conductimetric measurements were also used to study the template polymerization of acrylic acid onto poly(N-vinylpyrrolidone). Results indicate that the reaction proceeds according to a pick-up mechanism. Complexes between poly(acrylic acid) and poly(N-vinylpyrrolidone) were always isolated in equimolar composition of the two polymers, regardless of the polymerization mixture composition. Spectroscopic evidence of the existence of strong interaction and intimate mixing of the two polymers in the complexes was found. An influence of the template molecular weight on the chain length of the forming poly(acrylic acid) was detected by means of viscometry. © 2000 Society of Chemical Industry

Poly(ester urethane) Guides for Peripheral Nerve Regeneration.

A biocompatible and elastomeric PU was synthesized from low-molecular-weight PCL as macrodiol, CMD as chain extender and HDI as chain linker for applications in the field of peripheral nerve repair. PU cast films supported in vitro attachment and proliferation of NOBEC. The in vitro adhesion and proliferation of S5Y5 neuroblastoma cells on the inner surface of uncoated, gelatin- and PL-coated PU guides were compared. Due to their superior in vitro performance, PL-coated PU guides were tested in vivo for the repair of 1.8 cm-long defects in rat sciatic nerves. The progressive regeneration was confirmed by EMG and histological analysis showing the presence of regenerating fibers in the distal stumps.

Molecularly imprinted poly(ethylene-co-vinyl alcohol) membranes for the specific recognition of phospholipids.

In this paper we concentrated on the possibility of adopting molecular imprinting technology for the preparation of polymeric membranes imprinted with phosphatidylcholine, one of the main phospholipids found in the cell membrane and lipoproteins, via phase inversion, with the intention of applying this method in the ongoing research into the regression of atherosclerosis. The polymer matrix was based on poly(ethylene-co-vinyl alcohol) with an ethylene molar content of 44% and the amount of template molecule was varied so as to obtain three different kinds of membrane. We found that they possessed elevated binding capabilities (78.6% of the initial amount of phosphatidylcholine was found to be adsorbed by the membrane) united with a very high selectivity. Similar phospholipids (phosphatidylinositol and phosphatidylethanolamine) were found to be adsorbed only in very small quantities and mostly due to the porosity of the membrane itself and not to molecular imprinting.

Molecularly imprinted nanoparticles with recognition properties towards a laminin H–Tyr–Ile–Gly–Ser–Arg–OH sequence for tissue engineering applications

Nanotechnology is an emerging field that promises to revolutionize medicine and is increasingly used in tissue engineering applications. Our research group proposed for the first time molecular imprinting as a new nanotechnology for the creation of advanced synthetic support structures for cell adhesion and proliferation. The aim of this work was the synthesis and characterization of molecularly imprinted polymers with recognition properties towards a laminin peptide sequence and their application as functionalization structures in the development of bioactive materials. Nanoparticles with an average diameter of 200 nm were synthesized by precipitation polymerization of methacrylic acid in the presence of the template molecule and trimethylpropane trimethacrylate as the cross-linking agent. The imprinted nanoparticles showed good performance in terms of recognition capacity and selectivity. The cytotoxicity tests showed normal vitality of C2C12 myoblasts cultured in the medium that was put in contact with the imprinted polymers. After the deposition on the polymeric film surface, the imprinted particles maintained their specific recognition and rebinding behaviour, showing an even higher quantitative binding than free nanoparticles. Preliminary in vitro cell culture tests demonstrated the ability of functionalized materials to promote cell adhesion, proliferation and differentiation, suggesting that molecular imprinting can be used as an innovative functionalization technique.

Poly(ethylene‐co‐vinyl alcohol) Membranes with Specific Adsorption Properties for Potential Clinical Application

Abstract The preparation of novel polymeric systems through Molecular Imprinting Technology (MIT) for potential application in extracorporeal blood purification is described. Membranes based on poly(ethylene‐co‐vinyl alcohol) material, produced through a phase inversion method, were modified introducing in their structure specific binding sites for lipid and/or protein molecules. Membranes prepared are intended to selectively remove low density lipoproteins and cholesterol (LDL) from the plasma, by using interactions at a molecular level, between the molecularly imprinted membrane and specific target molecules, created during the preparation procedure. The binding performances of membranes and their potentiality as adsorbents for two different model target compounds, a phospholipid (phosphotidylcholine, PC) and a protein (α‐amylase enzyme) were investigated, showing improved adsorption capacity with respect to unmodified control membranes. In addition, molecularly imprinted poly(ethylene‐co‐vinyl alcohol) materials in the shape of microparticles, using the same templates, were prepared and studied for their potential use as adsorbers into a column.

Bioartificial polymeric materials: α-amylase, poly(2-hydroxyethyl methacrylate), poly(N-vinylpyrrolidone) system†

The preparation of a new bioartificial material based on poly(2-hydroxyethyl methacrylate) and the enzyme α-amylase by means of template polymerisation is described. The kinetics of the polymerisation reaction was monitored by dilatometry and a template influence on the initial rate evidenced. The morphological analysis indicated that the presence of the enzyme induced a porous structure of the obtained material, in which improved interactions between the two components could be detected with respect to its analogous prepared by simple blending of the preformed polymers. The negligible release of the active component (α-amylase) from the polycomplex suggested its possible application as an enzyme immobilisation system, since the enzyme showed to remain active in the polycomplex as well. In order to improve the properties of the system poly(N- vinylpyrrolidone) was added as an additional component in the polymerisation of 2-hydroxyethyl methacrylate in the presence of α-amylase. Higher biological activity of the immobilised enzyme was obtained for polycomplexes containing PVP, confirming the expected stabilising effect.

Molecularly imprinted bioartificial membranes for the selective recognition of biological molecules. Part 2: release of components and thermal analysis

Molecularly imprinted membranes imprinted for a large-molecular-weight protein were realised using a blend of natural and synthetic polymers. Bioartificial membranes of synthetic (poly(ethylene-co-vinyl alcohol)–EVAL, Clarene®) and biological (Dextran) polymers, molecularly imprinted with α-amylase as the template, were prepared and investigated. Dimethyl sulfoxide (DMSO) solutions of the α-amylase template, Clarene and Dextran were mixed under stirring in the desired proportions and dipped in DMSO (solvent)/water (non solvent) mixture, to obtain the phase separation. The release of Clarene, Dextran and α-amylase in the inversion baths was quantified by spectrophotometric methods and final composition of membranes was established. To study the interactions between the polymer components and between polymeric materials and the template, differential scanning calorimetry (DSC) and thermogravimetric analysis (TGA) were carried out. Results indicated that stable and continuous bioartificial membranes of Clarene and Dextran can be obtained, whereby calorimetric analysis suggested the presence of high interaction between α-amylase and the Clarene component.

Kinetics and reaction mechanism of template polymerization investigated by conductimetric measurements. 2. Radical polymerization of sodium methacrylate in the presence of poly(allylamine) hydrochloride

The polymerization of sodium methacrylate in water was investigated using conductimetry and dilatometry. Conductimetry was shown to be a reliable tool for the determination of the kinetic parameters of the blank polymerization (order of reaction with respect to monomer and initiator, activation energy). The conductimetric investigation of template polymerization of the acrylic monomer onto poly(allylamine) hydrochloride at low monomer concentration indicates that the reaction proceeds by a ‘zip’ mechanism. © 1999 Society of Chemical Industry