Preparation and characterization of alginate/gelatin blend films for cardiac tissue engineering

The aim of this work was the preparation of blends based on alginate and gelatin, with different weight ratio, to combine the advantages of these two natural polymers for application in cardiac tissue engineering. The physicochemical characterization, performed by Fourier transform infrared spectroscopy, differential scanning calorimetry and thermogravimetric analysis, revealed a good miscibility and the presence of interactions among the functional groups of pure biopolymers. Concerning the swelling and degradation tests, performed in different solutions simulating body fluids, both swelling degree and weight losses were higher in phosphate buffer saline (PBS) and for the blends with a higher content of gelatin. These results indicated a better stability of the blends in cell culture medium than in PBS and suggested a mainly hydrolytic degradation process. Cell culture tests, carried out using C2C12 myoblasts, showed a good cell proliferation for all the blends containing more than 60% of gelatin, with the alginate/gelatin 20:80 showing the best response. The same blend was the only one on which cell differentiation was observed. The results obtained in the biological characterization allow to select the alginate/gelatin 20:80 blend as a suitable material to prepare scaffolds for myocardial tissue engineering.     

Synthesis and characterization of copolymers of methylmethacrylate and 2-hydroxyethyl methacrylate for the aqueous solubilization of Paclitaxel

The aim of the present work is the modification of a hydrophobic polymeric macromolecule, polymethylmethacrylate, by introducing hydrophilic moieties of 2-hydroxyethyl methacrylate within the polymer chain. Synthesis, characterization, and drug delivery control capabilities exerted on a highly hydrophobic drug (Paclitaxel) are illustrated. In particular, the dependency of the drug delivery kinetic on the fraction of hydrophilic units inserted in the copolymer chain was studied. Results showed that it is possible to have an increase of the kinetic delivery introducing hydrophilic units. In addition, a double control, diffusive and due to the relaxation of the molecules, on drug delivery was obtained.     

Cardioprotection of PLGA/gelatine cardiac patches functionalised with adenosine in a large animal model of ischaemia and reperfusion injury: A feasibility study

The protection from ischaemia‐reperfusion‐associated myocardial infarction worsening remains a big challenge. We produced a bioartificial 3D cardiac patch with cardioinductive properties on stem cells. Its multilayer structure was functionalised with clinically relevant doses of adenosine. We report here the first study on the potential of these cardiac patches in the controlled delivery of adenosine into the in vivo ischaemic‐reperfused pig heart. A Fourier transform infrared chemical imaging approach allowed us to perform a characterisation, complementary to the histological and biochemical analyses on myocardial samples after in vivo patch implantation, increasing the number of investigations and results on the restricted number of pigs (n = 4) employed in this feasibility step. In vitro tests suggested that adenosine was completely released by a functionalised patch, a data that was confirmed in vivo after 24 hr from patch implantation. Moreover, the adenosine‐loaded patch enabled a targeted delivery of the drug to the ischaemic‐reperfused area of the heart, as highlighted by the activation of the pro‐survival signalling reperfusion injury salvage kinases pathway. At 3 months, though limited to one animal, the used methods provided a picture of a tissue in dynamic conditions, associated to the biosynthesis of new collagen and to a non‐fibrotic outcome of the healing process underway. The synergistic effect between the functionalised 3D cardiac patch and adenosine cardioprotection might represent a promising innovation in the treatment of reperfusion injury. As this is a feasibility study, the clinical implications of our findings will require further in vivo investigation on larger numbers of ischaemic‐reperfused pig hearts.     

Polymerisation onto Biological Templates,a New Way to Obtain Bioartificial Polymeric Materials

The radical polymerisation of synthetic monomers onto biological templates can be recognised as a technique for the straightforward preparation of bioartificial polymeric materials, polymer blends based materials in which interactions at the molecular level lead to enhanced properties. In the first part the polymerisations of different acrylic monomers in the presence of synthetic templates are presented as study models. The second part deals with preliminary experiments carried out using natural polymers as templates. Dilatometric and conductimetric measurements were used to study the kinetics of the template polymerisation reactions. Results indicate that the reactions proceed according to different mechanisms depending on the interactions between functional groups present in the systems. In many instances the results of the characterisation analyses have shown that the polymer complexes obtained by template polymerisation have a more ordered structure than the complexes prepared by simple mixing the two polymers. In addition conductimetry has revealed as a very simple diagnostic tool for identifying the template reaction mechanism.     

Three-dimensional microfabricated scaffolds with cardiac extracellular matrix-like architecture

In recent years, research in the field of myocardial tissue engineering has advanced thanks to the development of new biomaterials and a more clear understanding of processes that are at the basis of cardiac tissue growth. However, classical porous scaffolds developed during these years to try to reconstruct and mimic heart function have proven to be inadequate because they are not able to reproduce the typical myocardial environment. One approach to increase functionality of tissue-engineered constructs relies on attempts to mimic the microarchitecture of natural tissues, since it is well known that topology is one of the principal stimuli that cells need to activate their functions. The aim of this work was the realization of three-dimensional microfabricated scaffolds, with cardiac extracellular matrix (ECM)-like architecture. For this purpose, samples of pig myocardium were decellularized, embedded in paraffin wax and analyzed under an optical microscope, in order to evaluate the geometrical features of the cardiac ECM. On the basis of these data, a simplified model of the cardiac ECM microarchitecture was designed. Microfabricated scaffolds were realized with Soft Lithography technique, using a bioartificial blend, based on alginate, gelatin and a novel poly(N-isopropylacrylamide)-based copolymer, which we synthesized. The scaffolds were characterized in terms of topological and mechanical properties. Moreover, cell adhesion, proliferation, and differentiation tests were performed. The microfabricated scaffolds showed they matched the anisotropic mechanical properties of adult human left ventricular myocardium, while at the same time being able to promote myoblast alignment in the absence of external stimuli.     

The value of lung ultrasound monitoring in H1N1 acute respiratory distress syndrome

We present the case of a healthy young male who developed acute respiratory failure as a result of infection with influenza A/H1N1 of swine‐origin and in whom ventilatory support was optimised and recovery of lung function was monitored by the use of sequential chest ultrasound examinations. The potential pivotal role of bedside lung ultrasonography in H1N1‐induced respiratory failure is discussed.     

Molecularly imprinted bioartificial membranes for the selective recognition of biological molecules

Membranes of a synthetic (poly(ethylene-co-vinyl alcohol), Clarene) and a biological (dextran) polymer, imprinted with alpha-amylase, of different compositions were prepared by the phase-inversion process. Membrane morphologies were shown to be dependent on the synthetic-biological components composition. The removal of the template from the membranes was performed by extraction with water, while an aqueous solution of alpha-amylase was permeated across the membranes under pressure to obtain the rebinding of the template. The selectivity of alpha-amylase-imprinted membranes was investigated by the same uptake experiment using pepsin, albumin and amyloglucosidase, and the rebinding of these proteins was compared with that of the print molecule. Before and after template extraction and after the rebinding experiment, kinetic measurements of the imprinting molecule were conducted to estimate the activity of the enzyme immobilised in the polymer matrix. Results obtained revealed that the immobilised enzyme maintains a good functionality while in the membrane compared to the free enzyme and the imprinted 'bioartificial' dextran and Clarene membranes, obtained by the phase-inversion method, can establish efficient interaction with alpha-amylase as template molecule, as confirmed by the fair selectivity in rebinding tests.     

Polyelectrolyte complexes obtained by radical polymerization in the presence of chitosan

The radical polymerization of acrylic acid and sodium 4-styrenesulfonate in the presence of chitosan as a template gives insoluble products, identified as the polyelectrolyte complexes chitosan-poly(acrylic acid) and chitosan-poly(4-styrenesulfonate). Kinetic results do not permit to propose any mechanism in the first case, while suggest a “pick-up” one in the second. The polyelectrolyte complexes have been characterized by FT-IR spectroscopy, X-ray diffractometry, optical and scanning electron microscopies, differential scanning calorimetry and thermogravimetric analysis. The results obtained indicate an ordered structure for the first complex, while the second one appears similar to that obtained by reacting the parent polymers. Therefore, the template polymerization technique appears advantageous only for the synthesis of the chitosan-poly(acrylic acid) complex. The thermal analysis shows that the complexes undergo two successive modifications on heating. FT-IR analysis demonstrates that the first process is an esterification between the hydroxyls of chitosan and the acidic groups of both daughter polymers; the second one appears to be an amidation of the chitosan ammonium groups only with the sulfonate groups.