Mario D'Acunto

Morpho-functional characterization of human mesenchymal stem cells from umbilical cord blood for potential uses in regenerative medicine

Mesenchymal stem cells (MSCs) represent a promising source of progenitor cells having the potential to repair and to regenerate diseased or damaged skeletal tissues. Bone marrow (BM) has been the first source reported to contain MSCs. However, BM-derived cells are not always acceptable, due to the highly invasive drawing and the decline in MSC number and differentiative capability with increasing age. Human umbilical cord blood (UCB), obtainable by donation with a noninvasive method, has been introduced as an alternative source of MSCs. Here human UCB-derived MSCs isolation and morpho-functional characterization are reported. Human UCB-derived mononuclear cells, obtained by negative immunoselection, exhibited either an osteoclast-like or a mesenchymal-like phenotype. However, we were able to obtain homogeneous populations of MSCs that displayed a fibroblast-like morphology, expressed mesenchym-related antigens and showed differentiative capacities along osteoblastic and early chondroblastic lineages. Furthermore, this study is one among a few papers investigating human UCB-derived MSC growth and differentiation on three-dimensional scaffolds focusing on their potential applications in regenerative medicine and tissue engineering. UCB-derived MSCs were proved to grow on biodegradable microfiber meshes; additionally, they were able to differentiate toward mature osteoblasts when cultured inside human plasma clots, suggesting their potential application in orthopedic surgery.

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.

NEW BIOARTIFICIAL SYSTEMS AND BIODEGRADABLE SYNTHETIC POLYMERS FOR CARDIAC TISSUE ENGINEERING: A PRELIMINARY SCREENING

The aim of this work was the preparation and characterization of new polymeric biomaterials for application in myocardial tissue engineering. The attention was firstly focused on new bioartificial polymeric systems, with the aim to combine the features of synthetic polymers with the specific cell and tissue compatibility of biopolymers. In this work, alginate, collagen, and gelatin were used as the natural component and poly(N-isopropylacrylamide) was used as the synthetic component. The characterization included morphological, topographical, and mechanical analyses, thermogravimetric characterization, infrared spectroscopy, and cell culture tests. For the biological characterization, C2C12 myoblasts were cultured on different materials and cell adhesion, proliferation, and differentiation were evaluated. The morphological, topographical, and mechanical analyses, as well as the biological characterization, were also applied to a tri-block poly(ester-ether-ester) copolymer, obtained by reaction of preformed poly(ethylene glycol) with e-caprolactone, and a novel poly(ester urethane) obtained by using an L-lisine-derived diisocyanate, giving nontoxic degradation products. The encouraging results obtained in this work allowed us to select some of the new bioartificial polymers, the synthetic tri-block copolymer, and the polyurethane as potential good materials to prepare scaffolds for myocardial tissue engineering.

Boron nitride nanotube-functionalised myoblast/microfibre constructs: a nanotech-assisted tissue-engineered platform for muscle stimulation

In this communication, we introduce boron nitride nanotube (BNNT)‐functionalised muscle cell/microfibre mesh constructs, obtained via tissue engineering, as a three‐dimensional (3D) platform to study a wireless stimulation system for electrically responsive cells and tissues. Our stimulation strategy exploits the piezoelectric behaviour of some classes of ceramic nanoparticles, such as BNNTs, able to polarize under mechanical stress, e.g. using low‐frequency ultrasound (US). In the microfibre scaffolds, C2C12 myoblasts were able to differentiate into viable myotubes and to internalize BNNTs, also upon US irradiation, so as to obtain a nanotech‐assisted 3D in vitro model. We then tested our stimulatory system on 2D and 3D cellular models by investigating the expression of connexin 43 (Cx43), as a molecule involved in cell crosstalk and mechanotransduction, and myosin, as a myogenic differentiation marker. Cx43 gene expression revealed a marked model dependency. In control samples (without US and/or BNNTs), Cx43 was upregulated under 2D culture conditions (10.78 ± 1.05‐fold difference). Interactions with BNNTs increased Cx43 expression in 3D samples. Cx43 mRNA dropped in 2D under the ‘BNNTs + US’ regimen, while it was best enhanced in 3D samples (3.58 ± 1.05 vs 13.74 ± 1.42‐fold difference, p = 0.0001). At the protein level, the maximal expressions of Cx43 and myosin were detected in the 3D model. In contrast with the 3D model, in 2D cultures, BNNTs and US exerted a synergistic depletive effect upon myosin synthesis. These findings indicate that model dimensionality and stimulatory regimens can strongly affect the responses of signalling and differentiation molecules, proving the importance of developing proper in vitro platforms for biological modelling. Copyright

Acrylic copolymers as candidates for drug-eluting coating of vascular stents.

The aim of the present work is the synthesis and characterization of polymer materials showing good adhesion, drug loading, and delivery properties, for potential cardiovascular application. In particular, poly(methylmethacrylate-co-acrylic acid) copolymers are prepared in different compositions by a radical polymerization and investigated as potential materials to coat metallic stents and to carry out a local drug release. Films obtained by dissolving the copolymer in an appropriate organic solvent (also loaded with an anti-restenosis drug, such as tacrolimus) are investigated: physicochemical properties, adhesiveness to metallic stent material, and kinetics of drug release in physiological environment are studied.

Good Manufacturing Practices—Grade Preformed Ossicular Prostheses from Banked Bone via Computer Numerically Controlled Micromilling

Objectives: The aim of this study was the fabrication of ossicular replacement prostheses (ORPs) from decellularized banked cortical bone via computer numerically controlled (CNC) ultraprecision micromilling, in order to obtain preformed clinical-grade tissue products, reproducing shape, size, and details perfectly comparable to those of synthetic devices. Methods: Banked femoral compact bone was used to fabricate partial and total ORPs via CNC micromilling according to Good Manufacturing Practices procedures. Drawings of ORPs with different shapes and sizes were uploaded to the computer interface, and different surface-finish parameters were tested. The obtained products underwent dimensional, weight, and surface characterizations. A histologic analysis was pursued to compare the bone matrix compactness of the produced ORPs to that of the ear ossicles. Results: Banked-bone ORPs were produced with high dimensional accuracy. Partial ORP weights averaged (±SD) 31.2 ± 0.6 mg, and total ORP weights averaged 69.3 ± 0.7 mg. The best-finish mode allowed microscale or nanoscale roughness free from machinery textures to be obtained. Finally, the histologic analysis confirmed that the extracellular matrix compactness of the produced ORPs was suitable for ossicular chain replacement. Conclusions: This study assesses the fabrication feasibility of novel banked-bone ORPs of extremely high dimensional accuracy. Such devices are aimed at combining the most favorable aspects of both synthetic (reproducibility, convenience, and biosafety) and biological replacements (total biocompatibility).

Composites Between Alumina and an Ester‐Ether‐Ester Bioresorbable Copolymer

Composites between alumina and the biore-sorbable (poly(e-caprolactone)-block-poly(oxyethylene)-block-poly(e-caprolactone) copolymer were obtained by reacting e-caprolactone with preformed poly(ethylene glycol), in the presence of ceramic alumina powder, at 185°C under vacuum. The mechanical properties, tested by compression and flexural strengths and Youngs modulus, show that the copolymer interacts poorly with the alumina grains. Both scanning electron and atomic force microscopy show a scare wettability between alumina and copolymer, as well as the aggregation of alumina micro-particles into clusters of big size. Both mechanical and morphological tests seem to indicate a stronger interaction between the alumina micro-particles than between the alumina surface and the reaction mixture during the polymerization, as well as a compacting effet by alumina on the forming copolymer. The FT-IR spectra of the composites show both copolymer and alumina absorption bands. The FT-IR analysis on the fractions of an extraction which CHCl 3 indicates the presence of traces of poly(e-caprolactone), stably linked to alumina. The polymerization of e-caprolactone with alumina alone in the same conditions gives poly(e-caprolactone), mainly free and in minor part linked to the alumina surface. Two polymerization mechanisms, simultaneously occuring, are proposed. The most relevant result of this work is the lack of chemical inertness of alumina towards e-caprolactone, which leads to reconsider also the use of alumina as a biochemically inert material.