Advanced 3D Printing Strategies for the Controlled Delivery of Growth Factors.

The controlled delivery of growth factors (GFs) from tissue engineered constructs represents a promising strategy to improve tissue repair and regeneration. However, despite their established key role in tissue regeneration, the use of GFs is limited by their short half-life in the in vivo environment, their dose-dependent effectiveness, and their space- and time-dependent activity. Promising results have been obtained both in vitro and in vivo in animal models. Nevertheless, the clinical application of tissue engineered constructs releasing GFs is still challenging due to the several limitations and risks associated with their use. 3D printing and bioprinting, by allowing the microprecise spatial deposition of multiple materials and the fabrication of complex geometries with high resolution, offer advanced strategies for an optimal release of GFs from tissue engineered constructs. This review summarizes the strategies that have been employed to include GFs and their delivery system into biomaterials used for 3D printing applications to optimize their controlled release and to improve both the in vitro and in vivo regeneration processes. The approaches adopted to overcome the above-mentioned limitations are presented, showing the potential of the technology of 3D printing to get one step closer to clinical applications.

Mechanical Characterization of 3D-Printed Patterned Membranes for Cardiac Tissue Engineering: An Experimental and Numerical Study.

A myocardial infarction can cause irreversible damage to the heart muscle. A promising approach for the treatment of myocardial infarction and prevention of severe complications is the application of cardiac patches or epicardial restraint devices. The challenge for the fabrication of cardiac patches is the replication of the fibrillar structure of the myocardium, in particular its anisotropy and local elasticity. In this study, we developed a chitosan-gelatin-guar gum-based biomaterial ink that was fabricated using 3D printing to create patterned anisotropic membranes. The experimental results were then used to develop a numerical model able to predict the elastic properties of additional geometries with tunable elasticity that could easily match the mechanical properties of the heart tissue (particularly the myocardium).

Synergistic effect of CNTF and GDNF on directed neurite growth in chick embryo dorsal root ganglia.

It is often critical to improve the limited regenerative capacity of the peripheral nerves and direct neural growth towards specific targets, such as surgically implanted bioengineered constructs. One approach to accomplish this goal is to use extrinsic neurotrophic factors. The candidate factors first need to be identified and characterized in in vitro tests for their ability to direct the neurite growth. Here, we present a simple guidance assay that allows to assess the chemotactic effect of signaling molecules on the growth of neuronal processes from dorsal root ganglia (DRG) using only standard tissue culture materials. We used this technique to quantitatively determine the combined and individual effects of the ciliary neurotrophic factor (CNTF) and glial cell line-derived neurotrophic factor (GDNF) on neurite outgrowth. We demonstrated that these two neurotrophic factors, when applied in a 1:1 combination, but not individually, induced directed growth of neuronal processes towards the source of the gradient. This chemotactic effect persists without significant changes over a wide (10-fold) concentration range. Moreover, we demonstrated that other, more general growth parameters that do not evaluate growth in a specific direction (such as, neurite length and trajectory) were differentially affected by the concentration of the CNTF/GNDF mixture. Furthermore, GDNF, when applied individually, did not have any chemotactic effect, but caused significant neurite elongation and an increase in the number of neurites per ganglion.

Extraction and Physicochemical Characterization of Chitin from Cicada orni Sloughs of the South-Eastern French Mediterranean Basin.

Chitin is a structural polysaccharide of the cell walls of fungi and exoskeletons of insects and crustaceans. In this study, chitin was extracted, for the first time in our knowledge, from the Cicada orni sloughs of the south-eastern French Mediterranean basin by treatment with 1 M HCl for demineralization, 1 M NaOH for deproteinization, and 1% NaClO for decolorization. The different steps of extraction were investigated by Fourier Transform Infrared Spectroscopy (FTIR), X-Ray Diffraction (XRD), Thermogravimetric Analysis (TGA), and Scanning Electron Microscopy (SEM). Results demonstrated that the extraction process was efficiently performed and that Cicada orni sloughs of the south-eastern French Mediterranean basin have a high content of chitin (42.8%) in the α-form with a high degree of acetylation of 96% ± 3.4%. These results make Cicada orni of the south-eastern French Mediterranean basin a new and promising source of chitin. Furthermore, we showed that each step of the extraction present specific characteristics (for example FTIR and XRD spectra and, consequently, distinct absorbance peaks and values of crystallinity as well as defined values of maximum degradation temperatures identifiable by TGA analysis) that could be used to verify the effectiveness of the treatments, and could be favorably compared with other natural chitin sources.

Neurotrophic Effect of Fish-Lecithin Based Nanoliposomes on Cortical Neurons.

Lipids play multiple roles in preserving neuronal function and synaptic plasticity, and polyunsaturated fatty acids (PUFAs) have been of particular interest in optimizing synaptic membrane organization and function. We developed a green-based methodology to prepare nanoliposomes (NL) from lecithin that was extracted from fish head by-products. These NL range between 100-120 nm in diameter, with an n-3/n-6 fatty acid ratio of 8.88. The high content of n-3 PUFA (46.3% of total fatty acid content) and docosahexanoic acid (26%) in these NL represented a means for enrichment of neuronal membranes that are potentially beneficial for neuronal growth and synaptogenesis. To test this, the primary cultures of rat embryo cortical neurons were incubated with NL on day 3 post-culture for 24 h, followed by immunoblots or immunofluorescence to evaluate the NL effects on synaptogenesis, axonal growth, and dendrite formation. The results revealed that NL-treated cells displayed a level of neurite outgrowth and arborization on day 4 that was similar to those of untreated cells on day 5 and 6, suggesting accelerated synapse formation and neuronal development in the presence of NL. We propose that fish-derived NL, by virtue of their n-3 PUFA profile and neurotrophic effects, represent a new innovative bioactive vector for developing preventive or curative treatments for neurodegenerative diseases.