Structure-function assessment of 3D-printed porous scaffolds by a low-cost/open source fused filament fabrication printer.

Additive manufacturing encompasses a plethora of techniques to manufacture structures from a computational model. Among them, fused filament fabrication (FFF) relies on heating thermoplastics to their fusion point and extruding the material through a nozzle in a controlled pattern. FFF is a suitable technique for tissue engineering, given that allows the fabrication of 3D-scaffolds, which are utilized for tissue regeneration purposes. The objective of this study is to assess a low-cost/open-source 3D printer (In-House), by manufacturing both solid and porous samples with relevant microarchitecture in the physiological range (100-500 µm pore size), using an equivalent commercial counterpart for comparison. For this, compressive tests in solid and porous scaffolds manufactured in both printers were performed, comparing the results with finite element analysis (FEA) models. Additionally, a microarchitectural analysis was done in samples from both printers, comparing the measurements of both pore size and porosity to their corresponding computer-aided design (CAD) models. Moreover, a preliminary biological assessment was performed using scaffolds from our In-House printer, measuring cell adhesion efficiency. Finally, Fourier transform infrared spectroscopy - attenuated total reflectance (FTIR-ATR) was performed to evaluate chemical changes in the material (polylactic acid) after fabrication in each printer. The results show that the In-House printer achieved generally better mechanical behavior and resolution capacity than its commercial counterpart, by comparing with their FEA and CAD models, respectively. Moreover, a preliminary biological assessment indicates the feasibility of the In-House printer to be used in tissue engineering applications. The results also show the influence of pore geometry on mechanical properties of 3D-scaffolds and demonstrate that properties such as the apparent elastic modulus (Eapp) can be controlled in 3D-printed scaffolds.

Microporosity Clustering Assessment in Calcium Phosphate Bioceramic Particles.

There has been an increase in the application of different biomaterials to repair hard tissues. Within these biomaterials, calcium phosphate (CaP) bioceramics are suitable candidates, since they can be biocompatible, biodegradable, osteoinductive, and osteoconductive. Moreover, during sintering, bioceramic materials are prone to form micropores and undergo changes in their surface topographical features, which influence cellular physiology and bone ingrowth. In this study, five geometrical properties from the surface of CaP bioceramic particles and their micropores were analyzed by data mining techniques, driven by the research question: what are the geometrical properties of individual micropores in a CaP bioceramic, and how do they relate to each other? The analysis not only shows that it is feasible to determine the existence of micropore clusters, but also to quantify their geometrical properties. As a result, these CaP bioceramic particles present three groups of micropore clusters distinctive by their geometrical properties. Consequently, this new methodological clustering assessment can be applied to advance the knowledge about CaP bioceramics and their role in bone tissue engineering.

Molecular Regulation of Alternative Polyadenylation (APA) within the Drosophila Nervous System.

Alternative polyadenylation (APA) is a widespread gene regulatory mechanism that generates mRNAs with different 3'-ends, allowing them to interact with different sets of RNA regulators such as microRNAs and RNA-binding proteins. Recent studies have shown that during development, neural tissues produce mRNAs with particularly long 3'UTRs, suggesting that such extensions might be important for neural development and function. Despite this, the mechanisms underlying neural APA are not well understood. Here, we investigate this problem within the Drosophila nervous system, focusing on the roles played by general cleavage and polyadenylation factors (CPA factors). In particular, we examine the model that modulations in CPA factor concentration may affect APA during development. For this, we first analyse the expression of the Drosophila orthologues of all mammalian CPA factors and note that their expression decreases during embryogenesis. In contrast to this global developmental decrease in CPA factor expression, we see that cleavage factor I (CFI) expression is actually elevated in the late embryonic central nervous system, suggesting that CFI might play a special role in neural tissues. To test this, we use the UAS/Gal4 system to deplete CFI proteins from neural tissue and observe that in this condition, multiple genes switch their APA patterns, demonstrating a role of CFI in APA control during Drosophila neural development. Furthermore, analysis of genes with 3'UTR extensions of different length leads us to suggest a novel relation between 3'UTR length and sensitivity to CPA factor expression. Our work thus contributes to the understanding of the mechanisms of APA control within the developing central nervous system.

Identification and characterization of novel factors that act in the nonsense-mediated mRNA decay pathway in nematodes, flies and mammals.

Nonsense-mediated mRNA decay (NMD) is a surveillance mechanism that degrades mRNAs harboring premature termination codons (PTCs). We have conducted a genome-wide RNAi screen in Caenorhabditis elegans that resulted in the identification of five novel NMD genes that are conserved throughout evolution. Two of their human homologs, GNL2 (ngp-1) and SEC13 (npp-20), are also required for NMD in human cells. We also show that the C. elegans gene noah-2, which is present in Drosophila melanogaster but absent in humans, is an NMD factor in fruit flies. Altogether, these data identify novel NMD factors that are conserved throughout evolution, highlighting the complexity of the NMD pathway and suggesting that yet uncovered novel factors may act to regulate this process.