Leveling Up Hydrogels:Hybrid Systems in Tissue Engineering

Hydrogels can mimic several features of the cell native microenvironment and have been widely used as synthetic extracellular matrices (ECMs) in tissue engineering and regenerative medicine (TERM). However, some applications have specifications that hydrogels cannot efficiently fulfill on their own. Incorporating reinforcing structures like fibrous scaffolds or particles into hydrogels, as hybrid systems, is a promising strategy to improve their functionality. We describe recent advances in the fabrication and application of these hybrid systems, where structural properties and stimuli responsiveness of hydrogels are enhanced while their ECM-like features are preserved. Furthermore, we discuss how these systems can contribute to the development of more complex tissue engineered structures in the rapidly evolving field of TERM

Geometrically controlled liquefied capsules for modular tissue engineering strategies

A plethora of bioinspired cell-laden hydrogels are being explored as building blocks that once assembled are able to create complex and highly hierarchical structures recapitulating the heterogeneity of living tissues. Yet, the resulting 3D bioengineered systems still present key limitations, mainly related with limited diffusion of essential molecules for cell survival, which dictates the failure of most strategies upon implantation. To maximize the hierarchical complexity of bioengineered systems, while simultaneously fully addressing the exchange efficiency of biomolecules, the high-throughput fabrication of liquefied capsules is proposed using superhydrophobic-superhydrophilic microarrays as platforms to produce the initial structures with high fidelity of geometry and size. The liquefied capsules are composed by i) a permselective multilayered membrane; ii) surface-functionalized poly(ε-caprolactone) microparticles loaded into the liquefied core acting as cell adhesion sites; and iii) cells. It is demonstrated that besides the typical spherical liquefied capsules, it is also possible to obtain multi-shaped blocks with high geometrical precision and efficiency. Importantly, the internal gelation approach used to produce such blocks does not jeopardize cell viability, evidencing the mild conditions of the proposed cell encapsulation technique. The proposed system is intended to be used as hybrid devices implantable using minimally invasive procedures for multiple tissue engineering applications.publishe

Correction: Biofunctionalized pectin hydrogels as 3D cellular microenvironments

Correction for 'Biofunctionalized pectin hydrogels as 3D cellular microenvironments' by Sara C. Neves et al., J. Mater. Chem. B, 2015, 3, 2096–2108

Large-scale screening of unknown varieties in a grapevine intra-varietal variability collection

Since the last decade of the last century, it is known that many old grapevine varieties are descendants of other varieties through natural crossing. Portugal has an important program for the conservation of representative samples of intra-varietal variability of all autochthonous varieties, managed by the Portuguese Association for Grapevine Diversity (PORVID), which makes looking for genotypes with dubious identification an important activity from a perspective of its valorisation. This communication presents the results of the molecular analysis of 5,000 samples (accessions) from the PORVID’s collection, using nine microsatellite loci currently recommended by the International Organization of Vine and Wine (OIV) for genetic grapevine identification. The results obtained confirmed the molecular identity of 4,220 samples corresponding to 214 varieties present in the official list of Portuguese varieties. In 780 samples, 95 profiles with a plural number of accessions revealed not to be listed in the Vitis International Variety Catalogue (VIVC) database, corresponding to possible varieties either descendent from natural crossing from at least one known parental variety, or from undetermined origin. Furthermore, the need for a comprehensive strategy aimed at uncovering other hidden varieties is discussed to prevent their imminent loss, deepen understanding of their origin, and add economic value and sustainability to the vine and wine sector

Discriminating varieties throughout the wine chain: applying SNP markers and High Resolution Melting analysis to wine authenticity

Trabajo presentado en el IV International Meeting of the Portuguese Society of Genetics, celebrado en Braga (Portugal), los días 19 y 20 de octubre de 2023Wine authenticity assessment throughout different winemaking steps is a topic of interest for consumers and producers. The correct identification of grapevine varieties is a main goal of different authenticity strategies. PCR amplification and capillary electrophoresis of microsatellite markers is the standard DNA-based method for varietal identification. Although successfully applied to vines, it reveals to be unreliable when applied to DNA extracted from must and wine, mainly due to the degradation and contamination of the DNA present in these matrices. High Resolution Melting has been proposed as a PCR-based alternative for this purpose, but it could benefit in targeting fragments of small size, when analysing must or wine DNA samples. This study aimed to discriminate five grapevine varieties using DNA extracted from samples with different complexity, using HRM analysis of three SNP markers1 identified in fragments of approximately 100 bp. We used Cabernet Sauvignon as the international reference and four Portuguese varieties - Touriga Franca and Touriga Nacional, two grapevine varieties commonly used in Portugal, and Donzelinho Tinto and Rufete, two less utilized autochthonous varieties. HRM assays were performed using gDNA extracted from leaf, monovarietal must and monovarietal wine samples. Using the melting profiles obtained from leaf gDNA as reference, varietal discrimination was remarkably achieved in must gDNA samples from all varieties in study. In relation to wine gDNA, Cabernet Sauvignon and Rufete samples showed to be the most troublesome in giving reproducible results. The varieties were clustered into different groups according to their genotype, in line with the data from the ICVV-SNP database. Considering the sensitivity and precision of this methodology, our results show that this approach is promising to analyse samples collected throughout the wine production chain

Leveling Up Hydrogels: Hybrid Systems in Tissue Engineering

Hydrogels can mimic several features of the cell native microenvironment and have been widely used as synthetic extracellular matrices (ECMs) in tissue engineering and regenerative medicine (TERM). However, some applications have specifications that hydrogels cannot efficiently fulfill on their own. Incorporating reinforcing structures like fibrous scaffolds or particles into hydrogels, as hybrid systems, is a promising strategy to improve their functionality. We describe recent advances in the fabrication and application of these hybrid systems, where structural properties and stimuli responsiveness of hydrogels are enhanced while their ECM-like features are preserved. Furthermore, we discuss how these systems can contribute to the development of more complex tissue engineered structures in the rapidly evolving field of TERM

Identification of Portuguese traditional grapevines using molecular marker-based strategies

Precise identification is crucial to study and for the valorisation of ancient grapevine varieties, with many synonyms and misidentifications hampering this task. SSR-Multiplex PCR and SNP genotyping combined with HRM were used, to identify 19 varieties present in an ancient Portuguese vineyard. The combination of the methodologies allowed the identification of 18 grapevines, from which 10 were correctly identified by ampelography and the remaining were misidentified. SSR and SNP data retrieved for one sample did not match any profile publicly available, however pedigree analysis suggests it is a new variety. One sample identified as Cerceal Branco presented an unexpected red berry. A sample identified as Malvasia Preta/Preta Roxa also presented a berry colour non coincident with its molecular identification. This suggests they might represent new somatic variants of existent varieties. Three HRM assays were performed to test its robustness in comparison to SSR/SNP results. Molecular identification was successful, revealing possible new variants. HRM revealed to be useful in identifying varieties by melting curve profile comparison against reference material. The results provide a better knowledge of a valuable pool of genetic patrimony: ancient varieties that represent a piece of history and heritage from an important winegrowing region.This work was funded by the Portuguese Foundation for Science and Technology (FCT), Portugal, grant numbers UIDB/04138/2020, and UIDP/04138/2020. We also acknowledge FCT financial support to S.B. (SFRH/BD/146346/2019)

Additive manufactured polymeric 3D scaffolds with tailored surface topography influence mesenchymal stromal cells activity

Additive manufactured three-dimensional (3D) scaffolds with tailored surface topography constitute a clear advantage in tissue regeneration strategies to steer cell behavior. 3D fibrous scaffolds of poly (ethylene oxide terephthalate)/poly(butylene terephthalate) block copolymer presenting different fiber surface features were successfully fabricated by additive manufacturing combined with wet-spinning, in a single step, without any post-processing. The optimization of the processing parameters, mainly driven by different solvent/non-solvent combinations, led to four distinct scaffold types, with average surface roughness values ranging from 0.071 +/- 0.012 mu m to 1.950 +/- 0.553 mu m, average pore sizes in the x-and y-axis between 351.1 +/- 33.6 mu m and 396.1 +/- 32.3 mu m, in the z-axis between 36.5 +/- 5.3 mu m and 70.7 +/- 8.8 mu m, average fiber diameters between 69.4 +/- 6.1 mu m and 99.0 +/- 9.4 mu m, and porosity values ranging from 60.2 +/- 0.8% to 71.7 +/- 2.6%. Human mesenchymal stromal cells (hMSCs) cultured on these scaffolds adhered, proliferated, and produced endogenous extracellular matrix. The effect of surface roughness and topography on hMSCs differentiation was more evident for cells seeded at lower density, where the percentage of cells in direct contact with the surface was higher compared to more densely seeded scaffolds. Under osteogenic conditions, lower surface roughness values (0.227 +/- 0.035 mu m) had a synergistic effect on hMSCs behavior, while chondrogenesis was favored on rougher surfaces (1.950 +/- 0.553 mu m)

From the Field to the Bottle—An Integrated Strategy for Wine Authenticity

The wine sector is one of the most economically important agro-food businesses. The wine market value is largely associated to terroir, in some cases resulting in highly expensive wines that attract fraudulent practices. The existent wine traceability system has some limitations that can be overcome with the development of new technological approaches that can tackle this problem with several means. This review aims to call attention to the problem and to present several strategies that can assure a more reliable and authentic wine system, identifying existent technologies developed for the sector, which can be incorporated into the current traceability system