Sara C. Neves

Chitosan/Poly(ɛ-caprolactone) blend scaffolds for cartilage repair

Chitosan (CHT)/poly(ɛ-caprolactone) (PCL) blend 3D fiber-mesh scaffolds were studied as possible support structures for articular cartilage tissue (ACT) repair. Micro-fibers were obtained by wet-spinning of three different polymeric solutions: 100:0 (100CHT), 75:25 (75CHT) and 50:50 (50CHT) wt.% CHT/PCL, using a common solvent solution of 100 vol.% of formic acid. Scanning electron microscopy (SEM) analysis showed a homogeneous surface distribution of PCL. PCL was well dispersed throughout the CHT phase as analyzed by differential scanning calorimetry and Fourier transform infrared spectroscopy. The fibers were folded into cylindrical moulds and underwent a thermal treatment to obtain the scaffolds. μCT analysis revealed an adequate porosity, pore size and interconnectivity for tissue engineering applications. The PCL component led to a higher fiber surface roughness, decreased the scaffolds swelling ratio and increased their compressive mechanical properties. Biological assays were performed after culturing bovine articular chondrocytes up to 21 days. SEM analysis, live-dead and metabolic activity assays showed that cells attached, proliferated, and were metabolically active over all scaffolds formulations. Cartilaginous extracellular matrix (ECM) formation was observed in all formulations. The 75CHT scaffolds supported the most neo-cartilage formation, as demonstrated by an increase in glycosaminoglycan production. In contrast to 100CHT scaffolds, ECM was homogenously deposited on the 75CHT and 50CHT scaffolds. Although mechanical properties of the 50CHT scaffold were better, the 75CHT scaffold facilitated better neo-cartilage formation.

Biofunctionalized pectin hydrogels as 3D cellular microenvironments

In situ-forming hydrogels of pectin, a polysaccharide present in the cell wall of higher plants, were prepared using an internal ionotropic gelation strategy based on calcium carbonate/d-glucono-δ-lactone, and explored for the first time as cell delivery vehicles. Since no ultrapure pectins are commercially available yet, a simple and efficient purification method was established, effectively reducing the levels of proteins, polyphenols and endotoxins of the raw pectin. The purified pectin was then functionalized by carbodiimide chemistry with a cell-adhesive peptide (RGD). Its gelation was analyzed by rheometry and optimized. Human mesenchymal stem cells embedded within unmodified and RGD-pectin hydrogels of different viscoelasticities (1.5 and 2.5 wt%) remained viable and metabolically active for up to 14 days. On unmodified pectin hydrogels, cells remained isolated and round-shaped. In contrast, within RGD-pectin hydrogels they elongated, spread, established cell-to-cell contacts, produced extracellular matrix, and migrated outwards the hydrogels. After 7 days of subcutaneous implantation in mice, acellular pectin hydrogels were considerably degraded, particularly the 1.5 wt% hydrogels. Altogether, these findings show the great potential of pectin-based hydrogels, which combine an interesting set of easily tunable properties, including the in vivo degradation profile, for tissue engineering and regenerative medicine.

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 μm to 1.950 ± 0.553 μm, average pore sizes in the x- and y-axis between 351.1 ± 33.6 μm and 396.1 ± 32.3 μm, in the z-axis between 36.5 ± 5.3 μm and 70.7 ± 8.8 μm, average fiber diameters between 69.4 ± 6.1 μm and 99.0 ± 9.4 μm, 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 μm) had a synergistic effect on hMSCs behavior, while chondrogenesis was favored on rougher surfaces (1.950 ± 0.553 μm).

A laparoscopic surgery training interface

Laparoscopy is a surgical procedure on which operations in the abdomen are performed through small incisions using several specialized instruments. The laparoscopic surgery success greatly depends on surgeon skills and training. To achieve these technical high-standards, different apprenticeship methods have been developed, many based on in vivo training, an approach that involves high costs and complex setup procedures. This paper explores Virtual Reality (VR) simulation as an alternative for novice surgeons training. Even though several simulators are available on the market claiming successful training experiences, their use is extremely limited due to the economic costs involved. In this work, we present a low-cost laparoscopy simulator able to monitor and assist the trainees surgical movements. The developed prototype consists of a set of inexpensive sensors, namely an accelerometer, a gyroscope, a magnetometer and a flex sensor, attached to specific laparoscopic instruments. Our approach allows repeated assisted training of an exercise, without time constraints or additional costs, since no human artificial model is needed. A case study of our simulator applied to instrument manipulation practice (hand-eye coordination) is also presented.

Cyanobacterium-Derived Extracellular Carbohydrate Polymer for the Controlled Delivery of Functional Proteins.

The unicellular cyanobacterium Cyanothece sp. CCY 0110 is a highly efficient producer of extracellular polymeric substances (EPS), releasing up to 75% of the polymer to the culture medium. The carbohydrate polymer released to the medium (RPS) was previously isolated and characterized; it is composed of nine different monosaccharides including two uronic acids, and also containing peptides and sulfate groups. Here it is shown that the RPS spontaneously assembles with proteins at high concentrations leading to a phase transition. The proteins are released progressively and structurally intact near physiological conditions, primarily through the swelling of the polymer-protein matrix. The releasing kinetics of the proteins can be modulated through the addition of divalent cations, such as calcium. Notably, the polymer is not toxic to human dermal neonatal fibroblasts in vitro at RPS concentrations bellow 0.1 mg mL-1 . The results show that this polymer is a good candidate for the delivery of therapeutic macromolecules.

In vitro interaction of polymeric biomaterials with cells

The analysis of cell-material interactions is a key step in biomaterials development. It is fundamental for designing optimal biomaterial formulations and also required at the final stage of in vitro testing, providing rational criteria for the acceptance or rejection of candidate implant materials and biomedical devices. Cell response to a biomaterial or its leachates can be evaluated at different levels, namely, in terms of cell morphology, viability, metabolic activity, proliferation, migration, and differentiation. Yet, classical cell/molecular biology protocols, originally tailored for measuring events in cells cultured as monolayers, may be inadequate in experimental setups involving, for example, 3D culture on scaffolds. Adjustment, validation, and careful interpretation of obtained data are thus required, as discussed herein. The emergence of new or upgraded assays, advanced bioimaging tools and high-throughput screening platforms has provided important contributions to the field.