Calcium Carbonate Coating of 3D-Printed PLA Scaffolds Intended for Biomedical Applications.

The incorporation of ceramic additives is the most commonly used strategy to improve the biofunctionality of polymer-based scaffolds intended for bone regeneration. By embedding ceramic particles as a coating, the functionality improvement in the polymeric scaffolds can be concentrated on the cell-surface interface, thus creating a more favourable environment for the adhesion and proliferation of osteoblastic cells. In this work, a pressure-assisted and heat-induced method to coat polylactic acid (PLA) scaffolds with calcium carbonate (CaCO3) particles is presented for the first time. The coated scaffolds were evaluated by optical microscopy observations, a scanning electron microscopy analysis, water contact angle measurements, compression testing, and an enzymatic degradation study. The ceramic particles were evenly distributed, covered more than 60% of the surface, and represented around 7% of the coated scaffold weight. A strong bonding interface was achieved, and the thin layer of CaCO3 (~20 µm) provided a significant increase in the mechanical properties (with a compression modulus improvement up to 14%) while also enhancing the surface roughness and hydrophilicity. The results of the degradation study confirmed that the coated scaffolds were able to maintain the pH of the media during the test (~7.6 ± 0.1), in contrast to the pure PLA scaffolds, for which a value of 5.07 ± 0.1 was obtained. The ceramic-coated scaffolds developed showed potential for further evaluations in bone tissue engineering applications.

Effects of ceramic additives and bioactive coatings on the degradation of polylactic acid-based bone scaffolds under hydrolytic conditions.

Polylactic acid (PLA) has been extensively used for the manufacturing of scaffolds in bone tissue engineering applications. Due to the low hydrophilicity and the acidic degradation process of this biomaterial, different strategies have been proposed to increase the biofunctionality of the support structure. The use of ceramic particles is a generally preferred option to increase the osteoconductivity of the base material, while acting as buffers to maintain the pH level of the surroundings tissues. Surface modification is another approach to overcome the limitations of PLA for tissue engineering applications. In this work, the degradation profile of 3D-printed PLA scaffolds containing beta-tricalcium phosphate (ß-TCP) and calcium carbonate (CaCO3 ) particles has been studied under hydrolytic conditions. Composite samples treated with plasma and coated with Aloe vera extracts were also studied to evaluate the effect of this surface modification method. The characterization of the 3D structures included its morphological, calorimetric and mechanical evaluation. According to the results obtained, the proposed composite scaffolds allowed an adequate maintenance of the pH level of the surrounding medium, with no effects observed on the morphology and mechanical properties of these structures. Hence, these samples showed potential to be further investigated as candidates for bone tissue regeneration.

Translation through collaboration: practice applied in BAMOS project in in vivo testing of innovative osteochondral scaffolds.

Osteoarthritis is the most common chronic degenerative joint disease, recognized by the World Health Organization as a public health problem that affects millions of people worldwide. The project Biomaterials and Additive Manufacturing: Osteochondral Scaffold (BAMOS) innovation applied to osteoarthritis, funded under the frame of the Horizon 2020 Research and Innovation Staff Exchanges (RISE) program, aims to delay or avoid the use of joint replacements by developing novel cost-effective osteochondral scaffold technology for early intervention of osteoarthritis. The multidisciplinary consortium of BAMOS, formed by international leading research centres, collaborates through research and innovation staff exchanges. The project covers all the stages of the development before the clinical trials: design of scaffolds, biomaterials development, processability under additive manufacturing, in vitro test, and in vivo test. This paper reports the translational practice adopted in the project in in vivo assessment of the osteochondral scaffolds developed.

In vivo evaluation of additively manufactured multi-layered scaffold for the repair of large osteochondral defects.

The repair of osteochondral defects is one of the major clinical challenges in orthopaedics. Well-established osteochondral tissue engineering methods have shown promising results for the early treatment of small defects. However, less success has been achieved for the regeneration of large defects, which is mainly due to the mechanical environment of the joint and the heterogeneous nature of the tissue. In this study, we developed a multi-layered osteochondral scaffold to match the heterogeneous nature of osteochondral tissue by harnessing additive manufacturing technologies and combining the established art laser sintering and material extrusion techniques. The developed scaffold is based on a titanium and polylactic acid matrix-reinforced collagen "sandwich" composite system. The microstructure and mechanical properties of the scaffold were examined, and its safety and efficacy in the repair of large osteochondral defects were tested in an ovine condyle model. The 12-week in vivo evaluation period revealed extensive and significantly higher bone in-growth in the multi-layered scaffold compared with the collagen-HAp scaffold, and the achieved stable mechanical fixation provided strong support to the healing of the overlying cartilage, as demonstrated by hyaline-like cartilage formation. The histological examination showed that the regenerated cartilage in the multi-layer scaffold group was superior to that formed in the control group. Chondrogenic genes such as aggrecan and collagen-II were upregulated in the scaffold and were higher than those in the control group. The findings showed the safety and efficacy of the cell-free "translation-ready" osteochondral scaffold, which has the potential to be used in a one-step surgical procedure for the treatment of large osteochondral defects. Supplementary Information: The online version contains supplementary material available at 10.1007/s42242-021-00177-w.

Experimental Analysis of the Enzymatic Degradation of Polycaprolactone: Microcrystalline Cellulose Composites and Numerical Method for the Prediction of the Degraded Geometry.

The degradation rate of polycaprolactone (PCL) is a key issue when using this material in Tissue Engineering or eco-friendly packaging sectors. Although different PCL-based composite materials have been suggested in the literature and extensively tested in terms of processability by material extrusion additive manufacturing, little attention has been paid to the influence of the fillers on the mechanical properties of the material during degradation. This work analyses the possibility of tuning the degradation rate of PCL-based filaments by the introduction of microcrystalline cellulose into the polymer matrix. The enzymatic degradation of the composite and pure PCL materials were compared in terms of mass loss, mechanical properties, morphology and infrared spectra. The results showed an increased degradation rate of the composite material due to the presence of the filler (enhanced interaction with the enzymes). Additionally, a new numerical method for the prediction of the degraded geometry was developed. The method, based on the Monte Carlo Method in an iterative process, adjusts the degradation probability according to the exposure of each discretized element to the degradation media. This probability is also amplified depending on the corresponding experimental mass loss, thus allowing a good fit to the experimental data in relatively few iterations.

On the Effectiveness of Oxygen Plasma and Alkali Surface Treatments to Modify the Properties of Polylactic Acid Scaffolds.

Surface modification of 3D-printed PLA structures is a major issue in terms of increasing the biofunctionality and expanding the tissue engineering applications of these parts. In this paper, different exposure times were used for low-pressure oxygen plasma applied to PLA 3D-printed scaffolds. Alkali surface treatments were also evaluated, aiming to compare the modifications introduced on the surface properties by each strategy. Surface-treated samples were characterized through the quantification of carboxyl groups, energy-dispersive X-ray spectroscopy, water contact angle measurements, and differential scanning calorimetry analysis. The change in the surface properties was studied over a two-week period. In addition, an enzymatic degradation analysis was carried out to evaluate the effect of the surface treatments on the degradation profile of the 3D structures. The physicochemical characterization results suggest different mechanism pathways for each type of treatment. Alkali-treated scaffolds showed a higher concentration of carboxyl groups on their surface, which enhanced the enzymatic degradation rate, but were also proven to be more aggressive towards 3D-printed structures. In contrast, the application of the plasma treatments led to an increased hydrophilicity of the PLA surface without affecting the bulk properties. However, the changes on the properties were less steady over time.

Comparison between calcium carbonate and ß-tricalcium phosphate as additives of 3D printed scaffolds with polylactic acid matrix.

In this study, polylactic acid (PLA)-based composite scaffolds with calcium carbonate (CaCO3 ) and beta-tricalcium phosphate (ß-TCP) were obtained by 3D printing. These structures were evaluated as potential 3D structures for bone tissue regeneration. Morphological, mechanical, and biological tests were carried out in order to compare the effect of each additive (added in a concentration of 5% w/w) and the combination of both (2.5% w/w of each one), on the PLA matrix. The scaffolds manufactured had a mean pore size between 400-425 µm and a porosity value in the range of 50-60%. According to the results, both additives promoted an increase of the porosity, hydrophilicity, and surface roughness of the scaffolds, leading to a significant improvement of the metabolic activity of human osteoblastic osteosarcoma cells. The best results in terms of cell attachment after 7 days were obtained for the samples containing CaCO3 and ß-TCP particles due to the synergistic effect of both additives, which results in an increase in osteoconductivity and in a microporosity that favours cell adhesion. These scaffolds (PLA:CaCO3 :ß-TCP 95:2.5:2.5) have suitable properties to be further evaluated for bone tissue engineering applications.

Nanofibers and Microfibers for Osteochondral Tissue Engineering.

The use of fibers into scaffolds is a way to mimic natural tissues, in which fibrils are embedded in a matrix. The use of fibers can improve the mechanical properties of the scaffolds and may act as structural support for cell growth. Also, as the morphology of fibrous scaffolds is similar to the natural extracellular matrix, cells cultured on these scaffolds tend to maintain their phenotypic shape. Different materials and techniques can be used to produce micrfibers- and nanofibers for scaffolds manufacturing; cells, in general, adhere and proliferate very well on PCL, chitosan, silk fibroin, and other nanofibers. One of the most important techniques to produce microfibers/nanofibers is electrospinning. Nanofibrous scaffolds are receiving increasing attention in bone tissue engineering, because they are able to offer a favorable microenvironment for cell attachment and growth. Different polymers can be electrospun, i.e., polyester, polyurethane, PLA, PCL, collagen, and silk. Other materials such as bioglass fibers, nanocellulose, and even carbon fiber and fabrics have been used to help increase bioactivity, mechanical properties of the scaffold, and cell proliferation. A compilation of mechanical properties and most common biological tests performed on fibrous scaffolds is included in this chapter. HIGHLIGHTS: The use of microfibers and nanofibers allows for tailoring the scaffold properties. Electrospinning is one of the most important techniques nowadays to produce fibrous scaffolds. Microfibers and nanofibers use in scaffolds is a promising field to improve the behavior of scaffolds in osteochondral applications.