DESIGN AND FABRICATION OF ZONAL CARTILAGE CONSTRUCTS

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Extruded bioreactor perfusion culture supports the chondrogenic differentiation of human mesenchymal stem/stromal cells in 3D porous poly(ɛ-caprolactone) scaffolds

Publication . Silva, João C.; Moura, Carla; Borrecho, Gonçalo; Matos, António P. Alves de; Silva, Cláudia L. da; Cabral, Joaquim M. S.; Bártolo, Paulo J.; Linhardt, Robert J.; Ferreira, Frederico Castelo

Novel bioengineering strategies for the ex vivo fabrication of native-like tissue-engineered cartilage are crucial for the translation of these approaches to clinically manage highly prevalent and debilitating joint diseases. Bioreactors that provide different biophysical stimuli have been used in tissue engineering approaches aimed at enhancing the quality of the cartilage tissue generated. However, such systems are often highly complex, expensive, and not very versatile. In the current study, a novel, cost-effective, and customizable perfusion bioreactor totally fabricated by additive manufacturing (AM) is proposed for the study of the effect of fluid flow on the chondrogenic differentiation of human bone-marrow mesenchymal stem/stromal cells (hBMSCs) in 3D porous poly(ɛ-caprolactone) (PCL) scaffolds. hBMSCs are first seeded and grown on PCL scaffolds and hBMSC–PCL constructs are then transferred to 3D-extruded bioreactors for continuous perfusion culture under chondrogenic inductive conditions. Perfused constructs show similar cell metabolic activity and significantly higher sulfated glycosaminoglycan production (≈1.8-fold) in comparison to their non-perfused counterparts. Importantly, perfusion bioreactor culture significantly promoted the expression of chondrogenic marker genes while downregulating hypertrophy. This work highlights the potential of customizable AM platforms for the development of novel personalized repair strategies and more reliable in vitro models with a wide range of applications.

2020-02Journal article

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Chondrogenic differentiation of mesenchymal stem/stromal cells on 3D porous poly (ε-caprolactone) scaffolds: Effects of material alkaline treatment and chondroitin sulfate supplementation

Publication . Moura, Carla; Silva, João Carlos; Faria, Sofia; Fernandes, Paulo Rui; Silva, Cláudia Lobato da; Cabral, Joaquim Manuel Sampaio; Linhardt, Robert; Bártolo, Paulo Jorge; Ferreira, Frederico Castelo

Cartilage defects resultant from trauma or degenerative diseases (e.g., osteoarthritis) can potentially be repaired using tissue engineering (TE) strategies combining progenitor cells, biomaterial scaffolds and bio physical/chemical cues. This work examines promoting chondrogenic differentiation of human bone marrow mesenchymal stem/stromal cells (BMMSCs) by combining the effects of modified poly (ε-caprolactone) (PCL) scaffolds hydrophilicity and chondroitin sulfate (CS) supplementation in a hypoxic 5% oxygen atmosphere. 3D extruded PCL scaffolds, characterized by mCT, featured a 21 mmL1 surface area to volume ratio, 390 mm pore size and approximately 100% pore interconnectivity. Scaffold immersion in sodium hydroxide solutions for different periods of time had major effects in scaffold surface morphology, wettability and mechanical properties, but without improvements on cell adhesion. In-situ chondrogenic differentiation of BM-MSC seeded in 3D-extruded PCL scaffolds resulted in higher cell populations and ECM deposition along all scaffold structure, when chondrogenesis was preceded by an expansion phase. Additionally, CS supplementation during BM-MSC expansion was crucial to enhance aggrecan gene expression, known as a hallmark of chondrogenesis. Overall, this study presents an approach to tailor the wettability and mechanical properties of PCL scaffolds and supports the use of CSsupplementation as a biochemical cue in integrated TE strategies for cartilage regeneration.

2020-06Journal article

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Comparison of Three-dimensional Extruded Poly (ɛ-Caprolactone) and Polylactic acid Scaffolds with Pore size Variation

Publication . Monteiro de Moura, Carla Sofia; Ferreira, Frederico Castelo; Bártolo, Paulo Jorge

Additive manufacturing (AM) has become a prominent approach among the scientific community for the production of three-dimensional (3D) matrices able to support tissue engineering approaches, promoting cell adhesion, proliferation and organization aiming to repair different tissues, such as bone or cartilage. In this study we used an extrusion-based technique for the production of poly (ɛ-caprolactone) (PCL) and polylactic acid (PLA) scaffolds and performed a side-by-side scaffold characteristics comparison. Using this technique we were able to create fully 3D interconnected porous scaffolds with pore size variations ranging from 190 μm to 390 μm with both materials. These scaffolds were assessed for stiffness, wettability and cell adhesion using mesenchymal stem/stromal cells (MSC). Comparisons between these two materials were made. The compressive modulus obtained is on the same order of magnitude for both materials. However, PCL presents a statistically significant higher compressive modulus. Results confirmed that PCL is a more hydrophobic material, so it presents a lower wettability when compared to PLA. Interestingly cell adhesion is similar for PLA and PCL, therefore selection between these two materials for the use of this versatile platform can be defined according with biodegradability aimed.

2016Journal article

Open access