Browsing by Author "Granja, Pedro L."
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- Cell-instructive pectin hydrogels crosslinked via thiol-norbornene photo-click chemistry for skin tissue engineeringPublication . Pereira, Rúben; Barrias, Cristina C.; Bártolo, Paulo J.; Granja, Pedro L.Cell-instructive hydrogels are attractive for skin repair and regeneration, serving as interactive matrices to promote cell adhesion, cell-driven remodeling and de novo deposition of extracellular matrix compo nents. This paper describes the synthesis and photocrosslinking of cell-instructive pectin hydrogels using cell-degradable peptide crosslinkers and integrin-specific adhesive ligands. Protease-degradable hydro gels obtained by photoinitiated thiol-norbornene click chemistry are rapidly formed in the presence of dermal fibroblasts, exhibit tunable properties and are capable of modulating the behavior of embedded cells, including the cell spreading, hydrogel contraction and secretion of matrix metalloproteases. Keratinocytes seeded on top of fibroblast-loaded hydrogels are able to adhere and form a compact and dense layer of epidermis, mimicking the architecture of the native skin. Thiol-ene photocrosslinkable pec tin hydrogels support the in vitro formation of full-thickness skin and are thus a highly promising plat form for skin tissue engineering applications, including wound healing and in vitro testing mod
- Electrospun polycaprolactone (PCL) degradation: An in vitro and in vivo studyPublication . Dias, Juliana R.; Sousa, Aureliana; Augusto, Ana; Bártolo, Paulo J.; Granja, Pedro L.Polycaprolactone (PCL) is widely used in tissue engineering due to its interesting properties, namely biocompatibility, biodegradability, elastic nature, availability, cost efficacy, and the approval of health authorities such as the American Food and Drug Administration (FDA). The PCL degradation rate is not the most adequate for specific applications such as skin regeneration due to the hydrophobic nature of bulk PCL. However, PCL electrospun fiber meshes, due to their low diameters resulting in high surface area, are expected to exhibit a fast degradation rate. In this work, in vitro and in vivo degradation studies were performed over 90 days to evaluate the potential of electrospun PCL as a wound dressing. Enzymatic and hydrolytic degradation studies in vitro, performed in a static medium, demonstrated the influence of lipase, which promoted a rate of degradation of 97% for PCL meshes. In an in vivo scenario, the degradation was slower, although the samples were not rejected, and were well-integrated in the surrounding tissues inside the subcutaneous pockets specifically created.
- A single-component hydrogel bioink for bioprinting of bioengineered 3D constructs for dermal tissue engineeringPublication . Pereira, Rúben; Sousa, Aureliana; Barrias, Cristina C.; Bártolo, Paulo J.; Granja, Pedro L.Bioprinting is attractive to create cellularized constructs for skin repair. However, the vast majority of bioinks present limitations in the printing of chemically defined 3D constructs with controllable biophysical and biochemical properties. To address this challenge, a single-component hydrogel bioink with a controlled density of cell-adhesive ligands, tuneable mechanical properties and adjustable rheological behaviour is developed for extrusion bioprinting and applied for the biofabrication of 3D dermal constructs. A methacrylate modified pectin bioink is designed to allow the tethering of integrin-binding motifs and the formation of hydrogels by UV photopolymerization and ionic gelation. The rheological behaviour of a low polymer concentration (1.5 wt%) solution is adjusted by ionic crosslinking, yielding a printable bioink that holds the predesigned shape upon deposition for postprinting photocrosslinking. Printed constructs provide a suitablemicroenvironment that supports the deposition of endogenous extracellular matrix, rich in collagen and fibronectin, by entrapped dermal fibroblasts. This approach enables the design of chemically defined and cell-responsive bioinks for tissue engineering applications and particularly for the generation of biomimetic skin constructs.
- Ultrasound sonication prior to electrospinning tailors silk fibroin/PEO membranes for periodontal regenerationPublication . Serôdio, Ricardo; Schickert, Sónia L.; Costa-Pinto, Ana R.; Dias, Juliana R.; Granja, Pedro L.; Yang, Fang; Oliveira, Ana L.In this study, silk fibroin (SF)/poly(ethylene oxide) (PEO) membranes were designed and fabricated by combining ultrasound sonication prior to electrospinning (0 to 20 min) as a strategy to physically control the rheological properties of solutions (10 to 30% w/v PEO) and to improve the spinnability of the system. PEO has proved to be essential as a co-spinning agent to assure good membrane reproducibility and enough flexibility for clinical manipulation. The rheological tests indicated that sonication greatly increased the viscosity of SF/PEO solutions and further enhanced the quality of the produced electrospun fibers with consequent improved mechanical properties in dry and wet conditions. By tuning the viscosity of the solutions using a simple sonication step prior to electrospinning, it was possible to induce water stability in the as-electrospun matrix, as demonstrated by infra-red spectroscopy. This reduced complexity in the process since it was not necessary to concentrate silk prior to electrospinning while avoiding the use of toxic solvents to perform a post-processing stabilization treatment which usually causes dimensional changes to the SF materials. Sonication pre-treatment allowed for minimizing the amount of synthetic polymer used to achieve the desirable mechanical properties (with the modulus ranging between 90 and 170 MPa), while avoiding a further water stabilization treatment. It also had a positive impact in the in vitro cell behavior of human primary periodontal ligament cells (hPDLs), resulting in a marked increase in cell proliferation. The present developed work constitutes a step forward towards simplicity and a better fabrication control of viable electrospun SFbased membranes for periodontal regeneration.