for Guided Bone Regeneration (GBR). The pivotal step on GBR therapy is the insertion of a membrane for support and barrier functions. In Chapter 3, we studied the effect of silica nanoparticles (Si-NPs) incorporation in electrospun poly(ε-caprolactone) (PCL) membranes to improve the mechanical and osteoconductive properties of the membranes. To this end, Si-NPs were firstly synthesized and then suspended in PCL solutions containing a polar solvent (2,2,2-trifluoroethanol) and water with the addition of an anionic surfactant. Nanocomposite membranes were fabricated from the solutions through an electrospinning technique. Morphology, structure and chemical composition, and tensile properties of the membranes were analyzed. Membrane stability was determined by visual examination of the membranes after immersion in phosphate buffered saline (PBS). The effect of the materials on osteoblastic differentiation was evaluated by in vitro culture of the membranes with MC3T3-E1 osteoblastic cells. The results indicated that Si-NPs were successfully incorporated in the interior of the PCL electrospun fibers during the electrospinning process. Tensile modulus significantly increased for composition S50 and tensile strength significantly increased for compositions S25 and S50. Membranes containing Si- NPs showed to be cytocompatible. The results obtained demonstrate that the Si- NPs were homogeneously incorporated within the electrospun fibers, resulting in an improvement of the tensile properties of the prepared materials.
Can a thermal treatment lead to a more sustained release of the simvastatin incorporated in PLLA electrospun membranes, enabling their use as GBR electrospun membranes?
The incorporation of osteostimulatory compounds can improve the biofunctionality of electrospun membranes, making them active players toward bone regeneration. Simvastatin has shown to promote osteogenic differentiation both in vitro and in vivo. However, in most of these systems, simvastatin was quickly released, not matching the pace of bone regeneration. In Chapter 4 we developed poly(L-lactic acid) (PLLA) membranes containing simvastatin (SV) with an increased drug release rate, compatible with GBR applications. To this end, SV was mixed with PLLA and electrospun. The membranes were subjected to a thermal treatment to increase the crystallinity of PLLA. Morphological, structural and chemical properties of the electrospun membranes were characterized. The effect of the thermal treatment on the release profile of SV was evaluated by near physiological release experiments at 37 °C. The osteostimulatory potential was determined by in vitro culture of the membranes with rat bone marrow stromal cells (rBMSCs). The results confirmed that thermal treatment led to an increased polymer crystallinity and a more sustained release of SV. In vitro assays demonstrated cellular proliferation over time for all the membranes and a significant increase in ALP-activity expression for the cells
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cultured with the membranes containing SV subjected to thermal treatment. Can a layer-by-layer (LbL) strategy, based on the deposition of PAA and PLL, be used to functionalize PLLA electrospun fibers, facilitating the incorporation of small molecules?
In Chapter 5, post-spinning functionalization of PLLA electrospun membranes by a Layer-by-Layer (LbL) strategy was investigated. Electrospun poly(L-lactic acid) (PLLA) membranes are widely used for biomedical applications. A major drawback for such systems is PLLA lacking biological functionality, leading to the need of developing functionalization strategies that alter the physicochemical properties of PLLA, and hence enable the incorporation of reactive molecular groups or macromolecules on its surface. The aim of this study was to functionalize the fiber surface of electrospun PLLA membranes by LbL, creating a system that potentially finds application as a highly biocompatible substrate with the capacity to function as a reservoir for small molecules (e.g. pharmaceutical compounds). To this end, biocompatible polyelectrolytes, poly(L-lysine) (PLL) and poly(acrylic acid) (PAA), were used to functionalize PLLA electrospun membranes. Morphological and elemental examination of the membranes was performed. LbL functionalization was achieved, with a proportional increase in the amount of polyelectrolytes deposited with the number of immersion steps. Furthermore, two model drugs with a different surface charge at physiological pH (rhodamine B and clindamycin) were incorporated and their loading efficiency and release evaluated. Both drugs were incorporated, albeit with low loading efficiency. Rhodamine B was released after 8 hours of incubation in phosphate buffered saline (PBS) at 37 °C, while clindamycin release occurred up to 48 hours of incubation. Our data demonstrated the feasibility of the approach, but more studies are needed to improve drug loading efficiency and control drug release.
Can functionalized microcylinders be developed by a top-down modification of electrospun fibers?
Electrospinning is a versatile technique, which can be used not only to produce scaffolds and membranes but also for the development of new structures and components for composite biomaterials. In Chapter 6, we described a top-down approach for the development of new functionalized micrometric structures based on electrospun PLLA fibers, which can be used for the development of drug delivery systems or as building blocks for new biomaterials. A wide range of particles has been developed for different applications in drug delivery, tissue engineering, or regenerative medicine. In contrast to traditional spherical particles, non-spherical (e.g. cylindrical) particles possess several structural and morphological advantages that make them attractive for specific applications. In
Chapter 6, electrospun fibers were processed into micron-sized cylinders (i.e.
microcylinders) with a high specific surface area and with or without surface porosity. To obtain these microcylinders, PLLA solutions were subjected to electrospinning, followed by an aminolysis-based chemical scission procedure. The morphology, structure, and chemical composition of the microcylinders were then characterized. Specific surface area and surface porosity of the microcylinders were controlled by the volatility of the solvents and their length was dependent on the duration of the aminolysis reaction. During aminolysis, the microcylinders became functionalized with amine groups, enabling potential further modifications by grafting with compounds containing desired chemical groups, e.g. carboxyl, carbonyl or hydroxyl functional groups. Additionally, the microcylinders showed in vitro biocompatible properties related to cell viability. These data demonstrate that PLLA microcylinders with a high specific surface area, optional surface porosity, amine-based functional handles granting additional functionalization, and cytocompatible properties are candidate materials for future biomedical applications.
Does the incorporation of PLLA microcylinders in calcium phosphate cements (CPCs) have a positive effect on their mechanical properties?