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Los animales han sido utilizados desde hace tiempo como sujetos experimentales para apoyar el progreso científico y los avances de la medicina. A día de hoy, diversos protocolos incluyendo el uso de animales todavía están en vigor en múltiples campos científicos y aplicaciones, como por ejemplo el de la seguridad de los dispositivos médicos. El objetivo de dichos estudios es determinar si un dispositivo medico es seguro para su uso humano o si supone un riesgo para la salud del paciente. Las pruebas en animales para este propósito están motivadas por requisitos regulatorios, estipulando que el uso de este tipo de test es necesario y justificándolo en un contexto de gestión de riesgos. Los modelos animales ya no se usan de manera sistemática como en el pasado, pero siguen siendo una piedra angular en el proceso de toma de decisión delicado, relacionado con el marketing de los dispositivos médicos. Sin embargo, las pruebas en animales tienen limitaciones, como las preocupaciones éticas y la validez de los datos que cuestionan su uso para este fin. Una atención especial está dada a día de hoy a las alternativas libres de animales para respaldar mejor la aplicación del concepto de los 3R (replacement, reduction, refining) en este campo. Desgraciadamente, los métodos muy avanzados y tecnologías en desarrollo encuentran obstáculos importantes (p.ej. validación y estandarización) que impiden y ralentizan su implementación como alternativas aceptadas del punto de vista regulatorio. Para superar estas dificultades, el proceso de aceptación regulatoria de las alternativas se podría optimizar. Este se puede alcanzar por ejemplo mejorando la cooperación, la coordinación y la comunicación entre los diferentes actores, i.e. investigadores, organismos regulatorios certificadores e industrias. Generalmente, una atención especial se debe prestar al compromiso y a un esfuerzo común para mejorar la eficacia de este proceso, y para establecer un marco regulatorio de comprobación que no dependa más de los modelos animales. Animals have been used since a very long time as experimental subjects to support scientific progress and medical advances. Currently, various testing procedures implying animals are still being conducted within a broad range of scientific fields and applications, as for example the assessment of medical devices safety. The aim of this assessment is to investigate whether a medical device is safe for human use or poses a risk for patients' health. Animal testing for that purpose is especially encouraged by regulatory requirements, which prerequisite however that the use of such testing is deemed necessary and justified in the context of a risk management process. Thus, animal models are no longer systematically used as in the past, but they remain a cornerstone within the delicate decision-making process related to the marketing of medical devices. Though, animal testing has some critical limitations, such as ethical concerns and relevance of animal data, which challenge its further use to that objective. A particular attention is therefore currently given to the development of animal-free alternatives to better support the application of the 3Rs Principle (replacement, reduction, refining) within this field. Unfortunately, the highly-advanced methods and technologies being developed face barriers (e.g. validation and standardization) which hinder and slow down their implementation as regulatory-accepted alternatives. To overcome these difficulties, the process of regulatory acceptance of these alternatives may be optimized. This may be achieved for example by improving the cooperation, coordination and communication between the different stakeholders, i.e. researchers, regulatory approval bodies and industries. Mostly, a special focus may be set on commitment and shared effort to enhance the efficiency of that process and to establish a regulatory testing framework that no longer relies on animal models.

Nadège Sachot,1,2 Agata Roguska,3 Josep Anton Planell,1,2 Malgorzata Lewandowska,3 Elisabeth Engel,1,2,4 Oscar Castaño1,2,5,6 1Biomaterials for Regenerative Therapies, Institute forBioengineering of Catalonia (IBEC), Barcelona,2CIBER en Bioingeniería, Biomateriales yNanomedicina, CIBER-BBN, Zaragoza, Spain; 3Faculty of Materials Science and Engineering, Warsaw University of Technology, Warsaw, Poland; 4Department of Materials Science and Metallurgical Engineering, Universitat Politècnica de Catalunya (UPC), 5Department of Materials Science and Physical Chemistry, Universitat de Barcelona (UB), 6Department of Engineerings: Electronics, Universitat de Barcelona, Barcelona, Spain Abstract: The success of scaffold implantation in acellular tissue engineering approaches relies on the ability of the material to interact properly with the biological environment. This behavior mainly depends on the design of the graft surface and, more precisely, on its capacity to biodegrade in a well-defined manner (nature of ions released, surface-to-volume ratio, dissolution profile of this release, rate of material resorption, and preservation of mechanical properties). The assessment of the biological behavior of temporary templates is therefore very important in tissue engineering, especially for composites, which usually exhibit complicated degradation behavior. Here, blended polylactic acid (PLA) calcium phosphate ORMOGLASS (organically modified glass) nanofibrous mats have been incubated up to 4 weeks in physiological simulated conditions, and their morphological, topographical, and chemical changes have been investigated. The results showed that a significant loss of inorganic phase occurred at the beginning of the immersion and the ORMOGLASS maintained a stable composition afterward throughout the degradation period. As a whole, the nanostructured scaffolds underwent fast and heterogeneous degradation. This study reveals that an angiogenic calcium-rich environment can be achieved through fast-degrading ORMOGLASS/PLA blended fibers, which seems to be an excellent alternative for guided bone regeneration. Keywords: electrospinning, fast degradation, ORMOGLASSES, angiogenesis, nanofibers, calcium release&nbsp

Electrospinning is a method that can be used to efficiently produce scaffolds that mimic the fibrous structure of natural tissue, such as muscle structures or the extracellular matrix of bone. The technique is often used as a way of depositing composites (organic/inorganic materials) to obtain bioactive nanofibers which have the requisite mechanical properties for use in tissue engineering. However, many factors can influence the formation and collection of fibers, including experimental variables such as the parameters of the solution of the electrospun slurry. In this study, we assessed the influence of the polymer concentration, glass content and glass hydrolysis level on the morphology and thickness of fibers produced by electrospinning for a PCL-(Si-Ca-P-2) bioactive ormoglassorganically modified glassblend. Based on previous assays, this combination of materials shows good angiogenic and osteogenic properties, which gives it great potential for use in tissue engineering. The results of our study showed that blend preparation directly affected the features of the resulting fibers, and when the parameters of the blend are precisely controlled, fibers with a regular diameter could be produced fairly easily when 2,2,2-trifluoroethanol was used as a solvent instead of tetrahydrofuran. The diameter of the homogeneous fibers ranged from 360 to 620 nm depending on the experimental conditions used. This demonstrates that experimental optimization of the electrospinning process is crucial in order to obtain a deposit of hybrid nanofibers with a regular shape. (c) 2014 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 103B: 1287-1293, 2015.

Hybrid materials are being extensively investigated with the aim of mimicking the ECM microenvironment to develop effective solutions for bone tissue engineering. However, the common drawbacks of a hybrid material are the lack of interactions between the scaffold's constituents and the masking of its bioactive phase. Conventional hybrids often degrade in a non-homogeneous manner and the biological response is far from optimal. We have developed a novel material with strong interactions between constituents. The bioactive phase is directly exposed on its surface mimicking the structure of the ECM of bone. Here, polylactic acid electrospun fibers have been successfully and reproducibly coated with a bioactive organically modified glass (ormoglass, Si-Ca-P-2 system) covalently. In comparison with the pure polymeric mats, the fibers obtained showed improved hydrophilicity and mechanical properties, bioactive ion release, exhibited a nanoroughness and enabled good cell adhesion and spreading after just one day of culture (rMSCs and rEPCs). The fibers were coated with different ormoglass compositions to tailor their surface properties (roughness, stiffness, and morphology) by modifying the experimental parameters. Knowing that cells modulate their behavior according to the exposed physical and chemical signals, the development of this instructive material is a valuable advance in the design of functional regenerative biomaterials.

The introduction of hybrid materials in regenerative medicine has solved some problems related to the mechanical and bioactive properties of biomaterials. Calcium phosphates and their derivatives have provided the basis for inorganic components, thanks to their good bioactivity, especially in bone regeneration. When mixed with biodegradable polymers, the result is a synergic association that mimics the composition of many tissues of the human body and, additionally, exhibits suitable mechanical properties. Together with the development of nanotechnology and new synthesis methods, hybrids offer a promising option for the development of a third or fourth generation of smart biomaterials and scaffolds to guide the regeneration of natural tissues, with an optimum efficiency/cost ratio. Their potential bioactivity, as well as other valuable features of hybrids, open promising new pathways for their use in bone regeneration and other tissue repair therapies. This review provides a comprehensive overview of the different hybrid organic-inorganic scaffolding biomaterials developed so far for regenerative therapies, especially in bone. It also looks at the potential for research into hybrid materials for other, softer tissues, which is still at an initial stage, but with very promising results.

Electrospinning is a method that can be used to efficiently produce scaffolds that mimic the fibrous structure of natural tissue, such as muscle structures or the extracellular matrix of bone. The technique is often used as a way of depositing composites (organic/inorganic materials) to obtain bioactive nanofibers which have the requisite mechanical properties for use in tissue engineering. However, many factors can influence the formation and collection of fibers, including experimental variables such as the parameters of the solution of the electrospun slurry. In this study, we assessed the influence of the polymer concentration, glass content and glass hydrolysis level on the morphology and thickness of fibers produced by electrospinning for a PCL-(Si-Ca-P2 ) bioactive ormoglass-organically modified glass-blend. Based on previous assays, this combination of materials shows good angiogenic and osteogenic properties, which gives it great potential for use in tissue engineering. The results of our study showed that blend preparation directly affected the features of the resulting fibers, and when the parameters of the blend are precisely controlled, fibers with a regular diameter could be produced fairly easily when 2,2,2-trifluoroethanol was used as a solvent instead of tetrahydrofuran. The diameter of the homogeneous fibers ranged from 360 to 620 nm depending on the experimental conditions used. This demonstrates that experimental optimization of the electrospinning process is crucial in order to obtain a deposit of hybrid nanofibers with a regular shape.

In bone regeneration, silicon-based calcium phosphate glasses (Bioglasses) have been widely used since the 1970s. However, they dissolve very slowly because of their high amount of Si (SiO2 > 45%). Recently, our group has found that calcium ions released by the degradation of glasses in which the job of silicon is done by just 5% of TiO2 are effective angiogenic promoters, because of their stimulation of a cell-membrane calcium sensing receptor (CaSR). Based on this, other focused tests on angiogenesis have found that Bioglasses also have the potential to be angiogenic promoters even with high contents of silicon (80%); however, their slow degradation is still a problem, as the levels of silicon cannot be decreased any lower than 45%. In this work, we propose a new generation of hybrid organically modified glasses, ormoglasses, that enable the levels of silicon to be reduced, therefore speeding up the degradation process. Using electrospinning as a faithful way to mimic the extracellular matrix (ECM), we successfully produced hybrid fibrous mats with three different contents of Si (40, 52, and 70%), and thus three different calcium ion release rates, using an ormoglass-polycaprolactone blend approach. These mats offered a good platform to evaluate different calcium release rates as osteogenic promoters in an in vivo subcutaneous environment. Complementary data were collected to complement Ca2+ release analysis, such as stiffness evaluation by AFM, zeta-potential, morphology evaluation by FESEM, proliferation and differentiation analysis, as well as in vivo subcutaneous implantations. Material and biological characterization suggested that compositions of organic/inorganic hybrid materials with a Si content equivalent to 40%, which were also those that released more calcium, were osteogenic. They also showed a greater ability to form blood vessels. These results suggest that Si-based ormoglasses can be considered an efficient tool for calcium release modulation, which could play a key role in the angiogenic promoting process.

Surface properties of biomaterials play a major role in the governing of cell functionalities. It is well known that mechanical, chemical and nanotopo- graphic cues, for example, influence cell proliferation and differentiation. Here, we present a novel coating protocol to produce hierarchicallyengineered fibrous scaffolds with tailorable surface characteristics, which mimic bone extracellular matrix. Based on the sol–gel method and a succession of surface treatments, hollow electrospun polylactic acid fibres were coated with a silicon–calcium–phosphate bioactive organic–inorganic glass. Compared with pure polymeric fibres that showed a completely smooth surface, the coated fibres exhibited a nanostructured topography and greater roughness. They also showed improved hydrophilic properties and a Young’s modulus sixfold higher than non-coated ones, while remaining fully flexible and easy to handle. Rat mesenchymal stem cells cultured on these fibres showed great cellular spreading and interactions with the material. This protocol can be transferred to other structures and glasses, allowing the fabrication of var- ious materials with well-defined features. This novel approach represents therefore a valuable improvement in the production of artificial matrices able to direct stem cell fate through physical and chemical interactions

Although the impact of composites based on Ti-doped calcium phosphate glasses is low compared with that of bioglass, they have been already shown to possess great potential for bone tissue engineering. Composites made of polylactic acid (PLA) and a microparticle glass of 5TiO 2 –44.5CaO–44.5P 2 O 5 –6Na 2 O (G5) molar ratio have already demonstrated in situ osteo- and angiogenesis-triggering abilities. As many of the hybrid materials currently developed usually promote osteogenesis but still lack the ability to induce vascularization, a G5/PLA combination is a cost-effective option for obtaining new instructive scaffolds. In this study, nanostructured PLA-ORMOGLASS (organically modified glass) fibers were produced by electro- spinning, in order to fabricate extra-cellular matrix (ECM)-like substrates that simultaneously promote bone formation and vascularization. Physical–chemical and surface characterization and tensile tests demon- strated that the obtained scaffolds exhibited homogeneous morphology, higher hydrophilicity and enhanced mechanical properties than pure PLA. In vitro assays with rat mesenchymal stem cells (rMSCs) and rat endothelial progenitor cells (rEPCs) also showed that rMSCs attached and proliferated on the materials influenced by the calcium content in the environment. In vivo assays showed that hybrid composite PLA-ORMOGLASS fibers were able to promote the formation of blood vessels. Thus, these novel fibers are a valid option for the design of functional materials for tissue engineering applications