Suchergebnisse

Tendon tissues have limited healing capacity. The incidence of tendon injuries and the unsatisfactory functional outcomes of tendon repair are driving the search for alternative therapeutic approaches envisioning tendon regeneration. Cellular therapies aim at delivering adequate, regeneration-competent cell types to the injured tendon and toward ultimately promoting its reconstruction and recovery of functionality. Mesenchymal stem cells (MSCs) either obtained from tendons or from non-tendon sources, like bone marrow (BM-MSCs) or adipose tissue (ASCs), have been receiving increasing attention over the years toward enhancing tendon healing. Evidences from in vitro and in vivo studies suggest MSCs can contribute to accelerate and improve the quality of tendon healing. Nonetheless, the exact mechanisms underlying these repair events are yet to be fully elucidated. This review provides an overview of the main challenges in the field of cell-based regenerative therapies, discussing the role of MSCs in boosting tendon regeneration, particularly through their capacity to enhance the tenogenic properties of tendon resident cells. ; The authors acknowledge the financial support from the European Union Framework Programme for Research and Innovation HORIZON2020, under the TEAMING Grant agreement No 739572-The Discoveries CTR, the ERC Grant CoG MagTendon nr 772817, and the Achilles Grant nr 810850, FCT-Fundacao para a Ciencia e a Tecnologia for the PhD grant of IC (PD/BD/128088/2016); and the Project NORTE-01-0145-FEDER-000021: "Accelerating tissue engineering and personalized medicine discoveries by the integration of key enabling nanotechnologies, marine-derived biomaterials and stem cells", supported by Norte Portugal Regional Operational Programme (NORTE 2020), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF). ...

The adipose tissue was considered a reserve of energy until the '80s, when it was found that this tissue was involved in the metabolism of sex steroids such as estrogens. From then on, the importance attributed to this tissue radically changed as it was then considered an active organ, involved in important functions of the human body. In 2001, for the first time, the existence of stem cells within this tissue was reported, and since then, this tissue has been gaining an increased importance as a stem cell source for a wide range of potential applications in cell therapies and=or tissue engineering and regenerative medicine strategies, mainly due to its wide availability and easy access. This manuscript provides an overview on adipose stem cells (i.e., adipose tissue–derived stem cells, ASCs) considering the tissue of origin, the niche of the ASCs, and their phenotype in all aspects. In this paper it is also discussed the markers that have been used for the characterization of these cells, their differentiation properties, and their immunological reactivity, reporting studies from 2001 until this date. The ASCs are also compared with bone marrow stem cells (BMSCs), until now considered as the gold standard source of stem cells, underlining the common characteristics and the differences between the stem cells obtained from these two sources, as well as the advantages and disadvantages of their potential use in different applications. Finally, this review will also focus on the potential application of ASCs in tissue engineering applications, particularly in the regeneration of bone and cartilage, commenting on the progress of this approach and future trends of the field. ; T. Rada thanks the European Marie Curie EST Project (Alea Jacta Est) for the Ph. D. fellowship. The authors acknowledge the Portuguese Foundation for Science and Technology (FCT) for the partial financial support through funds from POCTI and/or FEDER programs and to the European Union funded STREP Project HIPPOCRATES (NMP3-CT-2003-505758) and ...

This manuscript provides an overview of the in vitro and in vivo studies reported in the literature focusing on seaweed polysaccharides based hydrogels that have been proposed for applications in regenerative medicine, particularly, in the field of cartilage tissue engineering. For a better understanding of the main requisites for these specific applications, the main aspects of the native cartilage structure, as well as recognized diseases that affect this tissue are briefly described. Current available treatments are also presented to emphasize the need for alternative techniques. The following part of this review is centered on the description of the general characteristics of algae polysaccharides, as well as relevant properties required for designing hydrogels for cartilage tissue engineering purposes. An in-depth overview of the most well known seaweed polysaccharide, namely agarose, alginate, carrageenan and ulvan biopolymeric gels, that have been proposed for engineering cartilage is also provided. Finally, this review describes and summarizes the translational aspect for the clinical application of alternative systems emphasizing the importance of cryopreservation and the commercial products currently available for cartilage treatment. ; Authors report no declarations of interest. Authors thank the Portuguese Foundation for Science and Technology (FCT) for the PhD fellowship of Elena G. Popa (SFRH/BD/64070/2009) and research project (MIT/ECE/0047/2009). The research leading to these results has received funding from the European Union's Seventh Framework Programme (FP7/2007-2013) under grant agreement no ...

Musculoskeletal diseases are increasing the prevalence of physical disability worldwide. Within the body, musculoskeletal soft and hard tissues integrate through specific multitissue transitions, allowing for body movements. Owing to their unique compositional and structural gradients, injuries challenge the native interfaces and tissue regeneration is unlikely to occur. Tissue engineering strategies are emerging to emulate the physiological environment of soft-to-hard tissue interfaces. Advances in biomaterial design enable control over biophysical parameters, but biomaterials alone are not sufficient to provide adequate support and guide transplanted cells. Therefore, biological, biophysical, and biochemical tools can be integrated into a multifactorial toolbox, steering prospective advances toward engineering clinically relevant soft-to-hard tissue interfaces. ; The authors acknowledge the financial support from the European Union Framework Programme for Research and Innovation HORIZON2020 (TEAMING Grant agreement, No 739572 - The Discoveries CTR), the ERC Grant CoG MagTendon (nr 772817), Fundação para a Ciência e a Tecnologia (FCT) for the PhD grant of I.C. (PD/BD/128088/2016), and the Project NORTE-01-0145-FEDER-000021 through the European Regional Development Fund ...

Magnetic nanoparticles (MNPs) are attractive tools to overcome limitations of current regenerative medicine strategies, demonstrating potential to integrate therapeutic and diagnostic functionalities in highly controlled systems. In traditional tissue engineering (TE) approaches, the MNPs association with stem cells in a three-dimensional (3D) template offers the possibility to achieve a mechano-magnetic responsive system, enabling remote control actuation. Herein, we propose to study the role of MNPs integrated in κ-carrageenan (κC) hydrogels in the cellular response of human adipose-derived stem cells (hASCs) aiming at cartilage TE applications. The results indicated that the concentration of MNPs in the κC hydrogels influences cellular behavior, tuning a positive effect on cell viability, cell content and metabolic activity of hASCs, with the most promising outcomes found in 5% MNP-κC matrices. Although hASCs laden in MNPs-free- and MNPs-κC hydrogels showed similar metabolic and proliferation levels, MNPs κC hydrogels under magnetic actuation evidenced an instructive effect on hASCs, at a gene expression level, towards chondrogenic phenotype even in basic medium cultures. Therefore, the MNPs-based systems developed in this study may contribute to advanced strategies towards cartilage-like engineered substitutes. ; The authors wish to acknowledge the financial support of the Portuguese Foundation for Science and Technology for the PhD grant(SFRH/BD/64070/2009) and Post-Doc grant (SFRH/BPD/91072/2012). In addition, the research leading to these results has received funding from the European Union's Seventh Framework Programme (FP7/2007-2013) under grant agreement n REGPOT-CT2012-316331-POLARIS and under the projects MIT/ECE/0047/2009 and RL3-TECT-NORTE-07-0124-FEDER-000020 co-financed by ON.2 (NSRF), through ...

Cells microencapsulated in biocompatible semi-permeable polymeric membranes are effective as cell delivery systems while protecting the host against immune responses. In this study, cell encapsulation membranes were prepared based on carrageenan and alginate, two natural cationic polymers. Different formulations/conditions were explored to optimize the microcapsules which were characterized with respect to their morphology, mechanical stability, and cytotoxicity. Spherical-shaped microcapsules were obtained from all the polymeric systems. The iota-carrageenan/sodium alginate microcapsules exhibited the best stability and permeability, and therefore, these were selected for the cell encapsulation. These capsules provided an environment that supported cell proliferation and have the potential for tissue engineering as well as other cell-based therapy applications. ; One of the authors (SML) acknowledges the support of the Programme Alssan-the European Union Programme of High Level Scholarships for Latin America (scholarship no. E04M041362CO). This work was partially supported by the European STREP HIPPOCRATES (NMP3-CT-2003-505758) and by the Fundacao para a Ciencia e Tecnologia (project PTDC/QUI/68804/2006) and carried out under the scope of European NoE EXPERTISSUES ...

Tendon-to-bone interfaces exhibit a hierarchical multitissue transition. To replicate the progression from mineralized to nonmineralized tissue, a novel 3D fibrous scaffold is fabricated with spatial control over mineral distribution and cellular alignment. For this purpose, wet-spun continuous microfibers are produced using polycaprolactone (PCL)/ gelatin and PCL/gelatin/hydroxyapatite nano-to-microparticles (HAp). Higher extrusion rates result in aligned PCL/gelatin microfibers while, in the case of PCL/gelatin/HAp, the presence of minerals leads to a less organized structure. Biological performance using human adipose-derived stem cells (hASCs) demonstrates that topography of PCL/gelatin microfibers can induce cytoskeleton elongation, resembling native tenogenic organization. Matrix mineralization on PCL/gelatin/HAp wet-spun composite microfibers suggests the production of an osteogenic-like matrix, without external addition of osteogenic medium supplementation. As proof of concept, a 3D gradient structure is produced by assembling PCL/gelatin and PCL/gelatin/HAp microfibers, resulting in a fibrous scaffold with a continuous topographical and compositional gradient. Overall, the feasibility of wet-spinning for the generation of continuously aligned and textured microfibers is demonstrated, which can be further assembled into more complex 3D gradient structures to mimic characteristic features of tendon-to-bone interfaces. ; The authors thank to Hospital da Prelada (Porto, Portugal) for providing lipoaspirate tissue (Plastic Surgery Department) samples; Dr. Rui Domingues and Dr. Pedro Babo for the technical help with confocal microscopy and micro-CT analysis. The authors acknowledge the financial support from the European Union Framework Programme for Research and Innovation HORIZON2020, under the TEAMING Grant agreement No 739572 – The Discoveries CTR, the ERC Grant CoG MagTendon nr 772817, FCT-Fundação para a Ciência e a Tecnologia for the PhD grant of IC (PD/BD/128088/2016); and the Project ...

"Version of Record online: 29 OCT 2015" ; Bone formation relies on complex processes that require well-orchestrated interactions between several cell types, such as bone-forming cells (osteoblasts, OBs) and endothelial cells (ECs). Their co-culture has been proved relevant to mimicking specific features of the bone niche. Here we propose the co-culture of microvascular-like ECs and pre-OBs, both derived from the SSEA-4+ cell subpopulation from the stromal vascular fraction of human adipose tissue (SSEA-4+ hASCs), to define the conditions in which cells synergistically communicate to support the full differentiation of pre-OBs and maintenance of the EC phenotype. Co-cultures of different ratios of the two cell types were established and maintained for up to 21 days in standard endothelial maintenance (ENDO) and osteogenic differentiation (OST) media, as well as in a mixture of these (MIX). The osteogenic maturation of pre-OBs (ALP activity, OPN and OCN expression, calcium deposition), the evolution of EC numbers (CD31+ cells) and maintenance of the endothelial phenotype (CD31 and vWF expression, LDL uptake) were assessed throughout the culture time as a function of cell ratio and culture media. The results obtained demonstrate that EC number has a significant effect on the osteogenic differentiation of pre-OBs, depending on the medium used. While in ENDO medium the osteogenic differentiation was not observed, in the OST and MIX media it was attained at similar levels, except for the co-culture with a higher number of ECs in MIX medium. These findings demonstrate that the use of SSEA-4+ hASCs as a single-cell source is promising to attain 3D bone-like models with the potential to promote vascularized bone tissue regeneration. ; The authors are grateful to the Portuguese Foundation for Science and Technology (FCT) for personal grants SFRH/BD/ 42968/2008 through the MIT–Portugal Programme (SMM). The research leading to these results has received funding from the MIT/ECE/0047/2009 project, the European Union Seventh ...

Great efforts have been made to introduce growth factors (GFs) onto 2D/3D constructs in order to control cell behavior. Platelet Lysate (PL) presents itself as a cost-effective source of multiple GFs and other proteins. The instruction given by a construct-PL combination will depend on how its instructive cues are presented to the cells. The content, stability and conformation of the GFs affect their instruction. Strategies for a controlled incorporation of PL are needed. Herein, PL was incorporated into nanocoatings by layer-by-layer assembling with polysaccharides presenting different sulfation degrees (SD) and charges. Heparin and several marine polysaccharides were tested to evaluate their PL and GF incorporation capability. The consequent effects of those multilayers on human adipose derived stem cells (hASCs) were assessed in short-term cultures. Both nature of the polysaccharide and SD were important properties that influenced the adsorption of PL, vascular endothelial growth factor (VEGF), fibroblast growth factor b (FGFb) and platelet derived growth factor (PDGF). The sulfated polysaccharides-PL multilayers showed to be efficient in the promotion of morphological changes, serum-free adhesion and proliferation of high passage hASCs (P>5). These biomimetic multilayers promise to be versatile platforms to fabricate instructive devices allowing a tunable incorporation of PL. ; Portuguese Foundation for Science and Technology is gratefully acknowledged for fellowships of Sara M. Oliveira. (SFRH/BD/70107/2010).r The research leading to these results has received funding from the European Union's Seventh Framework Programme (FP7/2007-2013) under grant agreement no REGPOT-CT2012-316331-POLARIS and FP7-KBBE-2010-4-266033 - SPECIAL. This work was also supported by the European Research Council grant agreement ERC-2012-ADG 20120216-321266 for the project ComplexiTE. The authors acknowledge Rogerio P. Pirraco for the Flow cytometry ...

Surface properties play a vita role in the functioning of a biomaterial. Cellular adherence and growth onto biomaterials can be enhanced in biomaterial modifications of their surface. In this work, the cell behaviour on chitosan membranes modified by argon and nitron-plasma treatments was investigated. Characterization of the membranes was performed using atomic force microscopy, contact angle measurements, and X-ray photoelectron spectroscopy. Cytotoxicity assessment and direct contact assay were carried out for untreated and treated chitosan membranes using L929 fibroblast-like cells. Cell morphology and cell viability were assessed to evaluate the cell attachment and proliferation. Changes in terms of roughness, surface chemistry, and hydrophilicity/hydrophobic balance of chitosan-modified membranes were observed. Regarding cell studies, the findings revealed that the extracts of all membranes do not induce cytotoxicity effects. Moreover, the in vitro assays evidenced an improvement of the L929 adhesion, and attachment when compared to untreated chitosan membranes. overall, the data obtained clearly demonstrated that plasma treatments constitute an effective way of improving the biocompatibility of chitosan membranes towards to their use in biomedical applications. ; S. M. Luna acknowledges the support of the Programme Alssan - The European Union Programme of High Level Scholarships for Latin America (scholarship No E04M041362CO) and Silva SS acknowledge the support of the Portuguese Foundation for Science and Technology, (SFRH/BPD/45307/2008). This work was also partially supported by the European Union-funded - STREP project HIPPOCRATES (NMP3-CT-2003-505758) and was carried out under the scope of European NoE EXPERTISSUES ...

Molecular imprinting (MI) has been explored as an increasingly viable tool for molecular recognition in various fields. However, imprinting of biologically relevant molecules like proteins is severely hampered by several problems. Inspired by natural antibodies, the use of epitopes as imprinting templates has been explored to circumvent those limitations, offering lower costs and greater versatility. Here, we review the latest innovations in this technology, as well as different applications where MI polymers (MIPs) have been used to target biomolecules of interest. We discuss the several steps in MI, from the choice of epitope and functional monomers to the different production methods and possible applications. We also critically explore how MIP performance can be assessed by various parameters. Last, we present perspectives on future breakthroughs and advances, offering insights into how MI techniques can be expanded to new fields such as tissue engineering. ; This work was supported by Project NORTE-01-0145-FEDER-000021 supported by the Norte Portugal Regional Operational Program (NORTE 2020), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF); by the European Union Framework Program for Research and Innovation HORIZON 2020, under the Twinning grant agreement no. 810850–Achilles, European Research Council grant agreement no. 772817; and by FCT/MCTES (Fundação para a Ciência e a Tecnologia/Ministério da Ciência, Tecnologia, e Ensino Superior) through PhD grant PD/BD/143039/2018 for S.P.B.T., financed through the Doctoral Program in Advanced Therapies for Health (PATH) (FSE/POCH/ PD/169/2013), project PTDC/NAN-MAT/30595/2017, and individual contract 2020.03410. CEECIND for R.M.A.D. N.A.P. acknowledges support from the Cockrell Family Chair Foundation; the Institute for Biomaterials, Drug Delivery, and Regenerative Medicine; and the UT-Portugal Collaborative Research ...

Most tissues of the human body are characterized by highly anisotropic physical properties and biological organization. Hydrogels have been proposed as scaffolding materials to construct artificial tissues due to their water-rich composition, biocompatibility and tunable properties. However, unmodified hydrogels are typically composed of randomly oriented polymer networks, resulting in homogeneous structures with isotropic properties different from those observed in biological systems. Magnetic materials have been proposed as potential agents to provide hydrogels with the anisotropy required for their use on tissue engineering. Moreover, the intrinsic properties of magnetic nanoparticles enable their use as magnetomechanic remote actuators to control the behavior of the cells encapsulated within the hydrogels under the application of external magnetic fields. In this review, we combine a detailed summary of the main strategies to prepare magnetic nanoparticles showing controlled properties with an analysis of the different approaches available to their incorporation into hydrogels. The application of magnetically-responsive nanocomposite hydrogels in the engineering of different tissues is also reviewed. ; We acknowledge European Union's Horizon 2020 Research and Innovation Programme—European Research Council Consolidator Grant MagTendon (Agreement No. 772817) and Project Achilles (Agreement No. 810850) and Fundação para a Ciência e Tecnologia (FCT) for Project SmarTendon-PTDC/NAN-MAT/30595/2017 and Project MagTT-PTDC/CTM-CTM/29930/2017, from NORTE-01-0145-FEDER-000021 supported by Norte Portugal Regional Operational Programme (NORTE 2020). We also thank Agencia Estatal de Investigación by Project MAT2016-80266-R and Xunta de Galicia (Grupo de Referencia Competitiva ED431C 2018/26 and ED431C 2018/09; Agrupación Estratégica en Materiales ED431E 2018/08). A.P. is grateful to Xunta de Galicia for his postdoctoral fellowship ED481B2019/025. ERDF funds are also ...

Platelet-derived products (PDPs) have gained popularity, mainly due to their high concentrations of bioactive molecules such as growth factors and cytokines, which play important roles in tissue healing and regeneration. PDPs are obtained through minimally invasive procedures and their therapeutic effect has been widely recognized. In veterinary medicine, however, the lack of standard protocols to generate PDPs is a major hurdle for assessing the clinical relevance of PDP-based therapies and for their widespread usage. The aim of this review is to analyze the technical and scientific specificities of PDPs in terms of preparation methodologies, classification categorization, nomenclature, and biological proprieties to advance their future biotechnological potential in veterinary contexts. ; This work was supported by Fundação para a Ciência e Tecnologia, from Portugal SFRH/BD/118871/2016 PhD grant. The authors acknowledge the financial support from the European Union Framework Program for Research and Innovation HORIZON 2020, under the FOOD4CELLS project (PTDC/CTM-BIO/4706/2014 - POCI-01-0145-FEDER ...

First published: 1 September 2013 ; The inefficacy of the currently used therapies in achieving the regeneration ad integrum of the periodontium stimulates the search for alternative approaches, such as tissue-engineering strategies. Therefore, the core objective of this study was to develop a biodegradable double-layer scaffold for periodontal tissue engineering. The design philosophy was based on a double-layered construct obtained from a blend of starch and poly-ε-caprolactone (30:70 wt%; SPCL). A SPCL fibre mesh functionalized with silanol groups to promote osteogenesis was combined with a SPCL solvent casting membrane aiming at acting as a barrier against the migration of gingival epithelium into the periodontal defect. Each layer of the double-layer scaffolds was characterized in terms of morphology, surface chemical composition, degradation behaviour and mechanical properties. Moreover, the behaviour of seeded/cultured canine adipose-derived stem cells (cASCs) was assessed. In general, the developed double-layered scaffolds demonstrated adequate degradation and mechanical behaviour for the target application. Furthermore, the biological assays revealed that both layers of the scaffold allow adhesion and proliferation of the seeded undifferentiated cASCs, and the incorporation of silanol groups into the fibre-mesh layer enhance the expression of a typical osteogenic marker. This study allowed an innovative construct to be developed, combining a three-dimensional (3D) scaffold with osteoconductive properties and with potential to assist periodontal regeneration, carrying new possible solutions to current clinical needs . ; The research leading to these results has received funding from the European Union's Seventh Framework Programme (FP7/2007-2013; under Grant Agreement No. REGPOT-CT2012-316331-POLARIS) and from the Portuguese Foundation for Science and Technology (FCT; Grant No, MIT/ECE/0047/2009). Joao Requicha acknowledges the FCT for his PhD scholarship ( Grant No. ...

How to surpass invitro stem cell differentiation, reducing cell manipulation, and lead the in situ regeneration process after transplantation, remains to be unraveled in bone tissue engineering (bTE). Recently, we showed that the combination of human bone marrow stromal cells with bioactive silicate nanoplatelets (sNPs) promotes the osteogenic differentiation without the use of standard osteogenic inductors. Even more, using SSEA-4(+) cell-subpopulations (SSEA-4(+)hASCs) residing within the adipose tissue, as a single-cellular source to obtain relevant cell types for bone regeneration, was also proposed. Herein, sNPs were used to promote the osteogenic differentiation of SSEA-4(+)hASCs. The interactions between SSEA-4(+)hASCs and sNPs, namely the internalization pathway and effect on cells osteogenic differentiation, were evaluated. SNPs below 100μg/mL showed high cytocompatibility and fast internalization via clathrin-mediated pathway. SNPs triggered an overexpression of osteogenic-related markers (RUNX2, osteopontin, osteocalcin) accompanied by increased alkaline phosphatase activity and deposition of a predominantly collagen-type I matrix. Consequently, a robust matrix mineralization was achieved, covering >90% of the culturing surface area. Overall, we demonstrated the high osteogenic differentiation potential of SSEA-4(+)hASCs, further enhanced by the addition of sNPs in a dose dependent manner. This strategy endorses the combination of an adipose-derived cell-subpopulation with inorganic compounds to achieve bone matrix-analogs with clinical relevance. ; Authors thank the Portuguese Foundation for Science and Technology (FCT) for the personal grant SFRH/BD/42968/2008 through the MIT-Portugal Program (SMM). The research leading to these results has received funding from the MIT/ECE/0047/2009 project and the European Union's Seventh Framework Programme (FP7/2007-2013) under grant agreement n degrees REGPOT-CT2012-316331-POLARIS and MIT/ECE/0047/2009 ...