Current management for diabetes has stimulated the development of versatile 3D-based hydrogels as in vitro platforms for insulin release and as support for the encapsulation of pancreatic cells and islets of Langerhans. This work aimed to create agarose/fucoidan hydrogels to encapsulate pancreatic cells as a potential biomaterial for diabetes therapeutics. The hydrogels were produced by combining fucoidan (Fu) and agarose (Aga), marine polysaccharides derived from the cell wall of brown and red seaweeds, respectively, and a thermal gelation process. The agarose/fucoidan (AgaFu) blended hydrogels were obtained by dissolving Aga in 3 or 5 wt % Fu aqueous solutions to obtain different proportions (4:10; 5:10, and 7:10 wt). The rheological tests on hydrogels revealed a non-Newtonian and viscoelastic behavior, while the characterization confirmed the presence of the two polymers in the structure of the hydrogels. In addition, the mechanical behavior showed that increasing Aga concentrations resulted in hydrogels with higher Young's modulus. Further, the ability of the developed materials to sustain the viability of human pancreatic cells was assessed by encapsulation of the 1.1B4HP cell line for up to 7 days. The biological assessment of the hydrogels revealed that cultured pancreatic beta cells tended to self-organize and form pseudo-islets during the period studied.

Microencapsulating pancreatic islets in immunoprotective alginate hydrogels is a promising strategy for treatment of type 1 diabetes. However, this strategy is limited by inflammation and hypoxia mediated oxidative stress, due to encapsulation and the hydrogel itself, leading to impaired insulin secretion and limited short and long term cell survival. Herein, the antioxidant effect of fucoidan, an algae derived polysaccharide, on beta cells, and its positive effects on encapsulated beta cell viability and function is presented. Fucoidan from Fucus vesiculosus (FF) exhibits a high total antioxidant capacity, and free radical scavenging activity, and is able to significantly alleviate intracellular oxidative stress in rat insolinoma beta cells (INS1E). In addition, FF significantly increases insulin secretion in a dose‐ and time‐dependent manner. When FF is incorporated in ultrapure alginate used for microencapsulation of primary rat islets, both viability and glucose responsiveness of rat islets in these socalled Fucogel microcapsules (Fucocaps) are found to be significantly higher compared to islets encapsulated in alginate alone. Similar results are obtained with INS1E pseudoislets and neonatal pig islets. Fucocaps can provide a redox‐modulatory niche and an immune barrier for islets and beta cells in the same time leading to significantly improved survival and endocrine function by mitigating oxidative stress.

Mankind has used natural resources for many different applications, from food and clothes to drugs, but only recently is giving the due attention to the need for a responsible management of these resources toward sustainability. One of the approaches is to take the most out of its resources, addressing the several components of natural resources, which can be used in many industrial fields. In this regard, many natural compounds have been studied for the evaluation of biological activities with relevance for human health and well-being. Among the explored resources, the survey of marine organisms has been growing, particularly regarding microbiota and seaweeds or macroalgae. Macroalgae are macroscopic algae that are usually found on rocky shores, exhibiting a great diversity of colors, shapes and sizes. They are divided in three large groups, essentially based on their color: green macroalgae, brown macroalgae and red macroalgae. In recent decades, seaweeds have been widely used as a sustainable source of sulfated polysaccharides, which exhibiting diverse chemical and biological properties aim to address the demands of Tissue Engineering and Regenerative Medicine (TERM), as well as of other areas, such as cosmetics and food. This manuscript provides an overview of the paradigmatic examples of this type of biopolymer that can be obtained from the different groups of algae, detailing the chemical structure, general biological characteristics and most revealing properties.

There is a current lack of fully efficient therapies for diabetes mellitus, a chronic disease where the metabolism of blood glucose is severely hindered by a deficit in insulin or cell resistance to this hormone. Therefore, it is crucial to develop new therapeutic strategies to treat this disease, including devices for the controlled delivery of insulin or encapsulation of insulin-producing cells. In this work, fucoidan (Fu) - a marine sulfated polysaccharide exhibiting relevant properties on reducing blood glucose and antioxidant and anti-inflammatory effects - was used for the development of versatile carriers envisaging diabetes advanced therapies. Fu was functionalized by methacrylation (MFu) using 8% and 12% (v/v) of methacrylic anhydride and further photocrosslinked using visible light in the presence of triethanolamine and eosin-y to produce hydrogel particles. Degree of methacrylation varied between 2.78 and 6.50, as determined by (HNMR)-H-1, and the produced particles have an average diameter ranging from 0.63 to 1.3 mm (dry state). Insulin (5%) was added to MFu solution to produce drug-loaded particles and the release profile was assessed in phosphate buffer solution (PBS) and simulated intestinal fluid (SIF) for 24 h. Insulin was released in a sustained manner during the initial 8 h, reaching then a plateau, higher in PBS than in SIF, indicating that lower pH favors drug liberation. Moreover, the ability of MFu particles to serve as templates for the culture of human pancreatic cells was assessed using 1.1B4 cell line during up to 7 days. During the culture period studied, pancreatic beta cells were proliferating, with a global viability over 80% and tend to form pseudo-islets, thus suggesting that the proposed biomaterial could be a good candidate as versatile carrier for diabetes treatment as they sustain the release of insulin and support pancreatic beta cells viability.

Implantation of biomaterials and hybrid constructs in tissue engineering approaches presents major limitations such as inflammatory reaction and the lack of vasculature integration. Therefore, new strategies are needed to enhance implant function, immune protection, and revascularization. In this work, we developed fibrous meshes composed of fucoidan (Fu), a sulfated polysaccharide extracted from brown algae, and polycaprolactone (PCL), a synthetic biodegradable polymer, using the airbrush technique. The chemical characterization by FTIR, EDS, and XPS confirmed the presence of the two polymers in the structure of airbrushed nanofibrous meshes (ANFM). Moreover, these nanofibrous exhibited good wettability and mechanical properties envisaging their application as templates for biomaterials and cell culture. The developed ANFM were directly cultured with human pulmonary microvascular endothelial (HPMEC-ST1.6R) cells for up to 7 days. Biological results demonstrated that ANFM comprising Fu promoted cellular attachment, spreading, and proliferation of human endothelial cells. The angiogenic potential of ANFM was further evaluated by onplantation of PCL and PCL/Fu ANFM in chick chorioallantoic membrane (CAM). In ovo and ex ovo results showed that the incorporation of Fu increased the pro-angiogenic potential of ANFM. Altogether, the results suggest that airbrush biocomposite meshes could be used as a biomaterial substrate to promote vascularization.

The combination of marine origin biopolymers for tissue engineering (TE) applications is of high interest, due to their similarities with the proteins and polysaccharides present in the extracellular matrix of different human tissues. This manuscript reports on innovative collagen-chitosan-fucoidan cryogels formed by the simultaneous blending of these three marine polymers in a chemical-free crosslinking approach. The physicochemical characterization of marine biopolymers comprised FTIR, amino acid analysis, circular dichroism and SDS-PAGE, and suggested that the jellyfish collagen used in the cryogels was not denatured (preserved the triple helical structure) and had similarities with type II collagen. The chitosan presented a high deacetylation degree (90.1%) that can strongly influence the polymer physicochemical properties and biomaterial formation. By its turn, rheology, and SEM studies confirmed that these novel cryogels present interesting properties for TE purposes, such as effective blending of biopolymers without visible material segregation, mechanical stability (strong viscoelastic character), as well as adequate porosity to support cell proliferation and exchange of nutrients and waste products. Additionally, in vitro cellular assessments of all cryogel formulations revealed a non-cytotoxic behavior. The MTS test, live/dead assay and cell morphology assessment (phalloidin DAPI) showed that cryogels can provide a proper microenvironment for cell culturing, supporting cell viability and promoting cell proliferation. Overall, the obtained results suggest that the novel collagen-chitosan-fucoidan cryogels herein presented are promising scaffolds envisaging tissue engineering purposes, as both acellular biomaterials or cell-laden cryogels.

Oral administration of drugs presents important limitations, which are frequently not granted the importance that they really have. For instance, hepatic metabolism means an important drug loss, while some patients have their ability to swell highly compromised (i.e. unconsciousness, cancer…). Sublingual placement of an accurate Pharmaceutical Dosage Form is an attractive alternative. This work explores the use of the β-chitosan membranes, from marine industry residues, composed with marine sediments for dual sublingual drug delivery. As proof of concept, the membranes were loaded with a hydrophilic (gentamicin) and a hydrophobic (dexamethasone) drug. The physico-chemical and morphological characterization indicated the successful incorporated of diatomaceous earth within the chitosan membranes. Drug delivery studies showed the potential of all formulations for the immediate release of hydrophilic drugs, while diatomaceous earth improved the loading and release of the hydrophobic drug. These results highlight the interest of the herein developed membranes for dual drug delivery.

Chitosan-based porous structures have been significantly studied across the world as potential tissue engineering scaffolds. Despite the differences in chitosan produced from squid pens or crustacean shells, with the former being more reactive and easily available with a higher degree of deacetylation (DD), most of the studies report the use of crab or shrimp chitosan as they are readily available commercial sources. The aim of this work was to highlight the great potential of chitosan produced from squid pens for biomedical application. From freeze-dried scaffolds for soft tissue engineering, we investigated the influence of the DD of chitosan and the freezing temperature during processing on their performance. Chitosan was obtained by deacetylation of β-chitin previously isolated from endoskeleton of giant squid Dosidicus gigas (DD 91.2%) and compared with a commercially available batch obtained from crab shells (DD 76.6%). Chitosan solutions were frozen at −80° C or −196° C and further freeze-dried to obtain 3D porous structures (scaffolds). Scaffolds prepared at −196° C have a compact structure with smaller pores, while those prepared at −80° C showed a lamellar structure with larger pores. The compressive modulus varied from 0.7 up to 8.8 MPa. Both types of scaffolds were stable on PBS, including in the presence of lysozyme, up to 4 weeks. Furthermore, the squid chitosan scaffolds processed at −80° C promoted ATDC5 chondrocyte-like cells adhesion and proliferation. The results suggest that the developed squid chitosan scaffolds might be further exploited for applications in cartilage tissue engineering.

Event Abstract Back to Event Biomedical potential of fucoidan, a seaweed sulfated polysaccharide: from a anticancer agent to a building block of cell encapsulating systems for regenerative therapies Tiago H. Silva1, 2, Catarina Oliveira1, 2, Lara L. Reys1, 2, Diana Soares Da Costa1, 2, Rámon Novoa-Carballal1, 2, Nuno M. Oliveira1, 2, Andreia S. Ferreira3, Cláudia Nunes3, Simone S. Silva1, 2, Albino Martins1, 2, Manuel A. Coimbra3, João F. Mano1, 2, Nuno M. Neves1, 2 and Rui L. Reis1, 2 1 University of Minho, 3B´s Research Group - Biomaterials, Biodegradables and Biomimetics, Portugal 2 ICVS/3B´s - PT Government Associated Laboratory, Portugal 3 Universidade de Aveiro, QOPNA, Departamento de Química, Portugal Introduction: Marine macroalgae or seaweeds synthesize a wide variety of polymers and smaller compounds with several bioactivities, among which the sulfated polysaccharides acquire greater relevance not only due to the reported antioxidant, antiviral and anticancer[1] activities, but also to the resemblance of extracellular matrix glycosaminoglycans found in the human body[2]. In this study, the potential of fucoidan (Fu) isolated from brown seaweed Fucus vesiculosus for therapeutical use has been evaluated, focusing in its performance as antitumoral agent (bioactive role) or as building block of cell encapsulating systems (structural role). Materials and Methods: The anticancer activity of Fu extracts was assessed by evaluating the cytotoxic behavior over two human breast cancer cell lines (MCF-7 and MDA-MB-231) in in-vitro culture, using human fibroblasts and endothelial cells (HPMEC-ST1 and MRC-5, respectively) as reference. Regarding the structural role, Fu was modified by methacrylation reaction (MFu) using methacrylic acid and further crosslinked using visible radiation and triethanolamine and eosin-y as photoinitiators. The photocrosslinking was performed on MFu solution droplets placed in a silica-based superhydrophobic surface[3], allowing the formation of particles[4] (since natural Fu is highly soluble in water and ion gelation is not effective). Biological performance of the developed particles was assessed by in vitro culture of fibroblasts and pancreatic cells (L929 and 1.1B4, respectively) in contact with MFu particles, up to 7 days. The ability of the developed materials to support adhesion and proliferation of cells was evaluated for both types of cells. Results and Discussion: The tested anticancer activity is not ubiquitous on Fu extracts, being dependent on its chemical features, with molecular weight (Mw) representing a particular role. Specifically, Mw values around 60 kDa exhibited cytotoxic effects to human breast cancer cell lines, while not affecting normal fibroblasts or endothelial cells (which represent the cells of the healthy tissue that would be closer to the tumor in a real situation). A concentration range of 0.2 to 0.3 mg mL-1 from the selected Fu extract could be considered as the therapeutic window for further studies. Regarding fucoidan’s role on innovative biomaterials, the developed MFu particles could support the proliferation of fibroblasts (L929), but also of human pancreatic beta cells (1.1B4), which tend to form pseudo-islets after 7 days in culture (Fig. 1). This pancreatic cells could be also successfully encapsulated, opening a new route for a diabetes mellitus type 1 therapeutic approach. Fig. 1: Confocal microscopy images of 1.1B4 cells cultured in the presence of fucoidan-based particles and organized in pseudo-islets (red – actin; blue – nuclei). Conclusion: The present work establishes fucoidan as a high performance building block for the development of advanced therapies for cancer (targeted therapy) or tissue and organ regeneration. It shed light on the relation between chemical structure and biological activity towards anti-cancer effect and proposes novel beta cell laden particles as injectable insulin producing systems to tackle diabetes. Funding from projects 0687_NOVOMAR_1_P (co-funded by INTERREG 2007-2013 / POCTEP), CarbPol_u_Algae (EXPL/MAR-BIO/0165/2013, funded by the Portuguese Foundation for Science and Technology, FCT), POLARIS (FP7-REGPOT-CT2012-316331) and ComplexiTE (ERC-2012-ADG 20120216-321266), funded by the European Union’s Seventh Framework Programme for Research and Development is acknowledged. ASF, SSS, NMO and DSC are also thankful to FCT for their individual fellowships.References:[1] Atashrazm F, Lowenthal RM, et al., “Fucoidan and Cancer: A Multifunctional Molecule with Anti-Tumor Potential”, Marine Drugs 13 (2015), 2327-2346.[2] Silva TH, Alves A, et al., “Marine algae sulfated polysaccharides for tissue engineering and drug delivery approaches”, Biomatter 2 (2012), 278-289.[3] Rial-Hermida MI, Oliveira NM, et al., "Bioinspired superamphiphobic surfaces as a tool for polymer-and solvent-independent preparation of drug-loaded spherical particles", Acta Biomaterialia 10 (2014), 4314–4322.[4] Reys LL, Silva SS, et al., “Fucoidan hydrogels photocrosslinked with visible radiation as matrices for cell culture”, submitted. Keywords: Tissue Engineering, Biomimetic, biomaterial, biomacromolecule Conference: 10th World Biomaterials Congress, Montréal, Canada, 17 May - 22 May, 2016. Presentation Type: Poster Topic: Naturally-derived materials and biopolymers Citation: Silva TH, Oliveira C, Reys LL, Soares Da Costa D, Novoa-Carballal R, Oliveira NM, Ferreira AS, Nunes C, Silva SS, Martins A, Coimbra MA, Mano JF, Neves NM and Reis RL (2016). Biomedical potential of fucoidan, a seaweed sulfated polysaccharide: from a anticancer agent to a building block of cell encapsulating systems for regenerative therapies. Front. Bioeng. Biotechnol. Conference Abstract: 10th World Biomaterials Congress. doi: 10.3389/conf.FBIOE.2016.01.00221 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 28 Mar 2016; Published Online: 30 Mar 2016. Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Tiago H Silva Catarina Oliveira Lara L Reys Diana Soares Da Costa Rámon Novoa-Carballal Nuno M Oliveira Andreia S Ferreira Cláudia Nunes Simone S Silva Albino Martins Manuel A Coimbra João F Mano Nuno M Neves Rui L Reis Google Tiago H Silva Catarina Oliveira Lara L Reys Diana Soares Da Costa Rámon Novoa-Carballal Nuno M Oliveira Andreia S Ferreira Cláudia Nunes Simone S Silva Albino Martins Manuel A Coimbra João F Mano Nuno M Neves Rui L Reis Google Scholar Tiago H Silva Catarina Oliveira Lara L Reys Diana Soares Da Costa Rámon Novoa-Carballal Nuno M Oliveira Andreia S Ferreira Cláudia Nunes Simone S Silva Albino Martins Manuel A Coimbra João F Mano Nuno M Neves Rui L Reis PubMed Tiago H Silva Catarina Oliveira Lara L Reys Diana Soares Da Costa Rámon Novoa-Carballal Nuno M Oliveira Andreia S Ferreira Cláudia Nunes Simone S Silva Albino Martins Manuel A Coimbra João F Mano Nuno M Neves Rui L Reis Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. 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Algae are abundant sources of bioactive components with extensive therapeutic properties, receiving much interest in recent years. The research on marine brown algae, namely one of its polysaccharide-fucoidan, has increased exponentially. Fucoidan is a sulfated cell-wall polysaccharide with several reported biological properties including anticancer, antivirus, anticoagulant, antioxidant and anti-inflammatory effects. In this study, fucoidan was functionalized by grafting methacrylic groups in the chain backbone, photo-cross-linkable under visible light to obtain biodegradable structures for tissue engineering. The functionalization reaction was carried out by concentrations (8 and 12%) of methacrylic anhydride (MA). The modified fucoidan (MFu) was characterized by FTIR and 1HNMR spectroscopy to confirm the functionalization. Further, modified fucoidan was photo-cross-linked under visible light and using superhydrophobic surfaces, to obtain spherical particles with controlled geometries benefiting from the high repellence of the surfaces. When using higher concentrations of MA, the particles were observed to exhibit a smaller average diameter. Moreover, the behavior of L929 mouse fibroblast-like cells was evaluated when cultured in contact with photo-cross-linked particles was investigated, being observed up to 14 days in culture. The photo-cross-linking of MFu under visible light enables thus the formation of particles here suggested as potentially relevant in a wide range of biomedical applications.

funded by projects 0687_NOVOMAR_1_P (POCTEP), CarbPol_u_Algae (EXPL/MARBIO/0165/2013) , ComplexiTE(ERC-2012-ADG 20120216-321266). Portuguese Foundation for Science and Technology

This work was partially funded by projects 0687_NOVOMAR_1_P (POCTEP), CarbPol_u_Algae (EXPL/MAR-BIO/0165/2013) and POLARIS (FP7-REGPOT-CT2012-316331). Portuguese Foundation for Science and Technology is also gratefully acknowledged for doctoral grant of N. Oliveira and post-doc grants of S.S. Silva and D. Soares da Costa.

In recent years, much attention has been given to different marine organisms, namely as potential sources of valuable materials with a vast range of properties and characteristics. In this work, β-chitin was isolated from the endoskeleton of the giant squid Dosidicus gigas and further deacetylated to produce chitosan. Then, the squid chitosan was processed into membranes and scaffolds using solvent casting and freeze-drying, respectively, to assess their potential biomedical application. The developed membranes have shown to be stiffer and less hydrophobic than those obtained with commercial chitosan. On the other hand, the morphological characterization of the developed scaffolds, by SEM and micro-computed tomography, revealed that the matrices were formed with a lamellar structure. The findings also indicated that the treatment with ethanol prior to neutralization with sodium hydroxide caused the formation of larger pores and loss of some lamellar features. The in vitro cell culture study has shown that all chitosan scaffolds exhibited a non-cytotoxic effect over the mouse fibroblast-like cell line, L929 cells. Thus, chitosan produced from the endoskeletons of the giant squid Dosidicus gigas has proven to be a valuable alternative to existing commercial materials when considering its use as biomaterial.

Marine organisms are constituted by materials with a vast range of properties and characteristics that may justify their potential application within the biomedical field. Moreover, assuring the sustainable exploitation of natural marine resources, the valorisation of residues from marine origin, like those obtained from food processing, constitutes a highly interesting platform for development of novel biomaterials, with both economic and environmental benefits. In this perspective, an increasing number of different types of compounds are being isolated from aquatic organisms and transformed into profitable products for health applications, including controlled drug delivery and tissue engineering devices. This report reviews the work that is being developed on the isolation and characterisation of some polysaccharides, proteins, glycosaminoglycans and ceramics from marine raw materials. Emphasis is given to agar, alginates, carrageenans, chitin and chitosan, among other polysaccharides, collagen, glycosaminoglycans such as chondroitin sulphate, heparin and hyaluronic acid, calcium phosphorous compounds and biosilica. Finally, this report ends by reviewing the application of the previously mentioned materials on specific biomedical applications, in particular their participation on the development of controlled drug delivery systems and tissue engineering scaffolds.

Natural polymers are adequate renewable resources for the processability of well-defined architectures for several applications. Combinations of polysaccharides and proteins may mimic the naturally occurring environment of certain tissues. The main goal of this work renders the application of green chemistry principles, namely the use of ionic liquids (ILs) and biorenewable sources, such as chitosan (CHT) and silk fibroin (SF), to process new hydrogel-based constructs. Although the solubilization of both materials in ILs has been studied individually, this work reports, for the first time, the role of ILs as solvent, for the production of hydrogels from blends of chitosan and silk fibroin (CSF). These systems offer the advantage of being homogeneous and presenting easy and short dissolution time of both biomacromolecules. Moreover, the use of chitosan obtained from alpha- and beta-chitin allowed the production of blended hydrogels with distinct physical-chemical properties. In vitro assays demonstrated that these hydrogels supported the adhesion and growth of primary human dermal fibroblasts. Taken these properties together, the CSF hydrogels might be promising biomaterials to be explored for skin tissue engineering approaches.

Biomedical field is constantly requesting for new biomaterials, with innovative properties. Natural polymers appear as materials of election for this goal due to their biocompatibility and biodegradability. In particular, materials found in marine environment are of great interest since the chemical and biological diversity found in this environment is almost uncountable and continuously growing with the research in deeper waters. Moreover, there is also a slower risk of these materials to pose illnesses to humans. In particular, sulfated polysaccharides can be found in marine environment, in different algae species. These polysaccharides don’t have equivalent in the terrestrial plants and resembles the chemical and biological properties of mammalian glycosaminoglycans. In this perspective, are receiving growing interest for application on health-related fields. On this review, we will focus on the biomedical applications of marine algae sulfated polymers, in particular on the development of innovative systems for tissue engineering and drug delivery approaches.