Fucoidan-based hydrogels particles as versatile carriers for diabetes treatment strategies

There is a current lack of fully efficient therapies for diabetes mel-litus, a chronic disease where the metabolism of blood glucose isseverely hindered by a deficit in insulin or cell resistance to thishormone. Therefore, it is crucial to develop new therapeutic strat-egies to treat this disease, including devices for the controlleddelivery of insulin or encapsulation of insulin-producing cells. Inthis work, fucoidan (Fu)â a marine sulfated polysaccharide exhib-iting relevant properties on reducing blood glucose and antioxi-dant and anti-inflammatory effectsâ was used for thedevelopment of versatile carriers envisaging diabetes advancedtherapies. Fu was functionalized by methacrylation (MFu) using8% and 12% (v/v) of methacrylic anhydride and further photo-crosslinked using visible light in the presence of triethanolamineand eosin-y to produce hydrogel particles. Degree of methacryla-tion varied between 2.78 and 6.50, as determined by1HNMR, andthe produced particles have an average diameter ranging from0.63 to 1.3mm (dry state). Insulin (5%) was added to MFu solutionto produce drug-loaded particles and the release profile wasassessed in phosphate buffer solution (PBS) and simulated intes-tinal fluid (SIF) for 24h. Insulin was released in a sustained man-ner during the initial 8 h, reaching then a plateau, higher in PBSthan in SIF, indicating that lower pH favors drug liberation.Moreover, the ability of MFu particles to serve as templates forthe culture of human pancreatic cells was assessed using 1.1B4cell line during up to 7 days. During the culture period studied,pancreatic beta cells were proliferating, with a global viabilityover 80% and tend to form pseudo-islets, thus suggesting thatthe proposed biomaterial could be a good candidate as versatilecarrier for diabetes treatment as they sustain the release of insulinand support pancreatic beta cells viability.We acknowledge ERDF for the financial support through POCTEP Project 0687_NOVOMAR_1_P, under the scope of INTERREG 2007-2013, and project 0302_CVMAR_I_1_P, under the scope of INTERREG Espana-Portugal 2014-2020, and Structured Projects NORTE-01-0145-FEDER-000021, NORTE-01-0145-FEDER-000023 and ATLANTIDA (ref. NORTE-01–0145-FEDER-000040), under the scope of Programa Operacional Regional do Norte (Norte 2020). Funding from the Portuguese Foundation for Science and Technology for doctoral grant (SFRH/BD/112139/2015) and post-doctoral grant (SFRH/BPD/85790/2012) is also acknowledge

Evaluation of the potential of fucoidan-based microparticles for diabetes treatment

Abstract INTRODUCTION: Marine organisms have in their constitution materials with a wide range of properties and characteristics inspiring their application within the biomedical field. One important example is fucoidan (Fu), an underexploited sulfated polysaccharide extracted from the cell wall of the brown seaweeds, with high solubility in water1. Fucoidan is composed of L- fucose and glucuronic acid including sulfate groups and has important bioactive properties such as antioxidative, anticoagulant, anticancer and in the reduction of blood glucose1,2. In this work, the biomedical potential of fucoidan was assessed by processing modified fucoidan (MFu) into microparticles by photocrosslinking using superhydrophobic surfaces and visible light3,4. Biological performance on the developed constructs using human pancreatic beta cells is currently under investigation. METHODS: To design the materials structures, fucoidan was modified by methacrylation reaction3. Briefly, Fu aqueous solution 4% w/v was mixed with methacrylated anhydride (MA) in volume of 12% v/v at 50oC to react for 6h. Further, MFu particles with and without insulin (0.5% w/v) were produced by pipetting a solution of 5% MFu v/v with triethanolamine and eosin-y (photoinitiators) onto superhydrophobic surfaces4 (Fig. 1A) and then photocrosslinking using visible light4. MFu and developed particles were characterized using 1HNMR, turbidimetry and SEM to assess their chemistry and morphology, respectively. Moreover, the insulin release was evaluated in phosphate buffered saline (PBS) solution at pH 7and simulated intestinal fluid (SIF) at pH 5. The ability of the developed materials to support adhesion and proliferation of cells was assessed by suspension culture of human pancreatic cells 1.1B4 (3.5x105 cells/ml) in contact with MFu microparticles during up to 7 days. RESULTS: The chemical modification performed on Fu was confirmed by the presence of vinyl and additional methyl peaks in the 1HNMR of modified fucoidan, not present in Fu spectrum. Methacrylated fucoidan was obtained with a methacrylation degree of 17%. The produced fucoidan particles have round shape and average diameter of 1.53 mm (Fig. 1B). The insulin release in PBS and SIF demonstrate that the particles can release insulin in a sustained manner under the studied period. It seems that the insulin release is slower for SIF (pH5, Fig. 1C), than for PBS. The biological tests regarding the culture of pancreatic beta cells demonstrate that cells show a round-like shape and tend to form pseudo-islets during the culture period studied (Fig. 1D). DISCUSSION & CONCLUSIONS: This work demonstrates the successful production of fucoidan- based-microparticles through the methacrylation of fucoidan, using visible light and superhydrophobic surfaces. The covalent crosslinking methacrylated fucoidan through visible light represents a promising method to obtain biocompatible fucoidan particles with a uniform round shape. The obtained insulin release profiles are sensitive to different pH (pH7 and pH5), mimicking the normal physiological pathway for insulin release. Furthermore, the results suggest these systems could be used for treatment of type I diabetes mellitus as they sustain beta cells viability and proliferation. The response also suggested, that the MFu particles could be a good candidate as drug delivery vehicles for the diabetes mellitus treatment. REFERENCES: 1 Silva TH et al (2012), Biomatter 2(4): 278:289. 2Sezer Alidemir et al (2011), Fucoidan: A versatile biopolymer for biomedical applicatons (Springer Ber.Heid).pp377-406. 3Mihaila S.et al (2013), Adv. Health. Mat. 2(6): 895-907. 4Rial Hermida et al, Acta Biomater.(2014) 10(10) 4314-4322. ACKNOWLEDGEMENTS: This work was partially funded by projects 0687_NOVOMAR_1_P (POCTEP), CarbPol_u_Algae (EXPL/MAR- BIO/0165/2013), ComplexiTE (ERC-2012-ADG 20120216-321266). Portuguese Foundation for Science and Technology is also gratefully acknowledged for doctoral grants of L. Reys and N. Oliveira and post- doctoral grants of S.S. Silva and D. Soares da Costafunded 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 Technologyinfo:eu-repo/semantics/publishedVersio

Biomedical potential of fucoidan, a seaweed sulfated polysaccharide: from a anticancer agent to a building block of cell encapsulating systems for regenerative medicine

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 fucoidanâ 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 â actin; blue â 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

Angiogenic potential of airbrushed fucoidan/polycaprolactone nanofibrous meshes

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 proangiogenic potential of ANFM. Altogether, the results suggest that airbrush biocomposite meshes could be used as a biomaterial substrate to promote vascularization.Northern Portugal Regional Operational Programme (NORTE 2020), under the Portugal 2020 Partnership Agreement, through Structured projects NORTE-01-0145-FEDER-000021 and NORTE-01-0145-FEDER-000023. Financial support from the Portuguese Foundation for Science and Technology (FCT) is also acknowledged through PhD fellowship SFRH/BD/112139/2015 and R&D&I projects PTDC/CTM-CTM/29813/2017 and PTDC/BII-BIO/31570/2017

Building fucoidan/agarose-based hydrogels as a platform for the development of therapeutic approaches against diabetes

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 Youngâ 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.This research was funded by the Portuguese Foundation for Science and Technology (FCT), under the scope of individual fellowships/contracts (SFRH/BD/112139/2015, and SFRH/BPD/93697/ 2013, SFRH/BPD/85790/2012 and CEECIND/01306/2018) and research projects (PTDC/CTMCTM//29813/2017 and PTDC/BII-BIO/31570/2017); by the Northern Portugal Regional Operational Programme (NORTE 2020), under the scope of Structured Projects NORTE-01-0145-FEDER-000021 and NORTE-01-0145-FEDER-000023; and by European Union Transborder Cooperation Program Interreg España-Portugal 2014-2020 (POCTEP), under the scope of project 0302_CVMAR_I_1_P

Polissacarídeos de origem marinha para aplicação em estratégias de engenharia de tecidos

Os polissacarídeos de origem marinha tem recebido atenção crescente, quer dos cientistas, quer dos engenheiros a nível industrial, pelas várias propriedades que lhes são conhecidas, bem como pela variedade química que evidenciam. São originários de um meio ainda pouco explorado, mas ao qual se tem procurado aplicar estratégias sustentáveis de utilização, pelo que o seu potencial de aplicação é, de facto, impressionante. Neste capítulo, será dada particular ênfase a dois polissacarídeos – quitosano e fucoidana – a título de exemplo, em ambos os casos atendendo à sua semelhança aos glicosaminoglicanos presentes na matriz extracelular e por isso prometendo interesse muito significativo para a regeneração de tecidos

Sulfated seaweed polysaccharides

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.The authors would like to acknowledge the funding received from European Union through INTERREG España-Portugal 2014-2020 Programme, under the scope of the projects 0302_CVMAR_I_1_P and 0474_BLUEBIOLAB_1_E, as well as through ERDF with Programa Operacional Competitividade e Internacionalização (Portugal), under the scope of the project VALORMAR: Valorização Integral dos Recursos Marinhos: Potencial, Inovação Tecnológica e Novas Aplicações (POCI-024517-FEDER)

Dual delivery of hydrophilic and hydrophobic drugs from chitosan/diatomaceous earth composite membranes

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 beta-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.The research leading to these results has received funding from Erasmus Mundus Joint Programmes, ERDF / POCTEP 2007-2013 under project 0687_NOVOMAR_1_P, from the European Union Seventh Framework Programme (FP7/2007-2013) under grant agreement number REGPOT-CT2012-316331-POLARIS, and from the North Portugal Regional Operational Programme (ON.2 - O Novo Norte), within the National Strategic Reference Framework (QREN 2007-2013) under the project NORTE-01-0124-FEDER-000018. Portuguese Foundation for Science and Technology is also acknowledged for the post-doctoral fellowship SFRH/BPD/112140/2015, for the doctoral fellowship SFRH/BD/112139/2015 and for the funds provided under the program Investigador FCT 2012 (IF/00423/2012). Dr. Helder Santos (University of Helsinki) is also acknowledged for valuable discussions on the concept.The research leading to these results has received funding from Erasmus Mundus Joint Programmes, ERDF / POCTEP 2007–2013 under project 0687_NOVOMAR_1_P, from the European Union Seventh Framework Programme (FP7/2007–2013) under grant agreement number REGPOT-CT2012-316331-POLARIS, and from the North Portugal Regional Operational Programme (ON.2 – O Novo Norte), within the National Strategic Reference Framework (QREN 2007-2013) under the project NORTE-01-0124-FEDER-000018. Portuguese Foundation for Science and Technology is also acknowledged for the post-doctoral fellowship SFRH/BPD/112140/2015, for the doctoral fellowship SFRH/BD/112139/2015 and for the funds provided under the program Investigador FCT 2012 (IF/00423/2012). Dr. Hélder Santos (University of Helsinki) is also acknowledged for valuable discussions on the concept.info:eu-repo/semantics/publishedVersio