Materials Today Communications
Kefiran cryogels as potential scaffolds for drug delivery and tissue engineering applications
Graphical abstract
Introduction
Tissue engineering (TE) has emerged as a new approach involving the combination of cells, biomaterial scaffolds and bioactive agents to fabricate functional new tissues replacing the damaged ones. Polymeric scaffolds are fundamental components of TE approaches, since they provide an optimal cellular microenvironment, allowing 3D (three-dimensional) cellular adhesion, growth, and differentiation [1]. In addition, these scaffolds can promote high and efficient drug-loading to specific sites, providing local delivery of the appropriate dose of drug over a given time range, reducing the drug concentration at non-target sites, and, therefore promoting tissue regeneration [2]. In order to be functional for drug delivery and TE, polymeric scaffolds need to have particular physicochemical properties, and present easy-scalable and cost-effective manufacturing methods that will determine their success in TE approaches.
Polymeric scaffolds can form 3D cryogels by a freeze gelation technique. In fact, the most common processing methods for synthetizing cryogels are freeze-drying, salt leaching, particulate leaching and lyophilisation. However, recently freeze gelation has been reported and described as a time and energy efficient technique [3]. These cryogels are sponge-like networks with interconnected macropores that are formed in moderately frozen solutions of monomeric or polymeric precursors [4]. The unique structure of cryogels, in combination with their amazing chemical and physical properties, makes them attractive for several biomedical applications, especially as scaffolds for delivery systems and tissue engineering applications [5]. In fact, these cryogels are considered potential carriers for molecules of pharmaceutical interest, growth factors and cells [4,6].
A variety of polysaccharide-based scaffolds like alginate, dextran, hyaluronic acid, gellan gum and chitosan, among others, have been recently used in TE and regenerative medicine strategies due to their excellent biocompatibility combined with their potential biodegradability [6]. Among the remarkable polysaccharides used lately in several applications, it is possible to find the water-soluble branched glucogalactan Kefiran extracted from the flora of kefir grains [7]. This natural biopolymer has received increasing interest because of its safe status and its outstanding properties for biomedical applications [[7], [8], [9]]. However, achieving translational medicine together with product commercialization requires regenerative polymers to be safe and functional, with cost-effective and mass-production manufacturing methods. Therefore, we propose the development of a new advanced functional polymer, Kefiran, manufactured with a cost-effective and easy scalable method, freeze gelation.
Our research aimed to develop and characterise a novel 3D polymeric scaffold for drug delivery and TE applications based on Kefiran cryogels. The Kefiran scaffold was characterised in terms of the microstructure (micro-CT and SEM analysis), degradation assessment, and mechanical properties (rheometer analysis). Then, Kefiran-based cryogels were evaluated also for controlled delivery of diclofenac sodium. Finally, human Adipose Stem Cells (hASCs) were cultured onto the Kefiran scaffold, and their metabolic function was measured as indicator of their viability. Kefiran scaffolds are rarely reported as drug delivery systems [10]; however, a recent study described Kefiran-alginate gel microspheres for oral delivery [11]. Nevertheless, it is the first time to our knowledge that Kefiran cryogels are evaluated for delivery of diclofenac sodium in combination with its potential use as a TE scaffold.
Section snippets
Materials
AlamarBlue® reagent, Diclofenac sodium, Kefiran, Phosphate-buffered saline, Quant-iT PicoGreen dsDNA kit, Ultrapure water.
Preparation of diclofenac sodium solution
Diclofenac sodium is the sodium salt form of diclofenac, a benzene acetic acid derivate and nonsteroidal anti-inflammatory drug (NSAID) with analgesic, antipyretic and anti-inflammatory activity. Diclofenac sodium salt was provided by Sigma-Aldrich (cat 15307-79-6, D6899). Diclofenac sodium solution was prepared at a concentration of 0.5 mg/mL in ultrapure H2O.
Production of Kefiran cryogel
Kefiran
Results and discussion
Kefiran polysaccharide with its exceptional properties could be a potential candidate for future 3D biomaterial scaffold development. This Kefiran scaffold will interact with biological environment, deliver bioactive molecules such as drugs and growth factors, and act as cell adhesion mediators to cellular functioning and differentiation towards TE. Moreover, in TE, the 3D scaffolds should have an interconnected pore structure and high porosity to guarantee cellular penetration, proliferation
Conclusion
Kefiran cryogels were evaluated as scaffolds for TE and controlled drug delivery. The developed scaffold showed stability, elastic behavior and high porosity 3D structure, capable of controlled release of diclofenac drug for two weeks. Moreover, hASCs seeded on Kefiran scaffold revealed that this platform is biocompatible, sustaining cell metabolic activity for 72 h, a fundamental feature for tissue engineering and regenerative medicine. These features make Kefiran cryogels an appealing
Declaration of Competing Interest
The authors declare no conflicts of interest
Acknowledgements
Hajer Radhouani, Cristiana Gonçalves and F. Raquel Maia were supported by the Fundação para a ciência e a Tecnologia (FCT) from Portugal, with references CEECIND/00111/2017, SFRH/BPD/94277/2013 and SFRH/BPD/117492/2016, respectively. Diana Bicho was supported through the M-ERA-NET/0001/2014 project (FCT). Joaquim. M. Oliveira would like to thank the FCT for the fund provided under the program Investigador FCT 2015 (IF/01285/2015). This work was funded by the R&D Project KOAT – Kefiran
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