Cross-linked gelatin-nanocellulose scaffolds for bone tissue engineering
Introduction
Recent advances in the field of bone tissue engineering (BTE) present encouraging approaches to solve challenges associated with current bone augmentation techniques [1]. Hydrogels are a group of polymers that can absorb water up to thousand folds their dry weight, creating a gel-like environment mimicking stem cell niche [1]. Recently, a promising class of hydrogels have been produced from wood-based cellulose nanofibrils (CNFs). The CNFs have, due to their nanostructure, both high specific surface area and high surface reactivity, but are slowly degradable in vivo [2]. Insufficient degradation rate will cause the host to respond with a foreign body reaction [16]. A new hydrogel has been developed by blending gelatin (Gel) with CNF, to better meet requirements for BTE [6]. Mechanical properties and the degradation rate of the Gel-CNF hydrogel were further adjusted with different cross-linking approaches [6].
In this study, Gel-CNF hydrogels were cross-linked by either dehydrothermal treatment (DHT), or by a combination of hexamethylenediamine (HMDA), genipin, and DHT. The DHT is a physical treatment that includes high temperature and vacuum to remove the water from the matrix, resulting in the formation of intermolecular cross-links [4]. The HMDA is a diamine which is commonly used with biomaterials containing carboxyl groups, creating covalent bonds between oxidized CNFs [5]. The biodegradable molecule genipin reacts spontaneously with amino acids, and can cross-link the gelatin chains [6]. However, there is a risk of introducing cytotoxicity to the material when using chemical reagents as cross-linkers.
The present study was undertaken to investigate the osteogenic potential of these cross-linked scaffolds in vitro. Human bone marrow mesenchymal stem cells (hBMSCs) were cultured on scaffolds and their initial biological responses studied in terms of cell attachment, viability, and osteogenic differentiation.
Section snippets
Preparation of scaffolds
Oxidized CNFs were prepared by 2,2,6,6-tetramethylpiperidine-1-oxyl radical (TEMPO) according to Saito et al. [7]. To create interconnected porosity by ice crystal formation, the CNF suspension was frozen at −20 °C for 24 h before freeze-drying for another 24 h. Porous Gel-CNF scaffolds cross-linked by DHT (Gel-CNF1) and porous Gel-CNF scaffolds cross-linked by HMDA, genipin, and DHT (Gel-CNF2) were prepared according to Campodini et al. [6].
Scaffolds structure and porosity
Assessment of the 3D architecture was performed by
Results and discussion
Architecture of the scaffolds strongly influences their mechanical properties. The freeze-drying method used, produced hydrogel scaffolds with variable porosity and interconnected pores as shown by µ-CT (Fig. 1A-C) and SEM (Fig. 1D-F). Both total porosity and pore sizes of CNF scaffolds were higher than that of the Gel-CNF samples, regardless the method of cross-linking (Fig. 1, Table 1). Previously, Campodoni et al. reported that porous scaffolds from Gel-CNF hydrogels, cross-linked by either
Conclusion
The cross-linking methods used to modify mechanical properties and degradation rates of the Gel-CNF scaffolds did not generate any adverse biological responses to hBMSCs and supported their osteogenic differentiation. The results of this study suggest that the cross-linked Gel-CNF hydrogels are promising materials for BTE.
CRediT authorship contribution statement
Ingeborg Elisabeth Carlström: Writing - original draft, Writing - review & editing. Ahmad Rashad: Conceptualization, Methodology, Writing - review & editing, Supervision. Elisabetta Campodoni: Conceptualization, Methodology, Writing - review & editing. Monica Sandri: Writing - review & editing. Kristin Syverud: Conceptualization, Writing - review & editing. Anne Isine Bolstad: Methodology, Writing - review & editing, Supervision. Kamal Mustafa: Conceptualization, Methodology, Writing - review &
Declaration of Competing Interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgment
This work has been funded by the Research Council of Norway through the NORCEL project, (Grant no. 228147) and Trond Mohn Foundation (TMS, project no. BFS2018TMT10).
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These authors contributed equally to this study.