Materials Today Chemistry
3D graphene-containing structures for tissue engineering
Graphical abstract
Introduction
Replication and restoration of tissues/organs via modern tissue engineering techniques promise to mitigate the global health crisis because of insufficient tissue/organ donation, compounded by an aging population and need to treat newly discovered or incurable diseases and disabilities. Specific physicochemical properties of biomaterials combined with novel approaches to their application place them center stage as next-generation medical devices for regenerative medicine.
In 2004, monolayer carbon material, i.e. graphene, was discovered and prepared by Geim and Novoselov using a scotch tape method [1,2]. Arranged in a honeycomb lattice, graphene shows exceptional surface-to-volume ratio, high carrier mobility, and good electrical, optoelectronic, and mechanical properties (Fig. 1) [3], paving the way toward a postsilicon era [4,5]. The exceptional electronic properties of graphene are because of its special crystal lattice structure, whereby bonding between each carbon atom is hybridized sp2 with additional π orbitals [6]. There are two π orbitals in each unit cell of graphene, which is dispersed to form two π bonds, both of which could be considered as bonding and antibonding in nature. These unique properties make pristine graphene a mixture of gapless metal and semiconductor [7].
As a soft layered structure, graphene can be decorated with hydroxyl, epoxide, and carboxyl groups on the edge and basal plane through oxidization, rendering graphene oxide (GO). Graphene is hydrophobic and inclined to aggregate, whereas GO is hydrophilic and well dispersible in various solutions, including water. These characteristics make GO easy to process and possess high affinity to biomolecules. To expand the bionic applications of graphene and its derivatives, a more detailed comprehension of their properties in the biological context is needed. Most graphene and derivatives have proven to be cytocompatible in vitro and in vivo [8]. However, the physicochemical properties of the 2D materials, including structure shape, size, surface functionality, concentration, and aggregation state, have a major influence on cytocompatibility [9]. Flat 2D graphene materials are considered less toxic than tubular forms (e.g. single-walled carbon nanotubes [SWCNTs]), with cell membrane integrity being retained at up to a 1000 fold higher graphene concentration compared with SWCNT, in part because of their softer nature [10]. In addition, it has been demonstrated that penetration of cell membranes is dominated by the graphene edge shape, and so graphene with sharp edge features is more likely to damage the cell membrane [11]. Nanoscale graphene materials, especially in 10–100 nm size, can induce cytotoxicity, inflammation, and even genotoxicity when they are translocated into cells and cell nuclei with less steric hindrance compared with larger sized graphene materials [12,13]. Besides the intrinsic physical properties, the concentration and aggregation state of graphene also affect cytocompatibility, with diluted and less aggregated graphene solutions being more compatible [10,12]. Surface chemistry of graphene can also affect its interaction with cells by modulation of hydrophilicity and hydrophobicity, which subsequently affects aggregation. Nanoscale graphene decorated with more oxygen-containing groups is more easily internalized by cells, while the influence on the perturbation of the cell membrane becomes more irregular [14]. Although there are generalized effects of graphene on cells, cell specific effects should also be considered, e.g. different endocytic pathways of GO nanosheets are observed with osteoblasts, macrophages, and hepatocytes [15].
Structural defects in graphene layers, such as vacancies, adatoms, and substitutional impurities, introduced during preparation have significant influence on graphene properties [16]. Epitaxial growth [17], chemical vapor deposition (CVD) [18], micromechanical exfoliation [2], and chemical synthesis [19] are the four principal methods for graphene preparation. Graphene synthesized by epitaxial growth and CVD methods shows superior physicochemical properties and controllability over quality because of continuous bottom-up synthesis procedure [20], but the fabrication process is complicated requiring expensive specialized equipment and raw materials [21]. Micromechanical exfoliation can produce multilayer, bilayer, and single layer graphene with less defects on the sheet, but this method is hard to scale, and layer size is small [22]. Chemical synthesis of graphene is currently the most widely used and processable method for graphene preparation. This approach is more easily scaled than the other three methods, although graphene obtained by the other methods shows fewer defects and better controllability over graphene shape and azimuthal orientation [23]. For the chemical synthesis method, graphene is prepared through exfoliation and oxidation of graphite with additional chemical/thermal reduction treatment [24,25]. Typically, strong acid and oxidizing agents are employed for graphite expansion and oxidization, with further exfoliation of oxidized graphite to GO by mechanical treatment. The obtained GO can be reduced to graphene via a variety of techniques, such as microwave, thermal annealing, and chemical reduction [26]. Processability of the solution phase without the need for expensive facilities enable production of large quantities, making graphene more accessible in industry. However, the defects and non-uniform morphology of graphene made by chemical synthesis are possibly because of strong acid oxidation during the oxidization procedure and exfoliation during the ultrasonication process [25].
Graphene and its derivatives, as 2D materials, have found applications in many research fields, namely energy storage [27], drug delivery [28], tissue regeneration, etc. [29]. Transition from 2D structures to 3D provides new opportunities for graphene-containing structures. This is particularly so in the field of bioengineering, where the use of 3D structures has been shown to exhibit significant advantages over 2D [30], because the 3D graphene-containing structure (3DGCS) systems not only possess intrinsic graphene properties but also have high surface to volume ratio, larger surface area for decoration, abundant embedded binding sites, and many other remarkable properties. Especially, 3DGCSs with large surface area, micropores/channels, and improved biocompatibility, mechanical properties, and electrical conductivity are emerging as better platforms for tissue engineering and other bionic applications. In addition, cell activity can be enhanced by using 3D structures through enhanced cell adhesion, interaction, migration, proliferation, and differentiation [28,31].
Although graphene has received enormous attention in tissue engineering application recently, most of the published reviews are focused on introducing the properties of graphene-based materials and general biomedical application. Biomimetics has become a hot research topic, and there has been significant accomplishment in biomimetic 3DGCSs for tissue engineering, while there is still no systematic summary in this respect. In this review, application of 3DGCSs in tissue engineering and fabrication methods that enable production of 3DGCSs are summarized and discussed. Biosafety and biodegradability issues are also discussed with future perspectives provided at the end of the article.
Section snippets
Application of 3DGCSs for tissue engineering
Tissue engineering represents an advanced tool for treating and reconstructing defective tissues/organs using, for example, biomimetic scaffolds, while traditional therapy strategies have limitations, such as matchable donor shortage and post-transplant immune monitoring. Tissue engineering scaffolds are required to be biocompatible and present various cues to guide cell proliferation or differentiation toward diverse lineages both in vitro and in vivo. 3DGCSs with abundant embedded cell
Fabrication
The majority of methods devised for fabricating 3DGSCs involve graphene synthesized by chemical method. Importantly, a chosen fabrication strategy will impact the performance and clinical approvability and utility of a construct. In this section, principle fabrication strategies and resulting 3DGCSs are summarized and discussed.
Biodegradability of 3DGCS
The advent of graphene has advanced bionic research in the past several years because of its prominent physicochemical properties [28]. Unprecedented progress in bionic applications of graphene has raised concerns for its biopersistence in human tissue and organ, which needs to be addressed before clinical application.
Surface functionalization, lateral size, edge structure, composition, and administered doses are all important factors for graphene biocompatibility [204]. For example, cellular
Summary and outlook
During the past decade, a variety of 3DGCSs with diverse architectures, various compositions and different properties have been fabricated via a number of strategies. Among all these strategies, template assembly and self-assembly are the most efficient and widely used techniques. Except for the CVD method in template assembly strategy, other methods sourcing from processable graphene are amenable for large-scale production, which can significantly accelerate applications of bulky graphene
Acknowledgements
The authors gratefully acknowledge the Australian Research Council Centre of Excellence Scheme for provision of funding (CE140100012) and support from the Australian National Fabrication Facility—Materials Node. Prof. Gordon G. Wallace wishes to acknowledge the financial support from ARC through an ARC Laureate Fellowship (FL110100196). Prof. Gordon G. Wallace and Dr. Xiao Liu would like to acknowledge the support of ARC Industrial Transformation Training Centre in Additive Biomanufacturing (
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