Accelerating bioelectric functional development of neural stem cells by graphene coupling: Implications for neural interfacing with conductive materials
Introduction
It is critical to develop new materials for manipulating neural stem cell (NSC) behavior for neural regeneration and tissue engineering. Current strategies mostly focus on the different biochemical modifications of the materials, while less effort had been made to explore how the physical properties of the materials affect NSC behaviors. The clearest effect of the materials on cells is their ability to influence membrane structure or function. It is well known that ion channels and pumps in the membrane play critical roles in cell function, including proliferation, migration, and apoptosis, in a wide range of cells [1]. In particular, the current literature has described the regulatory role of bioelectric membrane gradients and signaling in a variety of tissues during development, maturation, and regeneration [2], suggesting the importance of being able to manipulate these bioelectric membrane properties in regenerative medicine.
Graphene, a 2-dimensional monolayer of carbon atoms, has been at the forefront of nanotechnology due to the material's unique electrical, mechanical, and thermal features. It has recently been considered to be a promising candidate for the fabrication of ultrafast nanoelectronic devices, transparent electrodes, nanocomposite materials, and biomedical materials [3]. It has already been utilized in a variety of biomedical applications, including cellular imaging and drug delivery [4], bio-analysis [5], stem cell research [6], [7], and even photothermal therapy for treating tumors [8]. Recently, we and other groups discovered the possibility of using graphene as a neural interfacing material because it could promote neurite sprouting and outgrowth in human neuroblastoma (SH-SY5Y) cell culture [9], PC-12 cells [10], primary cultures of hippocampal neurons [11], and direct NSC differentiation to neurons [12], [13], facilitate NSCs differentiation into neurons on graphene nanomesh semiconductors and form neuronal fibers [14], [15]. In addition, more and more studies have shown that graphene exhibits the potential capability of manipulating the fate of stem cells. For instance, graphene-based materials are capable of inducing NSC differentiation into the neuronal lineage [7], [16], controlling and even accelerating the differentiation of mesenchymal stem cells [6], [17], [18], [19], [20], [21], [22], and regulating the behaviors of other types of stem cells, including pluripotent stem cells and embryonic stem cells [23], [24], [25]. These pioneering studies clearly demonstrate the great potential of graphene-based materials in cell therapies. However, the underlying mechanisms behind the altered cell behaviors, such as enhanced differentiation and promoted cell growth, remain largely unknown.
The strong connections between cell functions and the cell membrane's bioelectric properties inspired us to investigate whether graphene can regulate NSC development and the maturation of their progeny by affecting the bioelectric properties of the cell. In this work, we studied the impact of graphene on the maturation of the electrophysiological state during NSC development, including the passive and the active bioelectric properties and subsequent choice of NSC fate.
Section snippets
Graphene film preparation
Graphene samples were synthesized according to the previously published CVD method [26]. Briefly, a thin copper foil (5 cm × 5 cm) was heated to 1000 °C and annealed for 20 min under H2 and Ar gases, followed by exposure to H2 and CH4 for 5 min. The films were then cooled down from 1000 °C to room temperature under H2 and Ar gases. Graphene films were removed from the copper foils by etching in an aqueous solution of iron nitrate. After the copper film was dissolved, a TCPS substrate was
Graphene substrate preparation
The graphene films were synthesized by the chemical vapor deposition (CVD) method [26] and then transferred to a tissue-culture polystyrene (TCPS) substrate (TCPS alone was used as the control substrate in the following experiments). The graphene films exhibited good characteristics of light transmission (Fig. 1A). Based on the optical transmittance (∼90% optical transmittance at wavelengths of 500–1000 nm) and room-temperature micro-Raman spectroscopy of the graphene films, we estimated that
Conclusion
In summary, graphene can accelerate the maturation of NSCs and their progeny by affecting the passive and active bioelectric membrane properties, including hyperpolarizing the VR, up-regulating the expression of TREK-1 channels, and increasing the probability of firing APs. Our results suggest that graphene as a neural interfacing material can remodel the passive and active bioelectric properties of NSCs and their progeny, which is further accompanied by increased differentiation of NSCs into
Conflict of interest
The authors declare no competing financial interests.
Acknowledgements
This work was supported by grants from the Major State Basic Research Development Program of China (973 Program) (2015CB965000), the National Natural Science Foundation of China (31571530, 81470692, 81500790, 31500852, 81570921, 31501194), the Natural Science Foundation from Jiangsu Province (BK20151404, BK20150022, BK20140620, BK20131290), the Yingdong Huo Education Foundation, the Open Research Funds of the State Key Laboratory of Genetic Engineering, Fudan University (SKLGE-1407) and the
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These authors contributed equally to this work.