The related article can be found in this link: https://www.nature.com/articles/s41467-018-05599-2

Considering both intrinsically limited regenerative capability of the central nervous system (CNS) and the complex inhibitory microenvironment of injured spinal cord, developing effective therapeutics for CNS diseases and injuries [e.g., Parkinson disease and spinal cord injury (SCI)] has been challenging. To this end, stem cell therapy can provide a promising solution. First, they can differentiate into neurons and restore the damaged neuronal circuits. Second, stem cells can secret tropic factors to modulate the inhibitory microenvironment of diseased or injured CNS areas. Nevertheless, low survival rate and incomplete differentiation control of stem cells in vivo remain critical barriers for stem cell therapy before its full potential can be realized. As such, there is an urgent need to develop an innovative approach for enhanced stem cell transplantation and for better controlling of stem cell fate in vivo. 

Motivated by these critical challenges, our group has been developing nanomaterial-based bioscaffolds (nanoscaffolds) for controlling neural stem cell (NSC) behaviors. More specifically, we successfully demonstrated several 2D carbon nanomaterial-based hybrid nanoscaffolds that guides NSCs differentiate into different neural cell lineages ^1-3.  Graphene and its derivatives-based nanoscaffolds are quite attractive for biomedical applications due to their unique biophysical properties such as strong binding towards biomolecules including extracellular matrix (ECM) proteins,  and their electrical/electrochemical properties (Fig. 1) ^4,5. However, a few years ago, we realized one drawback of these graphene-based 2D/3D nanomaterials for clinical stem cell transplantation: they are intrinsically non-degradable due to the strong sp2 carbon bonds. After realizing this potential barrier to translational research, we thought hard from scratch and asked ourselves a question: “Can we develop a biodegradable 2D/3D graphene mimic?” Then my student Letao Yang searched literature, tested several different 2D nanomaterials and finally identified MnO₂ nanosheet, a mild oxidant that has been widely applied used for batteries and recently developed for cancer applications as a promising candidate ^6,7. We hypothesized and proved that MnO₂ nanosheet can not only degrade by intracellular bioreductants, but also extracellular bioreductants such as vitamin C, which is essential for scaffold-based degradation. Unprecedentedly, even without any exogenous bioreductants, we observed significant biodegradation of MnO₂ 2D/3D nanosheets when they are assembled with cells. These results are very encouraging as most inorganic nanomaterials (e.g., TiO₂ nanotubes, carbon nanotubes, gold nanowires) haven’t shown such robust endogenous biodegradability. 

Fig. 1 Our previous work on Graphene-based hybrid nanoscaffolds for neural differentiation of stem cells. a, Scheme (image on the top), scanning electron microscope (SEM, image on the middle) and immunostaining image (image on the bottom) on a mature neuronal marker (MAP2) showing the enhanced axonal alignment of neurons differentiated from human NSCs on a silica nanoparticle-graphene oxide hybrid nanoscaffold. Scale bars: 10 μm and 50 μm for the middle and bottom images, respectively. b, Scheme (image on the top), SEM (image on the middle) and fluorescent images (image on the bottom) showing the enhanced adhesion and oligodendrocytic differentiation of GFP-labelled rat neural stem cells on a nanofiber-graphene oxide hybrid nanoscaffold. Scale bars: 10 μm and 20 μm for the middle and bottom images, respectively. c, Graphene-based hybrid nanoscaffolds for enhanced neuronal differentiation. The top image is a scheme illustrating the interactions between NSCs and nanoscaffolds. The middle optical microscope image characterizes a grid-shaped graphene oxide micropattern. The bottom immunostaining image on neuronal marker (TuJ1) indicates the neuronal differentiation from mesenchymal stem cells on the graphene oxide grid-shaped micropattern. Scale bars: 50 μm.

We then moved on to constructing a biodegradable hybrid inorganic (BHI) nanoscaffold using MnO2 nanosheet as building blocks, and we aimed to provide a favorable soluble and insoluble microenvironment for the transplanted stem cells (Fig. 2) in vitro as well as in vivo 8. More specifically, we assembled our nanoscaffolds from nanosheets, neurogenic drugs, and ECM proteins through a non-covalent crosslinking mechanism based on a strong binding between the nanosheet and ECM protein.  

Fig.2 BHI nanoscaffolds for advanced stem cell therapy. a, To develop an effective method for stem cell transplantation, we synthesized a BHI nanoscaffolds that simultaneously integrate advancements in 3D-hybrid nanomaterials and DFT simulation-based precision drug screening. Cells are labelled with green due to their green fluorescence protein labelling. Laminin proteins are colored in blue. Drugs are represented by red-colored dots. In the simulation scheme, blue colored atoms represent manganese and red colored atoms represent oxygen. b, By transplanting NSCs using 3D-BHI nanoscaffold, cell survival and neuronal differentiation were improved, which leads to a reduction of glialscar and enhanced functional recovery. c, Immunostaining images showing the survival and guided neuronal differentiation of NSCs in the SCI sites. Scale bar in the image on the left: 250 μm; Scale bar in images on the middle and right: 100 μm.

From our studies, we demonstrated several clear advantages of our BHI scaffold over conventional scaffolds: i) wide-range tunable biodegradation; ii) enhanced ECM-protein binding affinity; iii) highly efficient drug loading with sustained drug delivery capability; and iv) innovative magnetic resonance imaging (MRI)-based drug release monitoring and tracking of transplanted stem cells (Fig. 2). Taking advantage of their biodegradability, and incorporating their unique physiochemical properties into stem cell-based tissue engineering, we further used our MnO2 nanosheets-assembled 3D hybrid nanoscaffolds to better regulate stem cell adhesion, differentiation into neurons, and neurite outgrowth in vitro and for enhanced stem cell transplantation in vivo (Fig. 2). With these comprehensive results, we successfully demonstrate the potential of our biodegradable MnO2 nanoscaffold as a powerful tool for improving stem cell transplantation and advancing stem cell therapy to overcome the difficulties of generating a robust population of functional neurons and enhancing neuronal behaviors (neurite outgrowth and axon regeneration) in vivo. Collectively, our developed hybrid nanoscaffold-based approach to control stem cell behavior in vivo would provide an alternative to help overcome the critical barriers that limit cell therapy for many devastating disease and injuries. 

REFERENCES

1  Kim, T. H. et al. Controlling Differentiation of Adipose-Derived Stem Cells Using Combinatorial Graphene Hybrid-Pattern Arrays. ACS Nano 9, 3780-3790, (2015).

2  Shah, S. et al. Guiding stem cell differentiation into oligodendrocytes using graphene-nanofiber hybrid scaffolds. Adv. Mater. 26, 3673-3680, (2014).

3  Solanki, A. et al. Axonal alignment and enhanced neuronal differentiation of neural stem cells on graphene‐nanoparticle hybrid structures. Advanced Materials 25, 5477-5482, (2013).

4  Novoselov, K. S. et al. A roadmap for graphene. Nature 490, 192, (2012).

5  Yin, P. T., Shah, S., Chhowalla, M. & Lee, K.-B. Design, synthesis, and characterization of graphene–nanoparticle hybrid materials for bioapplications. Chemical reviews 115, 2483-2531, (2015).

6  Chen, Y. et al. Break-up of Two-Dimensional MnO2 Nanosheets Promotes Ultrasensitive pH-Triggered Theranostics of Cancer. Advanced Materials 26, 7019-+, (2014).

7  Zhao, Z. L. et al. Activatable Fluorescence/MRI Bimodal Platform for Tumor Cell Imaging via MnO2 Nanosheet-Aptamer Nanoprobe. J. Am. Chem. Soc. 136, 11220-11223, (2014).

8  Yang, L. et al. A biodegradable hybrid inorganic nanoscaffold for advanced stem cell therapy. Nature Communications 9, 3147, (2018).