Brain-on-chip (BOC) technology such as nanogrooves and microtunnel structures can advance in vitro neuronal models by providing a platform with better means to maintain, manipulate and analyze neuronal cell cultures. Specifically, nanogrooves have been shown to influence neuronal differentiation, notably the neurite length and neurite direction. Here, we have drawn new results from our experiments using both 2D and 3D neuronal cell culture implementing both flat and nanogrooved substrates. These are used to show a comparison between the number of cells and neurite length as a first indicator for valuable insights into baseline values and expectations that can be generated from these experiments toward design optimization and predictive value of the technology in our BOC toolbox. Also, as a new step toward neuronal cell models with multiple compartmentalized neuronal cell type regions, we fabricated microtunnel devices bonded to both flat and nanogrooved substrates to assess their compatibility with neuronal cell culture. Our results show that with the current experimental protocols using SH-SY5Y cells, we can expect 200 – 400 cells with a total neurite length of approximately 4,000–5,000 μm per 1 mm2 within our BOC devices, with a lower total neurite length for 3D neuronal cell cultures on flat substrates only. There is a statistically significant difference in total neurite length between 2D cell culture on nanogrooved substrates versus 3D cell culture on flat substrates. As extension of our current BOC toolbox for which these indicative parameters would be used, the microtunnel devices show that culture of SH-SY5Y was feasible, though a limited number of neurites extended into microtunnels away from the cell bodies, regardless of using nanogrooved or flat substrates. This shows that the novel combination of microtunnel devices with nanogrooves can be implemented toward neuronal cell cultures, with future improvements to be performed to ensure neurites extend beyond the confines of the wells between the microtunnels. Overall, these results will aid toward creating more robust BOC platforms with improved predictive value. In turn, this can be used to better understand the brain and brain diseases.
We thank the members of the Microfab/lab at the Eindhoven University of Technology for their experimental support. We especially thank Gülden Akçay for her contributions to the acquisition of part of the experimental data for the SH-SY5Y cells in microtunnel devices. Funding. This work was financially supported by the European Research Council (ERC; STG-grant no. 280281 and PoC-grant no. 713732), the FET Proactive CONNECT project (grant no. 824070), the Health∼Holland (grant no. LSHM19006), and the Eindhoven University of Technology.
|Horizon 2020 Framework Programme
|H2020 Future and Emerging Technologies
|European Research Council
|Eindhoven University of Technology
- microtunnel structures
- neurite length
- neuronal differentiation
- SH-SY5Y cells