Three-dimensional cell culture can be regulated by vibration: low-frequency vibration increases the size of olfactory...

Olfactory ensheathing cell (OEC) transplantation is emerging as a promising therapy for spinal cord injuries. However, outcomes are inconsistent, and the method needs improvement. Currently, cells are injected into the injury site as a suspension, and often fail to form a three-dimensional (3D) network crucial for both survival of the transplanted cells, and for regeneration of severed axons. 3D culture systems are therefore likely to improve the method. Of the many 3D culture systems available, the spheroid-producing naked liquid marble (NLM) technique is particularly advantageous compared to other platforms as it rapidly generates cell spheroids which can easily be extracted for further handling. To improve production of the spheroids, we designed and tested a device which allows fine control over vibrational stimuli to liquid marble cell cultures. We applied vibrational frequencies of 20, 60, and 80 Hz with consistent amplitude to NLM containing OECs and assessed the size and number of the 3D cell spheroids generated as well as the migratory capacity of cells cultured in the vibrated spheroids.

Vibrating the NLMs led to fewer and dramatically larger spheroids in comparison to non-vibrated NLMs. Of the frequencies tested, 60 Hz caused over 70-fold increase in spheroid volume. When transferred to a culture plate, the larger spheroids retained their structure after 72 h in culture, and cells that migrated out of the spheroids covered a significantly larger area compared to cells migrating out of spheroids formed at all the other frequencies tested.

We have shown that vibration can be used to regulate the formation of cell spheroids in NLM cultures. The ability to modulate the size of spheroids is useful for a range of 3D cell culture models and for preparing cells for in vivo transplantation.