Background

Directed and large-scale collective cell migration underlies key multicellular processes such as morphogenesis, wound healing, immune responses, and cancer progression. A tool allowing us to shepherd such migration would enable new possibilities across cell biology and biomedical engineering. The ideal requirements for such a tool are to a) be applicable across cell and tissue types, and b) allow for interactive spatiotemporal control. Up to now, no tools existed to interactively guide cell migration. Different pieces of evidence for a long time have pointed towards electrochemical cues as being the foundation for building cellular herding systems. It is now well-established that a) electric fields are natural, emergent responses to ionic imbalances that arise in vivo during morphogenesis, regeneration, and pathogenesis; b) cells transduce direct current (DC) electrical cues into navigational cues and migrate along the field gradient in a process called ‘electrotaxis’ or ‘galvanotaxis’ and c) electrotaxis is widespread across diverse systems including at least 20 diverse mammalian cell types. The electrotactic response is thought to derive from field-induced aggregation of as yet unknown membrane receptors that induce front-rear migratory polarity. Altogether, electrotaxis has exciting potential as a tool to manipulate cell migration, and as electric fields can be harnessed to direct cellular migration, modern electronic tools and approaches will likely enable unprecedented control over tissue dynamics. In their work, Zajdel et al present a new multi-electrode electro-bioreactor, SCHEEPDOG (Spatiotemporal cellular Herding with Electrochemical Potentials to Dynamically Orient Galvanotaxis), that harnesses electrotaxis by integrating multiple independent electrodes under computer control to dynamically program electric field patterns, and steer cell migration.

Key findings and developments

• In their work, Zajdel et al describe the design of the SCHEEPDOG system and validate it by programming large in vitrotissues to undergo complex migratory manouvers on-command (Figure 1, adapted from Zajdel et al 2019 (1)).

SCHEEPDOG design

• A limitation of all electrotaxis chamber designs is the use of a single anode/cathode pair to deliver a DC electric field to cells – this limits migration to a single axis of motion. SCHEEPDOG is unique, as it possesses 4 electrodes under continuous computer control to allow for true spatiotemporal programming of the electric field geometry in 2D.
• SCHEEPDOG is essentially a bioelectric flow chamber that incorporates three modules: 1) bioreactor architecture; 2) life support; and 3) dynamically programmable electric field generation:
• Bioreactor Architecture: the bioreactor housing is comprised of the culture substrate, cells, and layer-based microfluidic assembly.
• Life support: this module aims at accommodating cell/tissue metabolic needs and preventing detrimental electrochemical effects on migrating cells.
• Dynamically programmable electric field generation: Special care was taken to ensure field uniformity within a large stimulation zone to improve throughput. SCHEEPDOG uses closed-loop feedback to ensure a temporally stable field and monitors the channel voltage with a pair of probing electrodes.

 

Validation

Inducing collective cell migration

• The authors selected a 90° turn as an archetypal complex maneuver to validate bi-axial, programmable control over directed cell migration. The MDCK (Madin-Darby Canine Kidney) epithelium was selected as the initial model system.
• They developed a universal electrical stimulation scheme capable of programming arbitrary 2D migration maneuvers such as precise angular turns and directing cells to migrate in a complete circle.
• 10,000 single cells were tracked, and the trajectories were overlaid to capture the spread of individual cell responses. While the trajectories showed cell-to-cell variation, a clear L-shaped collective cell migration was identified.
• Altogether, these experiments validated that SCHEEPDOG can deliver independent X- and Y-axis commands to tissues and that cells can respond to dynamic, orthogonal migration cues, highlighting the surprising plasticity of large scale collective migration.

Electrotactic performance parameters: differences between cell types

• The authors then went on to compare electrotactic control in different cell types. For this, they used primary skin keratinocytes collected from neonatal mice, cultured in basal epidermal growth media.
• They found that MDCKs require nearly 70% more time than keratinocytes to reorient during a 90° turn. They hypothesize that this may reflect underlying differences in tissue mechanics (e.g. cell-cell and cell-substrate adhesions), and signal transduction.

Large scale tissue translation

• The authors then evaluated boundary outgrowth, whereby they tracked the leading edges of MDCK and keratinocyte monolayers undergoing electrotaxis. They found significant differences between both cell types tested: electrotaxing MDCK cells undergo behaviour akin to supra-cellular migration, while electrotaxing keratinocyte ensembles migrate more akin to ‘marching in formation’.

Universal migration control scheme based on electrotactic timescale

• The authors generated a control scheme with a fixed field strength, and then altered the relative duration of X-axis commands versus Y-axis commands.
• Cells appeared to migrate smoothly along a 45° trajectory despite only being stimulated ‘right’ or ‘up’ in quick succession. This seemingly confirms the hypothesis that cells effectively time-average the electrotactic commands and thus perceive a virtual command direction.
• This data implies that the field-sensing mechanism of electrotaxis must operate on a timescale far faster than the migration response.

Herding cells in a closed circle

• To explore the capabilities and limits of control offered by SCHEEPDOG, the authors programmed a complex maneuver into a keratinocyte monolayer-a closed circle, which requires continuously adjusted stimulation commands.
• Cells successfully tracked this continuously shifting electric field, completing a circular maneuver with an average perimeter of over 300um.
• Angular histograms during an 8-hour period show that a) there is consistent tracking of command vectors and b) there is a persistent lag between the command vector and the direction of average cell migration.
• Altogether, this work is the first to achieve a tissue obeying a prescribed, continuously varying 2D migrational cue.

 

What I like about this paper

I like that it identifies and addresses a missing tool for a very broad field, which is cell migration. As stated by the authors, it has been more than a century that the electrotactic properties of cells were described, yet a tool based on this property had not yet been developed to generate mass cell migration on command. I like the fact that the authors make SCHEEPDOG’s thorough design and validation data available to the field, and finally, I like that they put special emphasis on fabricating SCHEEPDOG to require only benchtop rapid prototyping. All things together, this work is in alignment with something I am fully supportive of which is open science.

 

Open questions

*Questions with author responses are found at the bottom of the page.

1. You mention in your introduction the comparative advantages of SCHEEPDOG with tools such as micropatterned proteins, surface topographies, chemotaxis, and optogenetics. Ultimately, cell migration in a living tissue depends on various factors. Can you use SCHEEPDOG in combination with other forms of migratory stimulation, to study the effect of each (i.e. electrical, chemical, structural) components of migration?
2. As I understand, the cells you use here are adherent cells. What can one expect of SCHEEPDOG, when using non-adherent cells such as erythrocytes, or free-moving cells such as ciliates or flagellates?
3. To what extent can you couple SCHEEPDOG for the study of cell biology at sub-cellular resolution, for instance, changes in nuclear positioning, changes in actin cytoskeleton, etc. during the induction of different types of cell migration?
4. Can you couple SCHEEPDOG to technologies such as organs on chip, to study cell migration of co-cultures of cells, resembling a physiological situation?
5. You mention in your discussion, you see SCHEEPDOG as a tool that allows to modify maneuver geometry and timescales, to isolate signalling, transcription and mechanical processes during migration. Can you expand slightly on what you envisage for the long term as SCHEEPDOG’s contribution to multiple fields?

Reference

1. Zajdel TJ, Shim G, Wang L, Rosello-Martinez A, Cohen DJ, On-demand spatiotemporal programming of collective cell migration via bioelectric stimulation, bioRxiv, 2019.

Acknowledgements

Many thanks to Daniel Cohen and his group, for their time and engagement, and to Mate Palfy for his helpful feedback.

 

doi: https://doi.org/10.1242/prelights.17093

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