Effect of pharmacological modulation of actin and myosin on collective cell electrotaxis

Abstract

Electrotaxis-the directional migration of cells in response to an electric field-is most evident in multicellular collectives and plays an important role in physiological contexts. While most cell types respond to applied electric fields of the order of a Volt per centimeter, our knowledge of the factors influencing this response is limited. This is especially true for collective cell electrotaxis, in which the subcellular migration response within a cell has to be coordinated with coupled neighboring cells. Here, we investigated the effect of the level of actin cytoskeleton polymerization and myosin activity on collective cell electrotaxis of Madin-Darby Canine Kidney (MDCK) cells in response to a weak electric field of physiologically relevant magnitude. We modulated the polymerization state of the actin cytoskeleton using the depolymerizing agent cytochalasin D or the polymerizing agent jasplakinolide. We also modulated the contractility of the cell using the myosin motor inhibitor blebbistatin or the phosphatase inhibitor calyculin A. While all the above pharmacological treatments altered cell speed to various extents, we found that only increasing the contractility and a high level of increase/stabilization of polymerized actin had a strong inhibitory effect specifically on the directedness of collective cell electrotaxis. On the other hand, even as the effect of the actin modulators on collective cell migration was varied, most conditions of actin and myosin pharmacological modulation-except for high level of actin polymerization/stabilization-resulted in cell speeds that were similar in the absence or presence of the electric field. Our results led us to speculate that the applied electric field may largely impact the cellular apparatus specifying the polarity of collective cell migration, rather than the functioning of the migratory apparatus. Bioelectromagnetics. 39:289-298, 2018. © 2018 Wiley Periodicals, Inc.

SCHEEPDOG: programming electric cues to dynamically herd large-scale cell migration

“…We selected a 90°turn as an archetypal complex maneuver to validate bi-axial, programmable control over directed cell migration. Here, we selected the MDCK epithelium as our initial model system as it is a standard model both for studying large-scale, collective cell migration and for collective electrotaxis in 1D (Cohen et al, 2014;Bashirzadeh et al, 2018;Li et al, 2012). We patterned 5x5 mm MDCK monolayers into SCHEEPDOG, then used automated phase-contrast microscopy to capture 1 h of control data (field OFF) before stimulating at~2 V cm −1 for 2 h along the X-axis ('right') followed by 2 h along the Y-axis ('up').…”

“…optogenetics). Further, the vast majority of electrotaxis studies focus on single cells due to experimental complexities, and the few collective-level electrotaxis studies (Li et al, 2012;Cohen et al, 2014;Lalli and Asthagiri, 2015;Bashirzadeh et al, 2018;Cho et al, 2018) have focused on elucidating biological or biophysical questions rather than advancing the infrastructure or exploring the limits of the phenomenon. We believe that modernized bioelectric interfaces will make the field far more approachable and substantially increase the utility of electrotaxis itself, bringing us much closer to being able to deploy electrotaxis as a useful tool to direct cell migration for both fundamental migration research and applied contexts such as expedited healing.…”

SCHEEPDOG: Programming Electric Cues to Dynamically Herd Large-Scale Cell Migration

“…To capture a range of phenotypes, we tested with both the MDCK kidney epithelial cell line and primary, neonatal mouse skin keratinocytes. While both cell lines undergo electrotaxis in 1D (Nishimura et al, 1996;Li et al, 2012;Cohen et al, 2014;Guo et al, 2015;Bashirzadeh et al, 2018), primary keratinocytes cultured in basal media have weak cell-cell adhesions leading to more individualistic behavior in counterpoint to the strong cadherin-mediated cell-cell adhesions in an MDCK epithelium (Li et al, 2012). We patterned large, 5 3 5 mm monolayers of either cell type into SCHEEPDOG, then used automated phase-contrast microscopy to capture 1 h of control data (field OFF) before stimulating at 2 V cm À1 for 2 h along the x axis (''right'') followed by 2 h along the y axis (''up'').…”

“…The potential of 2D stimulation is not realized by the handful of existing dual-axis studies due to lack of programmable control of field direction (Gokoffski et al, 2019) and production of cytotoxic byproducts (Riedel-Kruse et al, 2011). Further, the vast majority of electrotaxis studies focus on single cells due to experimental complexities, while the smaller body of collective-level studies (Cooper and Keller, 1984;Nishimura et al, 1996;Zhao et al, 1996;Cao et al, 2011;Li et al, 2012;Cohen et al, 2014;Lalli and Asthagiri, 2015;Bashirzadeh et al, 2018;Cho et al, 2018;Gokoffski et al, 2019) focused on specific biological or biophysical questions rather than pushing the envelope of electrotaxis to explore both its limits and capabilities as a tool. A next-gen electro-bioreactor is key to pushing electrotaxis research forward by enabling large-scale, precise, and repeatable perturbations for cell motility research.…”

Electrotaxis of Glioblastoma and Medulloblastoma Spheroidal Aggregates

“…This suggests that the dynamics of electrotaxis-induced gene expression are likely subtler than the previously seen dynamic changes in motility machinery, as our observed gene expression changes will more likely relate to the change of a particular cue, than a complete change in machinery for a different mode of invasion. Our observation is supported by recent work by Bashirzadeh, et al ., who by using pharmacological modulation of actin and myosin on cells undergoing electrotaxis, showed that electrical fields likely induce changes in polarization and not motility 45 . Based on this evidence, we would expect not to see dramatic changes in motility machinery when the cells are motile prior to dcEF application, which is exactly what we have observed.…”