Our studies show that normal blood lymphocytes and monocytes respond to a steady electric field in transwell assays. All lymphocyte subsets examined, including naïve and memory CD4, CD8 T cells, and B cells migrated towards the cathode, as did monocytes. Electrotactic migration is highly directional: that in studies monitoring the migratory path of memory T cells on fibronectin-coated glass, almost all cells migrated cathodally, and the directionality index is comparable to or higher than the optimal directionality index observed in comparable chemotaxis assays. We also confirm that electric field exposure induces Erk1/2 and Akt activation in T cells, consistent with activation of the MAP Kinase and PI3 Kinase signaling pathways implicated in coordinated cell motility. Finally, we show that an applied electric field induced the electrotactic migration of endogenous lymphocytes in mouse skin. Our results thus define electrotaxis as potentially an additional mechanism for the control of lymphocyte and monocyte migration.
Electrotaxis of eukaryotic cells has been reported in a number of studies (4
). Particularly, electrotaxis of epithelial cells plays an important role in controlling wound healing (i.e. the wound generates an inward DC electric field and electrotaxis of epithelial cells toward the cathode of the electric field helps closing the wound) (4
). Our results show that memory T cells migrate toward the cathode of an electric field applied with strength similar to wound generated electric field. Therefore, electrotaxis of lymphocytes and other immune cells may be involved in the process of wound healing as well. Furthermore, our study shows that immune cell positioning can be regulated in vivo by externally applied electric fields.
An interesting aspect of our results is the uniformity in the electrotactic responses of different mononuclear leukocyte subsets as assessed in the transwell assay. This contrasts with responses to known chemoattractants, whose cell surface receptors are differentially expressed and direct subset-selective migration. The uniform migration of circulating lymphocytes suggests that other leukocyte subsets (e.g. tissue memory cells) may undergo electrotaxis as well. On the other hand, not all GFP+ cells in mouse skin responded: rather the electrotactic responses appeared limited to cells that were intrinsically motile, i.e. cells that displayed random migration prior to field application. Thus, leukocyte responses to electric potential gradients may be regulated primarily by the migratory state of competence of the cells, rather than (as in the case of chemotactic response) by subset specificity among migratory populations.
Indeed, as suggested by the diverse cell types that respond, from Dictyostelium amoebae to epithelial cells, metastatic cancer cells and lymphocytes, electrotaxis may be a nearly universal competency of motile cells (4
). Regulation of electrotaxis may be possible in terms of directionality, however. Although lymphocytes, like many other mammalian cells, undergo cathode-directed electrotaxis, some cells (e.g. corneal endothelial cells, human vascular endothelial cells) migrate preferentially to the anode. Given the absence of subset selectivity in lymphocyte responses, however, it seems unlikely that electric fields operate in vivo to control the recruitment of lymphocytes from the blood or direct their microenvironmental interactions during physiologic immune responses. Instead, physiologic electric fields may serve to support cellular translocation events in settings where cell type specificity is not a critical issue. One example would be the bulk cell population movements that occur early in embryogenesis, when the principal distinction is between motile and non-motile undifferentiated cells; indeed the role of electric fields in cell movement was implicated in studies of early embryogenesis (4
). A more pertinent example is the cellular response to a penetrating wound, during which the rapid recruitment of any motile cell (fibroblasts, macrophages and other immune cells) from the immediately surrounding tissue would seem beneficial or at least not deleterious. As mentioned in the introduction, epithelial surfaces such as the skin are well documented batteries, maintaining an electrical potential gradient that is short circuited upon disruption of the epithelial barrier. Wounds in bovine cornea or in guinea pig and human skin, for example, generate local electric fields of ~0.4–1.4V cm−1
with the wound itself being the cathode. This field is well within the range for efficient electrotactic recruitment of lymphocytes and other motile cells into the wound. While in the case of lymphocytes and other immune cells it may be difficult in such a setting to isolate the effects of electrotaxis from those of the chemoattractants induced, our studies clearly show that lymphocytic cells resident in skin respond with directional migration when presented with an applied electric field.
Previous studies have found that electric fields activate intracellular signaling processes including kinase pathways involved in cellular motility. We have confirmed here that lymphocytes respond to electric fields with activation of Erk kinases and Akt, which are involved in chemoattractant receptor signaling and in electrotactic signaling in other cells (16
). Activation of these pathways suggests that electrotaxis and chemotaxis engage common intracellular cell motility programs in responding lymphocytes. On the other hand, in keeping with findings in Dictyostelium amoebae in which electric fields did not generate a rapid burst of calcium signaling (but only a slow, prolonged calcium response at the population level), (32
), we did not observe a robust, immediate calcium flux in lymphocytes in response to electrical stimulation, and delayed calcium elevation assessed by flow cytometric analysis of Fluo-4 labeled cells was variable (data not shown). The kinetics of MAP kinase and Akt signaling also distinguishes the electric field response, since phosphorylation of Erk and Akt was significant after 60 minutes, but was not appreciable by phospho-flow analyses at 1 or 10 minutes, contrasting with the rapid triggering of signaling cascades by chemoattractant receptors. In conclusion, our study demonstrates electrotaxis of lymphocytes, and shows that electric fields can direct lymphocyte migration in vitro and in vivo. Electrotaxis represents an additional mechanism for the control of leukocyte migration. It is likely to play a role in sites of epithelial injury, and may permit novel approaches for manipulating the positioning of lymphocytes and other immunocytes to enhance vaccine or anti-tumor responses.