Roles of microtubules, cell polarity and adhesion in electric-field-mediated motility of 3T3 fibroblasts

Abstract

Direct-current electric fields mediate motility (galvanotaxis) of many cell types. In 3T3 fibroblasts, electric fields increased the proportion, speed and cathodal directionality of motile cells. Analogous to fibroblasts' spontaneous migration, we initially hypothesized that reorientation of microtubule components modulates galvanotaxis. However, cells with intact microtubules did not reorient them in the field and cells without microtubules still migrated, albeit slowly, thus disproving the hypothesis. We next proposed that, in monolayers wounded and placed in an electric field, reorientation of microtubule organizing centers and stable, detyrosinated microtubules towards the wound edge is necessary and/or sufficient for migration. This hypothesis was negated because field exposure mediated migration of unoriented, cathode-facing cells and curtailed migration of oriented, anode-facing cells. This led us to propose that ablating microtubule detyrosination would not affect galvanotaxis. Surprisingly, preventing microtubule detyrosination increased motility speed, suggesting that detyrosination inhibits galvanotaxis. Microtubules might enhance adhesion/de-adhesion remodeling during galvanotaxis; thus, electric fields might more effectively mediate motility of cells poorly or dynamically attached to substrata. Consistent with this hypothesis, incompletely spread cells migrated more rapidly than fully spread cells. Also, overexpression of PAK4, a Cdc42-activated kinase that decreases adhesion, enhanced galvanotaxis speed, whereas its lack decreased speed. Thus, electric fields mediate fibroblast migration via participation of microtubules and adhesive components, but their participation differs from that during spontaneous motility.

Competitive Guidance Cues Affect Fibroblast Cell Alignment: Electric Fields vs. Contact Guidance

“…Cells grown on flat, non-grooved surfaces were exposed to an EF of 200mV/mm, From the cell population (n=339), 84% of cells aligned perpendicular, at angles of >800, to the EF vector, in agreement with other studies [6][7][8]. Figure 2 summarises these difference in cell alignment between cells exposed to grooves only (depth of 347 nm and width of 2 1m) and EFs only.…”

“…Another physiologically relevant guidance cue is an electric field (EF), which is present in many tissues during development and wound repair [5]. Fibroblasts respond to a d.c. EF by aligning their long axes perpendicular to the direction of the EF vector [6][7][8], and migrating towards the cathode [6,7], although a recent study showed that EFs do not direct the migration of fibroblast cells cultured on collagen I [9].…”

Microtubule regulation of corneal fibroblast morphology and mechanical activity in 3-D culture

“…However, microtubules are also known to regulate other aspects of cell behavior including cell polarization (Finkelstein, 2004;Sloboda, 1980;Watanabe, 2005). In order to assess these Rho kinase independent functions in corneal fibroblasts, we studied the effects of microtubule disruption in the presence of the specific Rho-kinase inhibitor Y-27632.…”

Harnessing the Electric Spark of Life to Cure Skin Wounds

“…Several cell types that are implicated in wound repair increase their speed and direction of migration (electrotaxis) in response to an EF, as summarized in Table 1. 14,[19][20][21][22][23][24][25][26][27][28][29][30] Along with the antibacterial effect that EFs have shown in several studies, 3 cells involved in the immune response such as neutrophils, lymphocytes, and monocytes show cathodal migration (toward the wound center) in an EF. 14,19 Granulocytes migrate toward the anode at 2.5 mM Ca 2 + and toward the cathode at 0.1 mM Ca 2 + .…”

“…Several studies have shown that fibroblasts can show both cathodal and anodal electrotactic responses. 14,23,[27][28][29][30] Additional studies on animal models and in cell cultures of dermal fibroblasts stimulated with a direct current (DC) EF demonstrated a significant increase in the ability of fibroblasts to synthesize collagen, produce DNA, and synthesize protein. In addition, the speed of epithelialization in the wound was noticeably increased.…”