Multiple rows of cells behind an epithelial wound edge extend cryptic lamellipodia to collectively drive cell-sheet movement

Abstract

The mechanism by which epithelial, endothelial and other strongly cell-cell adhesive cells migrate collectively as continuous sheets is not clear, even though this process is crucial for embryonic development and tissue repair in virtually all multicellular animals. Wound closure in Madin-Darby canine kidney (MDCK) epithelial cell monolayers involves Rac GTPase-dependent migration of cells both at and behind the wound edge. We report here for the first time that cells behind the margin of wounded MDCK cell monolayers, even hundreds of microns from the edge, extend `cryptic' lamellipodia against the substratum beneath cells in front of them, toward the wound, as determined by confocal, two-photon and transmission electron microscopy. These so-called submarginal cells nevertheless strictly maintain their more apical cell-cell contacts when they migrate as part of a coherent cell sheet, hiding their basal protrusions from conventional microscopy. The submarginal protrusions display the hallmarks of traditional lamellipodia based on morphology and dynamics. Cells behind the margin therefore actively crawl, instead of just moving passively when cells at the margin pull on them. The rate of migration is inversely proportional to the distance from the margin, and cells move co-ordinately, yet still in part autonomously, toward the wound area. We also clarify the ancillary role played by nonprotrusive contractile actin bundles that assemble in a Rho GTPase-dependent manner at the margin after wounding. In addition, some cell proliferation occurs at a delay after wounding but does not contribute to closure. Instead, it apparently serves to replace damaged cells so that intact spread cells can revert to their normal cuboidal morphology and the original cell density of the unbroken sheet can be restored.

Collective migration of an epithelial monolayer in response to a model wound

“…The active implication of the cells several rows behind the border has been identified in the MDCK system (9,20). Indeed, these references report cells that move actively within the epithelium, in a fibroblast way, by using a so-called ''cryptic'' lamellipodium that finds its way under the other cells of the monolayer.…”

“…This migration is associated with small GTPases of the Rho family whose activation is not restricted to the border cells but extends as far as 4-10 cell rows within the epithelium (20). At the same time, as already mentioned, the epithelia keep their integrity; the cell-cell contacts are maintained in particular by cadherins (21), although it has been observed that MDCK cells migrate actively within the monolayer by developing active ''cryptic'' lamellipodia under the other cells (20). Interestingly, although the onset of migration significantly depends on the initial cell density (22), cell division does not play an active role but rather appears to redensify the monolayer (20).…”

NADPH oxidases in lung biology and pathology: Host defense enzymes, and more

“…1), which are critically involved in cell growth and differentiation and morphogenesis (13,38,52,110,111). Because the airway epithelium is continuously subjected to injury due to environmental stress, it possesses an exceptional capacity to repair itself (112,113), through a coordinated response involving rapid spreading and migration of neighboring cells, cell proliferation and redifferentiation, to restore the original mucociliary epithelium (103,114,115). Using in vitro wound models in cultured airway epithelial systems, DUOX1 was found to play a critical role in cell migration as part of the wound repair response (87,108), illustrating an additional function of DUOX1 in maintenance of epithelial integrity.…”

The role of laser fluence in cell viability, proliferation, and membrane integrity of wounded human skin fibroblasts following helium‐neon laser irradiation

“…The central scratch has been used with multiple cell types and, as the monolayers heal the wound in a characteristic manner, they have been used to study cell polarization, matrix remodeling, cell migration, and numerous other processes [12]. Kornyei et al (2000) [14] used an in vitro scratch-wound model to investigate astroglial responses to mechanical injury while Saga et al (2003) [15], Liang et al (2004) [16], Farooqui et al (2004) [17], and Walker et al (2004) [18] used the cell scratch-wound model to study the proliferative and migratory responses of different cells. Lau et al (2001) [19] focused on the production of four inflammatory cytokines in primary culture using an injury model which simulated in vivo mechanical trauma and reported that all four cytokines began to increase 1 hour post-scratch and remained at high levels throughout the experiment [19].…”