Collective epithelial cell migration in response to two-dimensional geometrical extracellular matrix patterns

Abstract

Collective cell migration is a fundamental process in various physiological phenomena, including morphogenesis and wound healing. Failure to restore tissue integrity can lead to chronic inflammation and impact a wide range of medical practices such as surgery and trauma management, while abnormal cell migration is associated with numerous diseases, including cancer. In contrast to individual cells that undergo random movement, collective cells exhibit organized migration patterns influenced by their biophysical and biochemical interactions with neighbouring cells and the environment. Moreover, the geometrical extracellular matrix (ECM) patterns on substrates for two-dimensional epithelial cell cultures significantly influence the collective migration mode during the invasion of free spaces or the recovery of the discontinuities in cell monolayers.

One critical mechanism identified by researchers for closing non-cell-adhesive gaps is the accumulation of actin cables around concave edges and the resulting purse-string constriction. However, the studies to date have not separated the gap-edge curvature effect from the gap size effect. Here, we fabricate micropatterned hydrogel substrates with long, straight, and wavy non-cell-adhesive stripes of different gap widths to investigate the stripe edge curvature and stripe width effects on the reepithelialization of Madin–Darby canine kidney (MDCK) cells. Our results show that MDCK cell reepithelialization is closely regulated by the gap geometry and may occur through different pathways. Healing of narrow straight stripe is driven by dynamic purse-string contraction at the ends, which involves actin-cable assembly-disassembly cycles throughout the gap closure process. Healing of wavy stripes, however, involves gap bridging either via cell protrusion or by lamellipodium extension as critical cellular and molecular mechanisms in addition to purse-string contraction. We have identified the following necessary/sufficient conditions for efficient gap closure: cell migration in the direction perpendicular to wound front, sufficiently small gap size to allow bridging, and sufficiently high negative curvature at cell bridges for actin cable constriction. Our experiments demonstrate that straight stripes rarely induce cell migration perpendicular to wound front, but wavy stripes do; cell protrusion and lamellipodia extension can help establish bridges over gaps of about five times the cell size, but not significantly beyond. Such discoveries deepen our understanding of mechanobiology of cell responses to curvature and help guide development of biophysical strategies for tissue repair, plastic surgery, and better wound management.

To describe the collective cell migration mode, we employ particle image velocimetry (PIV) analysis to derive the velocity vector field. Furthermore, we utilize the spatial correlation function, widely used in the active matters community to determine the domain size of correlated cell motions, to evaluate the migration patterns. However, several definitions of the correlation function are employed in the field of biomechanics which are, sometimes, mutually inconsistent, leading to ambiguity in interpreting the results. We carry out a systematic, comprehensive investigation to evaluate the correlation functions and correlation lengths of several representative patterns of cell migration and alignment through their respective PIV vector fields and orientation director fields. By comparing the results obtained using different definitions of correlation functions, we propose a recommended definition of correlation function and correlation length. This contribution aims to provide the broader biophysical community with a comprehensive understanding of correlations in collective cell behaviour.

Our research on collective cell migration reveals the mechanobiology of collective cell responses to geometrical confinement and provide insights for interpretating correlation functions in the field of biomechanics. Specifically, we have observed a curvature-induced vortex cell migration pattern that promotes gap bridging and accelerates the wound healing process. These biomechanical strategies have potential applications in tissue repair, plastic surgery, and wound management. Additionally, our study provides perspectives on spacial correlation function and correlation length as means to evaluate collective cell migration behaviours.

Moving forward, we plan to introduce mechanical stretch forces to our current system and investigate how collective cell motions respond to pre-loading and post-loading forces. This study aims to enhance our understanding of the role of physical forces in regulating cellular physiology.