(722c) Paving the Way: Migrating Leader Cells Dissociate from Collectively Migrating Sheets Under Geometric Confinement

The migration of cells through tissues remains a fundamental and critical component of a wide range of biological processes including tissue repair, gastrulation, and the body’s innate immune response to pathogens. Depending on the cell type and the specific physiological context, cells traverse the body individually or as cohesive sheets of cells linked together by junctions. Single cell migration has been comprehensively investigated in vitro and in vivo in a variety of systems. In contrast, much less is known about collective migration and it is rapidly emerging as an attractive area of focus due to its many implications throughout development and disease progression. Collective migration plays a particularly critical role specifically within the context of cancer and the ability of cells to dissociate from the primary tumor in order to form metastases remains a major hallmark. Due to the fact that the specific mechanisms of tumor dissemination/invasion are poorly defined, we have designed assays to model tumor dissemination in vitro using PDMS and polyacrylamide-based microfluidic platforms to impart geometric constraints of prescribed dimensions to mimic tissue tracks and common routes of metastasis within the human body. Our data indicate that A431 squamous cell carcinoma cells transition from collectively migrating cells in the bulk monolayer to single migrating cells at a percentage that is proportional to the increasing degree of geometric confinement imposed by the walls of our microchannels. We hypothesized that this transition from collective to single migration modes could be due, in part, to a breakdown of the adherens junction that cohesively links cells together. Indeed, when the transmembrane protein E-cadherin is overexpressed, cells undergo much less single cell dissociation as compared to control cells which highlights the important role of E-cadherin in maintaining epithelial tissue integrity. Actomyosin contractility has also been shown to be an indispensable component of adherens junction formation and maintenance. Combining live cell imaging, confocal microscopy, Fluorescence Lifetime Imaging (FLIM), and lentiviral gene knockdowns, we have delineated a pathway mediated through cell contractility that facilitates the dissociation of cells from collectively migrating strands. In parallel, we are exploring the roles of intracellular tension, traction forces, and cell-cell signaling mechanisms that may regulate the activity or localization of the cells intrinsic contractile machinery to regulate the dissociation of cells transmitted to the adherens junctions in confining microchannels.