Abstract
Natural killer (NK) cell cytotoxicity in tissue is dependent on the ability of NK cells to migrate through the extracellular matrix (ECM) microenvironment. Traditional imaging studies of NK cell migration and cytotoxicity have utilized 2D surfaces, which do not properly reproduce the structural and mechanical cues that shape the migratory response of NK cells in vivo. Here, we have combined a microwell assay that allows long-term imaging and tracking of small, well-defined populations of NK cells with an interstitial ECM-like matrix. The assay allows for long-term imaging of NK–target cell interactions within a confined 3D volume. We found marked differences in motility between individual cells with a small fraction of the cells moving slowly and being confined to a small volume within the matrix, while other cells moved more freely. A majority of NK cells also exhibited transient variation in their motility, alternating between periods of migration arrest and movement. The assay could be used as a complement to in vivo imaging to study human NK cell heterogeneity in migration and cytotoxicity.
Highlights
Natural killer (NK) cells, a class of innate lymphoid cells (ILCs), are large granular lymphocytes that have the capacity to kill virus-infected and transformed cells
Single cell tracking and detection of cell-cell contacts was automated while accurate determination of target cell death was done by manual analysis
The studied NK cells were found to exhibit a range of different migration behaviors
Summary
Natural killer (NK) cells, a class of innate lymphoid cells (ILCs), are large granular lymphocytes that have the capacity to kill virus-infected and transformed cells. The main inhibitory signaling is provided by killer-cell immunoglobulin-like receptors (KIRs), which interact with cognate HLA class I molecules[2]. Cell migration is a fundamental ability underlying embryogenesis, immune function and wound healing. It is a highly complex, tightly regulated process involving systems of cell surface receptors, intracellular signaling pathways, the cytoskeleton, and it is highly dependent on interactions with the local extracellular microenvironment. The function of NK cells depends critically on their ability to extravasate from the circulation and migrate through the extracellular matrix (ECM) toward target tissues. A collagen-based assay where adherent target cells grown at the bottom of a culture plate were overlaid with a 3D collagen matrix containing T cells has been used to study the cytotoxic behavior of T cells[11]. ECM gels like collagen I and Matrigel have modest light scattering properties compared to tissue enabling optical imaging[13]
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