Abstract
Abstract Clustering of activating lymphocytes has been frequently observed during early stage of immune responses, but the importance of such multi-cellular clusters has not been systematically investigated due to the lack of tools. To address it, we fabricated cell-laden microwells that allowed us to precisely control multi-cellular interactionss among activating lymphocytes. The role of contact-mediated homotypic interactions between natural killer (NK) cells during IL-2 activation was examined using the cell-laden microwells. To assess the role of NK-NK homotypic interactions on the activation of NK cells, NK cells cultured in two different types of microwells, social microwells that allow contact-mediated multicellular interactions and lonesome microwells that inhibit cell-cell contact, were compared. Activation of NK cells by IL-2 was assessed by measuring phosphorylation of STAT-5, expression of CD25 and Ki-67. Immunofluorescence microscopy was performed to assess the levels of pSTAT-5, CD25, and Ki-67, and percentages of stained cells for each marker were examined and plotted. NK cells in social microwells expressed higher levels of all three markers than those in lonesome microwells, suggesting that homotypic contact-mediated interactions can enhance the activation of NK cells. Further analysis revealed that 2B4 is a key receptor mediating contact-mediated augmentation of NK cells in social miceowells by clustering and polarizing IL-2 receptor chains.
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