The impact of target stiffness on NK cell activation

Abstract

Abstract A diverse number of cell functions and behaviour are regulated by the mechanical properties of surrounding tissue. In addition, mechanical forces have been shown to impact synapse formation and signalling by T and B cells. Here, we set out to establish whether or not target cell tension impacts NK cell responses. To address this, human primary NK cells were placed on polyacrylamide substrates of varying rigidity coated with activating antibodies against NKp30 and LFA-1. NK cell activation measured by IFNγ secretion and degranulation were reduced on softer substrates. To better model the 3D interaction between NK cells and target cells, we synthesised sodium alginate cell-sized microbeads crosslinked with calcium chloride, where the inherent stiffness of the beads was controlled to be low (2-15kPa), medium (25-50kPa), or high stiffness (75-400kPa). Beads were coated with increasing concentrations of antibody to create surrogate targets with specific ligands and controlled stiffness. NK cells conjugated to stiff beads coated with mAb against NKp30, showed greater actin clustering, as well as perforin and MTOC polarisation towards the bead, to a far greater extent than those conjugated to softer beads. Thus, gel microbeads can be used to characterise the mechanosensitivity of immune synapses in 3D, and establish a role for target stiffness in NK cell activation.