Nanoscale Mechanosensing of Natural Killer Cells is Revealed by Antigen‐Functionalized Nanowires

Abstract

function of cells. [8] These forces have different origins, such as actin dynamics, [9] and play important roles at different stages of the lymphocyte immune activity. Initial sampling of antigens on the surface of antigen presenting cells (APCs), as well as activation of immunoreceptors, strongly depends on actin polymerization and dynamics. [10] Moreover, immunoreceptors recognize antigens under mechanical load to discriminate between high-affinity and low-affinity antigens. [11] Once activated, the receptor-antigen complexes on the lymphocyte-APC interface are driven by retrograde actin flow and myosin contraction into highly regulated structures termed immune synapse, whose forces affect the inside-out signaling of lymphocytes. Today, mechanical forces in immune system are a subject of emerging research, which has so far mostly focused on T cells and B cells. [12,13] Studying mechanical forces in cells is challenging, because these forces have relatively low magnitude -mostly at the nanoNewton scale, and often span over miniature regions sized down to the molecular scale. Existing tools include optical traps, [14,15] micropipettes, [16] and atomic force microscopy (AFM), [17,18] which, however, apply and detect forces only at single point on the cell membrane, and do not overview the mechanical behavior of the entire cell. Alternatively, traction force microscopy, which determines the displacement of microbeads embedded in hydrogel surface for cell spreading, maps forces of entire cells, [19][20][21] however, it can hardly detect the exact bead movement since the beads are distributed randomly, and their resting position is unknown. Furthermore, analysis of bead movement requires complex force calculations based on elasticity theory. [22] These constrains can be overcome by elastomeric micropillars for cell spreading, which allow facile mapping of force distribution within cells. [23] Furthermore, micropillars can be functionalized with biomolecules that yield chemical stimuli for various cell functions, such as adhesion [24,25] or immune response, [26] and thereby allow integration of mechanical and biochemical cues. However, the advantages of elastic micropillars come at the expense of their spatial and mechanical resolution. Indeed, poly(dimethyl siloxane) (PDMS) -material of choice for micropillar fabrication -is limited for the fabrication of pillars with micrometerscale size and aspect ratio of 3:1, for which sensing forces below Cells sense their environment by transducing mechanical stimuli into biochemical signals. Commonly used tools to study cell mechanosensing provide limited spatial and force resolution. Here, a novel nanowire-based platform for monitoring cell forces is reported. Nanowires are functionalized with ligands for cell immunoreceptors, and they are used to explore the mechanosensitivity of natural killer (NK) cells. In particular, it is found that NK cells apply centripetal forces to nanowires, and that the nanowires stimulate cell contraction. Based on the nanowire deformation, it is...

Mechanical Regulation of the Cytotoxic Activity of Natural Killer Cells

“…On the other hand, on the control surfaces, the effect of surface stiffness on the CD107a expression was significantly less pronounced. This result largely mirrors our previous report on the activation of NK cells on nanowires, where NK cells showed significant activation only on nanowires functionalized with MICA, while control surfaces lacking either nanowires, MICA, or both, did not induce a substantial activation of NK cells (23).…”

“…Early indication that mechanical forces also play a role in the regulation of NK cell immune activity was recently provided by Barda-Saad and Co, who showed that actin retrograde flow controls NK cell immune response via conformational changes of tyrosine phosphatase SHP-1 (22). Shortly after, we reported that NK cells stimulated onto vertical nanowires functionalized with activating ligands produced an enhanced immune response (23). In that study, the nanowiresnanomechanical objects with ultra-high aspect ratio and flexibilityexhibited extraordinary compliance to cellular forces as small as 10pN.…”

Advanced Materials and Devices for the Regulation and Study of NK Cells

“…The first evidence of mechanosensing in NK cells was recently provided by Barda-Saad and co., who showed that NK cell response is mediated by actomyosin retrograde flow [97]. Shortly afterwards, the authors of the present review provided a direct evidence for mechanosensing of NK cells [90]. For this purpose, we developed a novel platform for the study of the cell mechanical activity, which was based on nanowires—quasi 1D nanostructures whose aspect ratio can exceed a few orders of magnitude.…”

“…It should be noted that detecting mechanical forces in cells is challenging because these forces span over a small, often nanometric length scale, and have very low magnitude down to picoNewtons. Optical traps [80,81], micropipettes [82], and atomic force microscopy [89,90] have been used detect such forces, however they do so only at a single point of the cell membrane. Traction force microscopy based on measuring the dislocation of fluorescent beads in a hydrogel can map forces of an entire cell [91,92,93], however, the exact bead movement is hard to assess since the beads are distributed randomly, and their resting position is unknown.…”

Lymphocyte mechanotransduction: The regulatory role of cytoskeletal dynamics in signaling cascades and effector functions

“…Furthermore, it is necessary to explore whether other signaling molecules, and specifically inhibitory phosphatases, such as SHP‐2 or SHIP‐1, are also recruited to the cytoskeletal machinery and are impacted by actin dynamics during different NK cell responses. A recent study shed novel light on some of these issues by utilizing a nanowire based system coated with the NKG2D ligand MICA to measure force induced by NK cells, and found that NK cells displace a single nanowire with minimal force of 10 pN . This study showed that similarly to T‐cell mechanotransduction, the actin cytoskeleton was actively concentrated in areas of force exertion on the nanowires.…”