Dynamic Force Generation within the Immune Synapse

Increasing evidence suggests mechanical forces modulate T cell function. In this report, we investigate forces applied by mouse CD4+ T cells onto an underlying substrate as a model of the interface between T cells and antigen presenting cells. Traction force microscopy was carried out using microfabricated arrays of elastomer (polydimethylsiloxane, Sylgard 184, PDMS) pillars; cells induce deflections of the pillars which can be measured and used to estimate force applied to each structure. We chose a pillar geometry of 1 micrometer diameter and 5-9 micrometer height. These pillars were coated with a 1:1 mix of activating antibodies to CD3 and CD28, which ligate and activate the TCR complex and costimulatory CD28 signal. Traction force microscopy carried out on mouse naive CD4+ T cells 1 hour after seeding revealed that naive cells exert forces onto these pillar arrays with magnitude on the order of 50 pN per structure. Moreover, force application by a given cell is periodic, with a cycle on the order of minutes. To investigate the physiological implications of these forces, we measured IL-2 secretion by T cells seeded onto planar PDMS substrates of varying rigidity, which was controlled by varying the ratio of base : curing agent, yielding bulk moduli of 2 MPa to 25 kPa. Substrates were coated with activating antibodies to CD3 and CD28 prior to cell seeding, and the per area concentration of antibodies varied less than 10% across the different moduli. Activation of naive T cells, measured as IL-2 secretion over six hours, was 50% greater on the stiffest vs. softest elastomers, and each condition was statistically different from all others (Kruskal-Wallis methods, alpha = 0.05). These results demonstrate a functional impact of mechanical forces on T cell activation, and reveal new dynamics of the immune synapse.