Directed neutrophil migration in blood vessels and tissues is critical for proper immune function; however, the mechanisms that regulate three-dimensional neutrophil chemotaxis remain unclear. It has been shown that integrins are dispensable for interstitial three-dimensional (3D) leukocyte migration; however, the role of integrin regulatory proteins during directed neutrophil migration is not known. Using a novel microfluidic gradient generator amenable to 2D and 3D analysis, we found that the integrin regulatory proteins Kindlin-3, RIAM, and talin-1 differentially regulate neutrophil polarization and directed migration to gradients of chemoattractant in 2D versus 3D. Both talin-1-deficient and RIAM-deficient neutrophil-like cells had impaired adhesion, polarization, and migration on 2D surfaces whereas in 3D the cells polarized but had impaired 3D chemotactic velocity. Kindlin-3 deficient cells were able to polarize and migrate on 2D surfaces but had impaired directionality. In a 3D environment, Kindlin-3 deficient cells displayed efficient chemotaxis. These findings demonstrate that the role of integrin regulatory proteins in cell polarity and directed migration can be different in 2D and 3D.
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This work was supported by National Institutes of Health Grant RO1EB010039 (to A. H. and D. J. B.), AHA Grant 10POST3230031 (to P. J. C.), and University of Wisconsin Carbone Cancer Center Cancer Center, Support Grant P30 CA014520 (to D.J.B).
David J. Beebe holds equity in Bellbrook Labs, LLC, Tasso, Inc., and Salus Discovery, LLC. E. Berthier holds equity in Salus Discovery, LLC and Tasso, Inc. P. Cavnar holds equity in Salus Discovery, LLC.
YY, PC, EB, DB and AH designed the studies, analyzed data, and wrote the manuscript. YY, DAB, PC, EB performed experiments and analyzed data.
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