Technical Advance: Changes in neutrophil migration patterns upon contact with platelets in a microfluidic assay

Abstract

Neutrophils are traditionally regarded as the “first responders” of the immune system. However, recent observations revealed that platelets often respond earlier to recruit and activate neutrophils within sites of injury and inflammation. Currently, platelet–neutrophil interactions are studied by intravital microscopy. Although such studies provide exceptional, physiologic in vivo data, they are also laborious and have low throughput. To accelerate platelet–neutrophil interaction studies, we have developed and optimized an ex vivo microfluidic platform with which the interactions between platelets and moving neutrophils are measured at single-cell level in precise conditions and with high throughput. With the use of this new assay, we have evaluated changes in neutrophil motility upon direct contact with platelets. Motility changes include longer distances traveled, frequent changes in direction, and faster neutrophil velocities compared with a standard motility response to chemoattractant fMLP. We also found that the neutrophil–platelet direct interactions are transient and mediated by CD62P–CD162 interactions, localized predominantly at the uropod of moving neutrophils. This “crawling,” oscillatory neutrophil behavior upon platelet contact is consistent with previous in vivo studies and validates the use of this new test for the exploration of this interactive relationship.