Identification of sharpin as a molecule regulating leukocyte transmigration from siRNA screens (LB281)

Abstract

Migration of leukocytes into tissues is a hallmark of an acute inflammation. The migration is regulated by molecular interactions between adhesion molecules on endothelium and their counterparts on leukocytes. Here we have used siRNA‐based screens to search for new molecules regulating leukocyte transmigration through endothelium. The functional significance of the hits was confirmed using time‐lapse microscopy in in vitro flow chamber assays, in which human leukocytes were perfused with a defined shear stress over a confluent monolayer of cultured endothelial cells. In peripheral blood mononuclear cells a cytosolic protein, SHARPIN, was localized to uropods of migrating leukocytes. Moreover, using siRNA silencing we found that after the transmigration, the leukocytes without SHARPIN trapped close to transmigration site while control leukocytes moved onwards. In endothelial cells the use of siRNA to silence one ephrin type‐B receptor, reduced granulocyte transmigration through endothelium while overexpressing of it increased transmigration. As a conclusion we have found new molecules in leukocytes and in endothelium that regulate leukocyte transmigration.