The phosphoinositide 3-kinase (PI3K) catalytic subunit p110δ is expressed in neutrophils and is thought to play a role in their accumulation at sites of inflammation by contributing to chemoattractant-directed migration. We report here that p110δ is present in endothelial cells and participates in neutrophil trafficking by modulating the proadhesive state of these cells in response to tumor necrosis factor α (TNFα). Specifically, administration of the selective inhibitor of PI3Kδ, IC87114, to animals reduced neutrophil tethering to and increased rolling velocities on cytokine-activated microvessels in a manner similar to that observed in mice deficient in p110δ. These results were confirmed in vitro as inhibition of this isoform in endothelium, but not neutrophils, diminished cell attachment in flow. A role for PI3Kδ in TNFα-induced signaling is demonstrated by a reduction in Akt-phosphorylation and phosphatidylinositol-dependent kinase 1 (PDK1) enzyme activity upon treatment of this cell type with IC87114. p110δ expressed in neutrophils also contributes to trafficking as demonstrated by the impaired movement of these cells across inflamed venules in animals in which this catalytic subunit was blocked or genetically deleted, results corroborated in transwell migration assays. Thus, PI3Kδ may be a reasonable therapeutic target in specific inflammatory conditions as blockade of its activity reduces neutrophil influx into tissues by diminishing their attachment to and migration across vascular endothelium. (Blood. 2004;103:3448-3456)