BETA(2)-INTEGRIN ENGAGEMENT TRIGGERS ACTIN POLYMERIZATION AND PHOSPHATIDYLINOSITOL TRISPHOSPHATE FORMATION IN NONADHERENT HUMAN NEUTROPHILS

Beta2 integrins are involved in the adhesion of leukocytes to other cells and surfaces. Although adhesion is required for cell locomotion, little is known regarding the way beta2 integrin-receptors affect the actin network in leukocytes. In the present study filamentous actin (F-actin) levels in non-adherent human neutrophils have been measured by phalloidin staining after antibody cross-linking Of beta2 integrins. Antibody engagement Of beta2 integrins resulted in a rapid and sustained (146 and 131% after 30 and 300 s, respectively) increase in the neutrophil F-actin content. This is in contrast to stimulation with N-formyl-l-methionyl-l-leucyl-l-phenylalanine (fMLP), which causes a prompt and pronounced but rapidly declining rise in F-actin (214 and 127% after 15 and 300 s, respectively). Priming neutrophils with 1 nM PMA, a low concentration that did not influence the F-actin content per se, increased the magnitude of the beta2 integrin-induced response but had no effect on the kinetics (199% after 30 s and 169% after 300 s). Removal of extracellular Ca2+ only marginally affected the beta2 integrin-induced F-actin response for cells that were pretreated with PMA whereas the response for nonprimed cells was reduced by half. This suggests that even though extracellular Ca2+ has a modulatory effect it is not an absolute requirement for beta2 integrin-induced actin polymerization. Beta2 integrin engagement did not affect the resting cellular level of cAMP arguing against a role of cAMP in beta2 integrin-induced actin assembly. The lack of a cAMP signal might instead explain, at least in part, the prolonged F-actin response triggered by beta2 integrins, since addition of cAMP and 1-isobutyl-methylxanthine (IBMX) caused a prompt reversal of the beta2 integrin-induced F-actin elevation in electropermeabilized neutrophils. Engagement Of beta2 integrins, as previously shown for activation of the chemotactic peptide receptor, resulted in a significant formation of PtdInsP3. The capacity Of beta2 integrins and chemotactic peptide receptors to induce phosphatidylinositol trisphosphate (PtdInsP3) formation correlated with their ability to induce actin polymerization.