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 ce lls and surfaces. Although adhesion is required for cell locomotion, l ittle is known regarding the way beta2 integrin-receptors affect the a ctin 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. A ntibody engagement Of beta2 integrins resulted in a rapid and sustaine d (146 and 131% after 30 and 300 s, respectively) increase in the neut rophil F-actin content. This is in contrast to stimulation with N-form yl-l-methionyl-l-leucyl-l-phenylalanine (fMLP), which causes a prompt and pronounced but rapidly declining rise in F-actin (214 and 127% aft er 15 and 300 s, respectively). Priming neutrophils with 1 nM PMA, a l ow concentration that did not influence the F-actin content per se, in creased the magnitude of the beta2 integrin-induced response but had n o effect on the kinetics (199% after 30 s and 169% after 300 s). Remov al of extracellular Ca2+ only marginally affected the beta2 integrin-i nduced F-actin response for cells that were pretreated with PMA wherea s 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 polymerizatio n. Beta2 integrin engagement did not affect the resting cellular level of cAMP arguing against a role of cAMP in beta2 integrin-induced acti n 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 p rompt reversal of the beta2 integrin-induced F-actin elevation in elec tropermeabilized neutrophils. Engagement Of beta2 integrins, as previo usly shown for activation of the chemotactic peptide receptor, resulte d in a significant formation of PtdInsP3. The capacity Of beta2 integr ins and chemotactic peptide receptors to induce phosphatidylinositol t risphosphate (PtdInsP3) formation correlated with their ability to ind uce actin polymerization.