INTRACELLULAR CALCIUM AND CALCINEURIN REGULATE NEUTROPHIL MOTILITY ONVITRONECTIN THROUGH A RECEPTOR IDENTIFIED BY ANTIBODIES TO INTEGRINS ALPHA-V AND BETA-3

Buffering of intracellular calcium ([Ca2+](i)) or inhibition of the ca lcium/calmodulin-dependent phosphatase, calcineurin, results in neutro phils being unable to detach from vitronectin with a consequent loss o f motility, Treatment of [Ca2+](i)-buffered or calcineurin-inhibited n eutrophils with monoclonal antibodies (MoAbs) to beta 3 or alpha v bet a 3 integrins allowed neutrophils to detach and restored motility. Qua ntitative immunofluorescence and flow cytometry showed that MoAbs spec ific for beta 3, alpha v, or alpha v beta 3 integrins bind to neutroph ils. Immunolocalization studies using antibodies to the highly conserv ed cytoplasmic domains of alpha v and beta 3 also identified the recep tor on neutrophils. Whereas antibodies to cuv, alpha v beta 3, and bet a 3 recognized the receptor in intact cells, only the beta 3 MoAb immu noprecipitated the receptor from a neutrophil cell lysate. The cu subu nit co-immunoprecipitated by the beta 3 antibody reacted with an antib ody to alpha v by Western blot, Peptide maps of V8 protease digests sh owed a strong similarity in alpha and beta chains precipitated by anti bodies to beta 3 from neutrophils and endothelial cells. These results indicate that [Ca2+](i) and calcineurin regulate neutrophil motility on vitronectin through an alpha v beta 3-like receptor. Although we ca nnot rule out the possibility that neutrophils have an isoform of alph a v, such an isoform would have to be similar enough to react with alp ha v- and alpha v beta 3-specific MoAbs in intact cells. (C) 1996 by T he American Society of Hematology.