CDB15:0000064 ANXA1 — FPR2

ABSTRACT: Recent investigations conducted with human neutrophils have indicated an involvement for the receptor for formylated peptides, termed FPR, and its analog FPRL1 (or ALXR because it is the receptor for the endogenous ligand lipoxin A(4)) in the in vitro inhibitory actions of the glucocorticoid-regulated protein annexin 1 and its peptidomimetics. To translate these findings in in vivo settings, we have used an ischemia/reperfusion (I/R) procedure to promote leukocyte-endothelium interactions in the mouse mesenteric microcirculation. In naive mice, the annexin 1 mimetic peptide Ac2-26 (20 to 100 microg administered intravenously prior to reperfusion) abolished I/R-induced cell adhesion and emigration, but not cell rolling. In FPR-deficient mice, peptide Ac2-26 retained significant inhibitory actions (about 50% of the effects in naive mice), and these were blocked by an FPR antagonist, termed butyloxycarbonyl-Phe-Leu-Phe-Leu-Phe, or Boc2. In vitro, neutrophils taken from these animals could be activated at high concentrations of formyl-Met-Leu-Phe (30 microM; fMLP), and this effect was blocked by cell incubation with peptide Ac2-26 (66 microM) or Boc2 (100 microM). FPR-deficient neutrophils expressed ALXR mRNA and protein. Both ALXR agonists, lipoxin A(4) and peptide Ac2-26, provoked detachment of adherent leukocytes in naive as well as in FPR-deficient mice, whereas the CXC chemokine KC or fMLP were inactive. The present findings demonstrate that endogenous regulatory autocoids such as lipoxin A(4) and annexin 1-derived peptides function to disengage adherent cells during cell-cell interactions.

ABSTRACT: We have tested the effects of annexin 1 (ANXA1) and its N-terminal peptide Ac2-26 on polymorphonuclear leukocyte (PMN) recruitment under flow. Differential effects of the full-length protein and its peptide were observed; ANXA1 inhibited firm adhesion of human PMNs, while Ac2-26 significantly attenuated capture and rolling without effect on firm adhesion. Analysis of the effects of ANXA1 and Ac2-26 on PMN adhesion molecule expression supported the flow chamber results, with Ac2-26 but not ANXA1 causing l-selectin and PSGL-1 shedding. ANXA1 and its peptide act via the FPR family of receptors. This was corroborated using HEK-293 cells transfected with FPR or FPRL-1/ALX (the 2 members of this family expressed by human PMNs). While Ac2-26 bound both FPR and FPRL-1/ALX, ANXA1 bound FPRL-1/ALX only. ANXA1 and Ac2-26 acted as genuine agonists; Ac2-26 binding led to ERK activation in both FPR- and FPRL-1/ALX-transfected cells, while ANXA1 caused ERK activation only in cells transfected with FPRL-1/ALX. Finally, blockade of FPRL-1/ALX with a neutralizing monoclonal antibody was found to abrogate the effects of ANXA1 in the flow chamber but was without effect on Ac2-26-mediated inhibition of rolling. These findings demonstrate for the first time distinct mechanisms of action for ANXA1 and its N-terminal peptide Ac2-26.