Regulation of Human Polymorphonuclear Leukocytes Functions by the Neuropeptide Pituitary Adenylate Cyclase-Activating Polypeptide after Activation of MAPKs

Abstract

Anti-inflammatory activities of pituitary adenylate cyclase-activating protein (PACAP) are mediated in part through specific effects on lymphocytes and macrophages. This study shows that in human polymorphonuclear neutrophils (PMNs), PACAP acts as a proinflammatory molecule. In PMNs, vaso-intestinal peptide/PACAP receptor 1 (VPAC-1) was the only receptor found to be expressed by RT-PCR. Using VPAC-1 Ab, we found that VPAC-1 mRNA was translated into proteins. In PMNs, PACAP increases cAMP, inositol triphosphate metabolites, and calcium. It activates two of the three members of the MAPK superfamily, the ERK and the stress-activated MAPK p38. U73122, an inhibitor of phospholipase C (PLC), inhibits PACAP-induced ERK activation, whereas p38 MAPK phosphorylation was unaffected. Using specific pharmalogical inhibitors of ERK (PD098059) and p38 MAPK (SB203580), we found that PACAP-mediated calcium increase was ERK and PLC dependent and p38 independent. PACAP primes fMLP-associated calcium increase; it also primes fMLP activation of the respiratory burst as well as elastase release, these last two processes being ERK and PLC dependent and p38 MAPK independent. PACAP also increases membrane expression of CD11b and release of lactoferrin and metallo proteinase-9 (MMP-9). These effects were PLC dependent (CD 11b, lactoferrin, MMP-9), ERK dependent (CD 11b, lactoferrin, MMP-9), and p38 dependent (CD11b, lactoferrin). We conclude that PACAP is a direct PMN activator as well as an effective PMN priming agent that requires PLC, ERK, and p38 MAPK activities.

Pituitary Adenylate Cyclase-Activating Polypeptide 27 Is a Functional Ligand for Formyl Peptide Receptor-Like 1

“…These data suggest that PACAP27 specifically stimulates human neutrophils. In view of the fact that VPAC1 can be stimulated by VIP or PACAP38, as well as by PACAP27, our results were not consistent with those of a previous report which suggested that VPAC1 functions as a PACAP receptor in neutrophils (15). These data suggest that another receptor may be involved in the process of PACAP27-induced intracellular signaling in human neutrophils.…”

“…2E), but failed to inhibit PACAP27-induced cAMP elevation (Fig. 2G), indicating that WRW4 does not affect VPAC1, which has been reported to be expressed in human neutrophils (15). ERK phosphorylation was also completely inhibited by pretreating with WRW4, indicating that this ERK phosphorylation is also a part of the FPRL1-dependent signaling cascade (Fig.…”

“…Previously, it has been reported that PACAP27 primes neutrophil response to the fMLP (15). Bacterial fMLP can activate FPR and FPRL1 at high concentrations, but fMLP activates only FPR at low FIGURE 5.…”

“…Previously, PACAP27 has been reported to prime fMLP-induced calcium signaling (15). To determine the FPRL1 dependency, we examined the effect of PACAP27 on fMLP-induced calcium signaling with or without WRW4.…”

“…PACAP27 has been found to stimulate the surface expression of CD11b, a known mediator of neutrophil adhesion and mobilization (15). Purified neutrophils were incubated with PACAP27, and analyzed by flow cytometry, as shown by the dot plots in Fig.…”