Salbutamol delays human eosinophil apoptosis via a cAMP-dependent mechanism

Abstract

Eosinophils play a major role in asthma. One described mechanism leading to the impaired clearance of these cells from the lung is the delay in their programmed cell death (apoptosis). β 2-Adrenoceptor agonists have been shown to prolong survival and delay apoptosis of eosinophils. The aim of the present study was to evaluate the mechanisms, especially the role of cAMP pathway, in the prolongation of human eosinophil survival by a selective β 2-agonist salbutamol. Isolated human peripheral blood eosinophils were cultured in the absence or presence of a β 2-agonist salbutamol and the indicated antagonists/inhibitors under sterile conditions. Apoptosis was measured by using the relative DNA fragmentation assay and Annexin-V binding. Salbutamol prolonged survival of human eosinophils and it was inhibited by a β-receptor antagonist propranolol and mimicked by cell-permeant cAMP analogues dibutyryl- and 8-bromo-cAMP. Pharmacological inhibitors of adenylyl cyclase (SQ-22,536) and protein kinase A (Rp-8-CPT-cAMPS) antagonized the effects of salbutamol. The survival-prolonging action of salbutamol was potentiated by a phosphodiesterase inhibitor rolipram (EC 50 for the salbutamol effect was 13.6 ± 4.0 and 8.1 ± 3.1 nM in the absence and presence of rolipram, respectively; p = 0.0142, n = 10). In contrast, inhibition of Ca 2+-activated K +-channels by apamin, charybdotoxin, iberiotoxin or paxilline did not affect the ability of salbutamol to prolong eosinophil survival. Taken together, the present results suggest that salbutamol at clinically relevant concentrations decreases apoptosis in human eosinophils by activating the cannonical β 2-receptor-adenylyl cyclase–cAMP-protein kinase A pathway.