PARADOXICAL BLOCKADE OF BETA-ADRENERGICALLY MEDIATED INHIBITION OF STIMULATED EOSINOPHIL SECRETION BY SALMETEROL

Salmeterol (SALM) is a long-acting beta(2) adrenoceptor agonist that c auses prolonged relaxation of airway smooth muscle. To determine wheth er this agent also causes prolonged inhibition of stimulated eosinophi l secretion, we studied interactions between SALM and albuterol (ALB) in inhibiting eosinophil peroxidase (EPO) secretion in human eosinophi ls in vitro. Peripheral blood eosinophils were isolated from 18 human volunteers by negative immunoselection, and secretion of EPO was elici ted with 10(-6) M formyl-met-leu-phe (fMLP) + 5 mu g/ml cytochalasin B (CytB) in aliquots of 10(5) cells. Eosinophils were pretreated with e ither 10(-8) M ALB, 10(-8) M SALM or SALM + ALB for 5 min to 18 hr at 37 degrees C. Pretreatment with ALB for 5 min caused inhibition of sti mulated secretion of EPO to 783 +/- 210 ng/10(6) cells vs. 1475 +/- 28 6 ng/10(6) cells for eosinophils not treated with ALB (P < .05; n = 5) . Inhibition of EPO secretion caused by ALB was sustained for 30 min ( 924 +/- 160 ng/10(6) cells; P < .05 vs. fMLP + CytB; n = 5). By contra st, SALM had no inhibitory effect on fMLP-induced secretion after incu bation for 5 min to 18 hr. In cells obtained from four separate isolat ions, pretreat ment with 10(-8)M SALM before addition of ALB blocked t he inhibition of EPO release caused by 10(-8) M ALB alone (486 +/- 28 ng/10(6) cells for ALB alone vs. 902 +/- 32 ng/10(6) cells for SALM ALB; P < .01; n = 4). Incubation with ALB caused an increase in intrac ellular concentration of cAMP at 5 min from 433 + 106 fmol/10(6) cells to 944 +/- 186 fmol/10(6) cells (P < .05; n = 4). In separate studies , SALM alone had no effect on eosinophil cAMP concentration (408 +/- 4 5.5 fmol/10(6) cells; P = N.S. vs. control; n = 4). Preincubation with the same concentration of SALM (10(-8)M) that blocked the inhibition of EPO secretion caused by 10(-8) M ALB also blocked the ALB-induced i ncrease in eosinophil cAMP concentration (373 +/- 77.0 fmol/10(6) cell s; P = N.S. vs. control; n = 4). We demonstrate that ALB, but not SALM , causes inhibition of the EPO secretion elicited by fMLP + CytB. We a lso demonstrate that SALM antagonizes both the increase in cAMP and th e inhibitory effects on fMLP-induced stimulation caused by ALB at 5 mi n. Our data demonstrate that SALM antagonizes the inhibition of EPO se cretion caused by ALB in human peripheral blood eosinophils.