LEUKOTRIENE-B4 LEVEL IN NEUTROPHILS FROM ALLERGIC AND HEALTHY-SUBJECTS STIMULATED BY LOW CONCENTRATION OF CALCIUM IONOPHORE A23187 - EFFECT OF EXOGENOUS ARACHIDONIC-ACID AND POSSIBLE ENDOGENOUS SOURCE

Peripheral blood neutrophils from patients with allergic rhinitis and from normal subjects were incubated for 5 min at 37-degrees-C with 0.15-mu-M calcium ionophore A23187 in the absence or presence of exogenous arachidonic acid (2.5 to 10-mu-M). In neutrophils from allergic patients, the leukotriene B4 (LTB4) level was significantly increased by exogenous arachidonic acid in a concentration-dependent manner (16.2 +/- 4.2 and 38.1 +/- 6.8 pmol / 5 min per 2.10(6) cells in the absence and presence of 10-mu-M arachidonic acid, respectively; P < 0.005; n = 8). The LTB4 level in neutrophils from healthy subjects was only 0.97 +/- 0.17 pmol / 5 min per 2.10(6) cells (n = 5) and was not enhanced by exogenous arachidonate. When cells from allergic patients were challenged in the presence of exogenous [1-C-14]arachidonic acid, released LTB4 was radiolabeled and the incorporated radioactivity increased with the labeled arachidonate concentration. Labeled LTB4 was never detectable after incubating neutrophils from normal donors with exogenous labeled arachidonate. When neutrophils were incubated with [1-C-14]arachidonate for 1 h, the different lipid pools of the two cell populations were labeled but both types of neutrophils produced unlabeled LTB4 in response to ionophore stimulation. The hydrolysis of choline and ethanolamine phospholipids into diacyl-, alkenylacyl- and alkylacyl-species revealed that solely the alkylacyl-subclass of phosphatidylcholine was unlabeled. We conclude (i) that neutrophils from allergic patients stimulated by low ionophore concentration produce more LTB4 than neutrophils from healthy subjects and incorporate exogenous arachidonate, (ii) that endogenous arachidonate converted to LTB4 by the 5-lipoxygenase pathway may provide only from 1-O-alkyl-2-arachidonoyl-glycero-3-phosphocholine.