Cytosolic phospholipase A2, increased and activated in the eosinophils of patients with hypereosinophilic syndrome in vivo, is involved in the augmented release of leukotriene C4.
OBJECTIVE
Eosinophils from patients with hypereosinophilic syndrome (HES) showed augmented release of leukotriene C4 (LTC4) by stimulation with A23187. In the present study, we investigated the involvement of cytosolic phospholipase A2 (cPLA2) in this phenomenon.


METHODS
Eosinophils from normal and HES donors (2.5 x 10(6) cells/ml) were incubated with A23187 (0.03-3 microM) for 60 min in the presence or absence of a cPLA2 inhibitor, AACOCF3. The LTC4 released from eosinophils was measured by enzyme immunoassay. Distribution of cPLA2 and 5-lipoxygenase (5-LO) proteins within the eosinophils were detected by Western immunoblotting.


RESULTS
The level of LTC4 released from the HES eosinophils by stimulation with A23187 was higher than that from normal eosinophils. The A23187-induced LTC4 release was inhibited by AACOCF3 in a dose-dependent manner. The amounts of cPLA2 seemed to be increased in the non-stimulated HES eosinophils by an analysis of immunoblotting. To be noticed was that cPLA2 was detected as a phosphorylated and membrane-bound form in the HES eosinophils, but not in the normal eosinophils. In contrast, localization of 5-LO within the eosinophils under A23187 stimulation was not different between normal and HES donors, while the amounts of 5-LO also seemed to be increased in the HES eosinophils.


CONCLUSIONS
These results suggest that cPLA2, increased and activated (phosphorylated and membrane-translocated) in vivo, is involved in the augmented release of LTC4 from the HES eosinophils.
Urasaki
T
T
Division of Hematology, University of Tsukuba, Ibaraki, Japan.
Ninomiya
H
H
Takasaki
J
J
Kawauchi
Y
Y
Nagasawa
T
T
Masuho
Y
Y
D001094:Arachidonate 5-Lipoxygenase / Q000378:metabolism; D015153:Blotting, Western; D000001:Calcimycin / Q000494:pharmacology; D002462:Cell Membrane / Q000201:enzymology / Q000378:metabolism; D003600:Cytosol / Q000201:enzymology*; D004789:Enzyme Activation; D004804:Eosinophils / Q000201:enzymology* / Q000378:metabolism*; D006801:Humans; D017681:Hypereosinophilic Syndrome / Q000097:blood / Q000201:enzymology*; D066298:In Vitro Techniques; D017997:Leukotriene C4 / Q000378:metabolism*; D010741:Phospholipases A / Q000378:metabolism*; D054467:Phospholipases A2