Arachidonic acid and the
calcium ionophore A23187 are known to provoke a pulmonary artery pressor response,
edema formation and release of
thromboxane B2 (TxB2) and 6-keto prostaglandin-F1 alpha (6-keto PGFl alpha) into the recirculating perfusion fluid of isolated blood-free perfused rabbit lungs. Here we investigated the release of
leukotrienes (LTs) by repetitive 0.1 microM
A23187 challenge in the presence or absence of
cyclooxygenase and
5-lipoxygenase inhibitors. RP-HPLC analysis of perfusion fluid extracts persistently showed peaks with retention times of authentic
LTC4, -D4, -E4 and -B4. Fractionated RP-HPLC eluate subjected to radioimmunoassay (RIA) with
LTC4 and
LTB4 antibodies showed two major peaks of immunoreactivity corresponding to those compounds and minor immunoreactivity with
LTD4 and
LTE4 in accordance with the stated cross-reactivities of the
LTC4 antibody. Good correlation for both
LTB4 and
LTC4 levels measured by RP-HPLC versus RIA of collected HPLC peaks was found. Five to ten min after
A23187 challenge,
LTC4, -D4 and -B4 levels ranged from 800 to 1600 pg/ml perfusate.
LTC4 reached a maximum level at 20 min whereas
LTB4 slightly increased over a 35 min period. Upon repeated
A23187 challenge, interrupted by rinsing phases with fresh perfusion fluid, the LT release was reproducible several times with increasing reaction strength. This performed in presence of increasing concentration of the
5-lipoxygenase inhibitors AA-861 or
U-60,257 caused a dose-dependent inhibition of the release of all LTs with an IC50 of approximately 10(-8) to 10(-7) M and 10(-6) M, respectively.
Cyclooxygenase inhibition with
acetylsalicylic acid at doses completely suppressing the
A23187 induced pressor response did not inhibit the peptidoleukotriene release and only slightly depressed
LTB4 release.
CONCLUSION: