Cultured epithelial cells release cyclooxygenase-dependent and cyclooxygenase-lndependent factors that inhibit cholinergic contraction of canine airway smooth muscles

We observed the effects of supernatants from cultured epithelial cellson the contraction of tracheal smooth muscle evoked by acetylcholine (ACh) or by electrical field stimulation (EFS). Cultured canine tracheal epithelial cells were incubated in Krebs solution with or without indomethacin (10^-5M) for 30 and 120 min. The amplitude of the tracheal smooth muscle contractions evoked by EFS or exogenously applied ACh were measured before and after the application of each supernatant in the combined presence of both indomethacin (10^-5M) and propranolol (10^-6M). The control supernatant incubated without indomethacin markedly suppressed the amplitude of the contraction evoked by EFS, but not by ACh. The supernatant incubated with indomethacin for 30 min did not show any effects on the contractile responses evoked by EFS or ACh. However, the supernatants from the cultured epithelial cells incubated for a longer period (120 min) in the presence of indomethacin significantly suppressed the contraction evoked by EFS, but not by ACh. The prostaglandin E₂(PGE₂) concentration was markedly higher in the supernatants incubated without indomethacin (1.39 ± 0.51 ng/ ml, 30 min incubation) than in those with indomethacin (0.02 ± 0.01 ng/ml, 30 min incubation, and 0.06 ± 0.01 ng/ml, 120 min incubation). To determine whether PGE₂is responsible for the inhibitory effect of the supernatants, we evaluated the effects of PGE₂on the resting tone, and EFS- or ACh-evoked contraction. 10^-12to 10^-6Mof PGE₂showed no significant effect on the resting tone. 10^-9M of PGE₂, corresponding to the concentration of the supernatants incubated without indomethacin, and 10^-11Mof PGE₂, to that of the supernatants incubated with indomethacin, showed a similar extent of inhibitory effects to the corresponding supernatants on the EFS-evoked contraction, and no effect on the ACh-evoked contraction. These results suggest that cultured airway epithelial cells release at least two factors spontaneously even without stimulation. One of these factors may be prostanoid (PGE₂), which acts prejunctionally to inhibit the contractile response. The other factor is distinct from prostanoid and inhibits smooth muscle contraction, presumably by suppressing ACh release from vagus nerve termini.