Relaxant effect of 1-butanol fraction from Elaeagnus pungens leaf through inhibiting l-type Ca2+ channel on guinea pig tracheal smooth muscle

The leaf of Elaeagnus pungens thunb. (Family Elaeagnaceae) has been documented as an effective herb for the treatment of asthma and chronic bronchitis in traditional Chinese medicine. In the past years, only a few of preliminary studies reported the chemical constituents and pharmacology effects of the herb, but their action on the tracheal relaxation has not been investigated. To investigate the relaxing effect and mechanism of the extracts from Elaeagnus pungens leaves on guinea pig tracheal smooth muscle and bronchi smooth muscle cells. Four fractions of different polarities from Elaeagnus pungens leaves were tested to the tracheal strips on the resting tension or pre-contracted by histamine (20μM) and acetylcholine (20μM). Inhibitory effects of the 1-butanol fraction (400mg/ml) on cumulative histamine and acetylcholine (0.2–20μM) induced contraction were measured. In order to determine the mediators on the 1-butanol fraction effect, the relaxing effect of the 1-butanol fraction was evaluated in the absence and presence of β-adrenoceptor antagonists (1μM propranolol), K+ channels-blockers (4-aminopyridine (2mM), tetraethylammonium chloride (5mM) or glibenclamide (10μM)), the cyclooxygenase inhibitor (indomethacin, 10μM), nitric oxide synthase inhibitor (Nω-nitro-l-arginine methyl ester, 100μM) or l-type Ca2+ channel inhibitor (nifedipine, 1μM). Moreover, [Ca2+]i in bronchi smooth muscle cells was analyzed by measuring the fluorescence intensity with confocal system. 1-Butanol fraction induced the highest relaxant effect among four fractions of different polarities from Elaeagnus pungens leaves, and significantly relaxed the tracheal strip in the concentration-dependent manner on the resting tension and pre-contracted by histamine phosphate and acetylcholine. It also produced an unparallel rightward shift of the cumulative concentration-response curve of histamine or acetylcholine. Furthermore, the relaxant effect of 1-butanol fraction was not affected by propranolol, glibenclamide, tetraethylammonium chloride, 4-aminopyridine, indomethacin and Nω-nitro-l-arginine methyl ester. However, 1-butanol fraction-induced relaxation decreased after adding nifedipine. It also concentration-dependently inhibited CaCl2-induced contraction in the Ca2+-free, 60mM K+-containing solution. Additionally, [Ca2+]i in the BSMCs significantly reduced after administration of the 1-butanol fraction. The 1-butanol fraction from Elaeagnus pungens leaves resulted in a relaxation i