{'fraction_non_alphanumeric': 0.04004881295762148, 'fraction_numerical': 0.026403372531617484, 'mean_word_length': 4.764066496163683, 'pattern_counts': {'":': 0, '<': 1, '<?xml version=': 0, '>': 0, 'https://': 0, 'lorem ipsum': 0, 'www.': 0, 'xml': 0}, 'pii_count': 0, 'substrings_counts': 0, 'word_list_counts': {'cursed_substrings.json': 0, 'profanity_word_list.json': 0, 'sexual_word_list.json': 0, 'zh_pornsignals.json': 0}}
Effect of leukotriene D4 on tracheal mucociliary transport velocity in quails.

We investigated the effect of leukotriene D4 (LTD4) on tracheal mucociliary transport in quails. Topical application of LTD4 (0.2-2 ng) to tracheal mucosa dose-dependently increased mucociliary transport velocity (MCTV) in 5 or 10 min after application. Forty minutes after application of 2 ng of LTD4, MCTV was decreased to about 84% of that in the control group. Both the transient increase and the subsequent decrease induced by 2 ng of LTD4 were blocked by ONO-1078 (Pranlukast: 4-oxo-8-[4-(4-phenylbutoxy)-benzoylamino]-2-(tetrazol-5-yl)- 4H- 1-benzopyran) (0.03-3 mg/kg, i.m.), a specific leukotriene antagonist. These results suggest that LTD4 possesses a biphasic effect on tracheal mucociliary transport through leukotriene receptors.

ABSTRACT-We investigated the effect of leukotriene D4 (LTD4) on tracheal mucociliary transport in quails. Topical application of LTD4 (0.2-2 ng) to tracheal mucosa dose-dependently increased mucociliary transport velocity (MCTV) in 5 or 10 min after application. Forty minutes after application of 2 ng of LTD4, MCTV was decreased to about 841o of that in the control group. Both the transient increase and the subsequent decrease induced by 2 ng of LTD4 were blocked by ONO-1078 (Pranlukast: 4-oxo-8-[4-(4 phenylbutoxy)-benzoylamino]-2-(tetrazol-5-yl)-4H-1-benzopyran) (0.03-3 mg/kg, i.m.), a specific leuko triene antagonist. These results suggest that LTD4 possesses a biphasic effect on tracheal mucociliary transport through leukotriene receptors. Keywords: Tracheal mucociliary transport, Leukotriene D4, ONO-1078 Peptide leukotrienes, 5-lipoxygenase products of arachidonic acid metabolism, induce potent bronchocon striction, increase vascular permeability leading to mu cosal edema, and enhance mucus secretion (1). Therefore, peptide leukotrienes may be crucial to pathogenesis in al lergic bronchial asthma (2). Although mucociliary trans port is an important pulmonary defense mechanism, effects of leukotrienes on mucociliary transport remain unconclusive. Some findings showed that leukotriene D4 (LTD4), receptors of which are abundant in the lung (3), impaired mucociliary transport in allergic and nonallergic sheep (4). On the other hand, there are contradictory reports that LTD4 increased tracheal ciliary beat fre quency in sheep and dogs (5, 6). Accordingly, in the present study, we examined the effect of LTD4 on tracheal mucociliary transport velocity (MCTV) using a newly-developed method (7) in which LTD4 is directly applied to the tracheal mucosa in quails.
Male quails weighing 100-120 g (Kyudo Co., Ltd., Fukuoka) were anesthetized with urethane (1 g/kg, i.p.), and then they were fixed on their back and the feathers on the larynx were cut off. The skin of the front neck was ripped opened 3 cm along the median line with two pairs of forceps and the traumatic margins were clipped with two celfins to keep the tramatic opening slightly open. The local blood vessels and connective tissues were care fully separated from the trachea to expose the trachea. A thread was laid under the exposed trachea so that the part of the trachea used for the test could be kept level. An incision (approx. 2 cm) in the trachea was made with a hot knife. As soon as the operation was over, the quail was inserted into an observation box. In the observation box, the quail tracheal mucosa was kept under about 38C and approximately 100% humidity by use of a humidifier (8,9). Ash powders were placed on the caudal side of the tracheal mucosa to select the site where the powder was carried at the fastest speed, and MCTV was measured at the same site throughout the experiment. When MCTV became constant, the time taken for the powder to move 10 mm was measured.
LTD4 was donated by Ono Pharmaceutical Co., Ltd.
(Osaka). The LTD4 stock, which was dissolved in 50% ethanol and stored at -601C, was diluted with phos phate-buffered saline without Ca 2+ and Mg2+ (PBS) to appropriate concentrations. In the observation box, the LTD4 solutions were applied directly to the trachea ac cording to the method developed in our laboratory (7). This method is advantageous because it is not only ac curate and stable but also simple. A 1-pl aliquot of LTD4 was splashed through a 21-gauge needle with a plastic sprayer on its tip, by applying 0.5 kg/cm2 of air pressure for 30 msec. The switch to apply air pressure was conducted by an electro-magnetic valve controlled by an electronic stimulator (SEN-3201; Nihon Kohden, Co., Ltd., Tokyo). In sham-treated animals, 1 % ethanol in PBS was applied. Ethanol or LTD4 was applied 15 min after the MCTV measurement started. ONO-1078 (Pranlukast: 4-oxo-8-[4-(4-phenylbutoxy)benzoylamino]-2 (tetrazol-5-yl)-4H-1-benzopyran; Ono Pharmaceutical Co., Ltd., Osaka), a specific leukotriene receptor antago nist (1), was dissolved freshly in 50% ethanol and then administered into the femoral muscle 15 min before etha nol or LTD4 was applied. In each experiment, MCTV was measured for 75 min through a microscope. The results were calculated by the following equation: VT/VO x 100% (VT = the average velocity in each 5 min and Vo=the aver age velocity in the last 5 min before application). MCTV values are expressed as means ±S.E. Statistical analyses were performed by Dunnett's t-test, and a P value <0.05 was considered significant.
The application of 0.2 to 2 ng of LTD4 significantly in creased MCTV in a dose-dependent manner, whereas 1 % ethanol had little effect on MCTV (Fig. 1) The increase induced by 2 ng of LTD4 was dose-depend ently inhibited by pretreatment with 0.03 3 mg/kg of ONO-1078, while ONO-1078 alone showed no effect on MCTV (Fig. 2). ONO-1078 also significantly reversed the LTD4-induced delayed decrease in a dose-dependent fashion.  Impaired mucociliary clearance is a typical characteris tic of asthmatic attack induced by allergen, which may be caused by release of autacoid mediators from the im munoglobulin (IgE)-antigen reaction with resident or inflammatory cells in the airway (2). The efficacy of ciliary action to propel inhaled particles and cellular debris in the airway toward the pharynx depends upon several characteristics including the frequency and coordination of ciliary beat, rheological properties of the airway sur face fluid and mucus secretion (5). It has been well recog nized that LTD4 can facilitate ciliary beat frequency (5, 6). In the present study, we observed that 2 ng of LTD4 increased MCTV within the first 5 min and the increase gradually subsided in the next 5 min. The time course of this increase in MCTV was similar to that of the increase in ciliary beat frequency (5). Therefore, the transient in crease in MCTV induced by LTD4 may result from the LTD4-induced increase of ciliary beat frequency.
We also found that 2 ng of LTD4 induced a decrease in MCTV 40 min after application. This result is consistent with the finding that the earliest decrease in MCTV was observed 30 min after airway challenge with LTD4 in al lergic and nonallergic sheep (4). LTD4 has been identified as a potent mucus secretagogue (4), affecting rheological properties of the airway surface fluid. Therefore, it is possible that the LTD4-induced decrease in MCTV could be related to the mucus secretagogue effect of LTD4 (4, 5). The syntheses and release of macromolecular mucous glycoproteins induced by LTD4 are a series of relatively prolonged biological processes (5, 10) and the time-con suming processes may explain the time lag between LTD4 application and appearance of the decreasing effect on MCTV.
ONO-1078 suppressed both the transient increase and the subsequent decrease induced by 2 ng of LTD4, sug gesting LTD4-receptor-mediated actions on tracheal mu cosa. LTD4 receptors are known to be coupled to phos pholipase C via a pertussis toxin-sensitive G-protein in airway smooth muscles (11). Consequently, LTD4 will in duce polyphosphoinositide hydrolysis and protein kinase C activation (11). Recently, we found that protein kinase C activation induced mucus secretion in hamster tracheal epithelial cells (12). At least in part, the excessive mucus secretion induced by protein kinase C activation may be involved in the subsequent decrease induced by LTD4.
In conclusion, using a newly-developed method for topical application, we demonstrated that LTD4 had a biphasic action on mucociliary transport function, con sisting of an initial transient increase that may be caused by ciliostimulation and a subsequent prolonged depres sion that may be due to excessive mucus secretions (5). In addition, if LTD4 can partly contribute to the impaired mucociliary transport function, ONO-1078 may possess some preventive roles in the impaired function without effects on normal mucociliary function. With respect to human tracheobronchial diseases, however, the sig nificance of the present results in quails remains to be tested by experiments involving patients and normal sub jects.