Pharmacological studies on the active factors derived from airway epithelial cells.

气道上皮细胞活性因子的药理学研究。

基本信息

批准号：
06454162
负责人：
ITO Yushi
金额：
$ 4.67万
依托单位：
Kyushu University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1994
资助国家：
日本
起止时间：
1994 至 1995
项目状态：
已结题

项目摘要

Airway epithelial cells do not simply act as diffusion barriers against airway stimulating substances such as toxic gases or antigens, but play an important role in regulation of the airway muscle tone by rekeasing multiple active factors. Namely, airway epitelial cells release more than two factors, one of which inhibits excitatory neuro-effector tranamission and an other which modulates the resting membrane potential of the airway smooth muscle cells. To identify these factors we cultured airway epithelial cells and used the cultuured supernatants. Cultured supernatants completely or partially suppressed the contractile response of epithelium-denuded bronchioles to electrical field stimulation and suppressed the excitatory junction potentials. Cultured epithelial cells produce prostaglandins, and exogenously applied PGE_2 inhibits excitatory neuro-effector transmission presynaptically in the dog trachea and bronchioles. Therefore we radioimmunoassayd the amounts of PGE_2 in the super … More natants, which ranged between 3.1-23.8ng/ml. A high concentration of PGE_2 in the supernatant was associated with aboliton of the twitch contraction. Indomethacin, which suppresses the synthesis of PGE_2 in the suprenatants, prevented the inhibitory effect of supernatants on the twitch contractions and excitatoty junction potentials. Thus we concluded that one of the factors which is released from the airway epithelial cells and inhibits excitatory neuro-effector transmission is PGE_2. In an attempt to identify the epithelium derived hyperpolarizing factor (EpDHF) , we freeze-dried the supernatants and then applied them to Gel-filtration chromatograohy on Sephacryl S-100HR and then high-perfoamace liquid chromatofraphy on a c_<18> reversed-phase column. Hyperpolarization assay of the chromatographic fractions showed that active factors eluted in peaks coincided with low molecular weights (10-20 KD). Enzymatic treatment with proteolytic enzymes of the HPLC-purified active fraction resuletd in the abolition of hyperpolarizing activity.The results suggest that the active factor (EpDHF) 1 that was present in the cultured epithelial supenatants was a peptide in nature. Less

气道上皮细胞不仅充当针对有毒气体或抗原等气道刺激物质的扩散屏障，而且通过重新释放多种活性因子在调节气道肌张力中发挥重要作用，即气道上皮细胞释放两种以上因子，其中一种抑制兴奋性神经效应器传递，另一种调节气道平滑肌细胞的静息膜电位。为了识别这些因素，我们培养了气道上皮细胞并使用了培养的上清液完全或部分抑制了上皮剥脱的细支气管对电场刺激的收缩反应，并抑制了培养的上皮细胞产生前列腺素，并且外源性应用的PGE_2抑制了狗气管和突触前的兴奋性神经效应器传递。因此我们对 PGE_2 的含量进行了放射免疫测定。在上清液中，其浓度范围为 3.1-23.8 ng/ml，上清液中高浓度的 PGE_2 与抽搐收缩的消除有关，吲哚美辛抑制了上清液中 PGE_2 的合成，从而阻止了抑制作用。由此我们得出结论，上清液对抽搐收缩和兴奋性连接电位的影响是从气道释放的因素之一。上皮细胞并抑制兴奋性神经效应器传递的是PGE_2。为了鉴定上皮源性超极化因子（EpDHF），我们将上清液冷冻干燥，然后将其应用于Sephacryl S-100HR上的凝胶过滤色谱，然后进行高效液相色谱。在c_ 18 反相柱上进行的perfoamace 液相色谱分析。色谱级分显示，活性因子在峰中洗脱，与低分子量（10-20 KD）一致，用 HPLC 纯化的活性级分的蛋白水解酶进行酶处理，导致超极化活性的消除。结果表明，活性因子（存在于培养的上皮上清液中的EpDHF)1本质上是一种肽。