Ro J Y, Yim Y N, Kim K H
Department of Pharmacology, Yonsei University College of Medicine, Seoul, Korea.
Yonsei Med J. 1996 Oct;37(5):325-38. doi: 10.3349/ymj.1996.37.5.325.
The inhibitory effect of cromakalim on the mediator release from mast cells caused by antigenantibody reactions was in controversy with the specific antigen used. However, it has recently been observed that cromakalim inhibits the release of mediators from superfused tracheal and parenchymal strips or lung mast cells after passive sensitization with the IgG1 antibody. An attempt, therefore, was made to determine the inhibitory mechanisms of cromakalim on the release of mediators such as histamine and leukotriene released by the activation of enzymes during mast cell activation. Guinea pig lung mast cells were purified through enzyme digestion, rough percoll and continuous percoll density gradients. The purified mast cells were prelabeled with [3H]palmitic acid. PLD activity was assessed more directly by the production of labeled phosphatidylethanol by PLD-mediated transphosphatidylation in the presence of ethanol. In the cells labelled with [3H]myristic acid, [3H] DAG production was measured. The methyltransferase activity was assessed by measuring the incorporation of [3H]methyl moiety into phospholipids in sensitized mast cells labelled with L-[3H] methylmethionine. cAMP level was measured by radioimmunoassay. Cromakalim resulted in a decrease in the amount of histamine and leukotrienes releases by 30% in the ovalumin-induced mast cell. Cromakalim had little effect on phospholipase D activity enhanced by the activated mast cell. Cromakalim inhibited the initial increase of diacylglycerol production during mast cell activations. Cromakalim inhibited the phospholipid methylation increased in the activated mast cell. These results show that cromakalim decreases histamine release by inhibiting the initial increase of 1,2-diacylglycerol during the mast cell activation, which is mediated via the phosphatidylinositide-phospholipase C system rather than the phosphatidylcholine-phospholipase D system. Furthermore, cromakalim reduces phosphatidylcholine production by inhibiting the methyltransferase, which decreases the conversion of phosphatidylcholine into arachidonic acid and inhibits the production of leukotrienes.
克罗卡林对抗原抗体反应引起的肥大细胞介质释放的抑制作用因所用的特异性抗原而异。然而，最近观察到，在用IgG1抗体被动致敏后，克罗卡林可抑制来自灌流气管和实质条带或肺肥大细胞的介质释放。因此，人们试图确定克罗卡林对肥大细胞活化过程中酶激活所释放的组胺和白三烯等介质释放的抑制机制。通过酶消化、粗制 Percoll 和连续 Percoll 密度梯度法纯化豚鼠肺肥大细胞。纯化的肥大细胞用[3H]棕榈酸进行预标记。通过在乙醇存在下 PLD 介导的转磷脂酰化作用产生标记的磷脂酰乙醇，更直接地评估 PLD 活性。在用[3H]肉豆蔻酸标记的细胞中，测量[3H]二酰甘油的产生。通过测量用 L-[3H]甲基甲硫氨酸标记的致敏肥大细胞中[3H]甲基部分掺入磷脂的情况来评估甲基转移酶活性。通过放射免疫测定法测量 cAMP 水平。在卵清蛋白诱导的肥大细胞中，克罗卡林使组胺和白三烯的释放量减少了30%。克罗卡林对活化肥大细胞增强的磷脂酶 D 活性几乎没有影响。克罗卡林抑制肥大细胞活化过程中二酰甘油产生的初始增加。克罗卡林抑制活化肥大细胞中增加的磷脂甲基化。这些结果表明，克罗卡林通过抑制肥大细胞活化过程中1,2-二酰甘油的初始增加来减少组胺释放，这是通过磷脂酰肌醇-磷脂酶 C 系统而非磷脂酰胆碱-磷脂酶 D 系统介导的。此外，克罗卡林通过抑制甲基转移酶来减少磷脂酰胆碱的产生，这减少了磷脂酰胆碱向花生四烯酸的转化并抑制白三烯的产生。
