The effects of cromakalim on the mediator releases from guinea pig lung mast cell activated by specific antigen-antibody reactions

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.

The effects of cromakalim on the mediator releases from guinea pig lung mast cell activated by specific antigen-antibody reactions

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.

Basic fibroblast growth factor does not initiate mitogenic signalling via phosphoinositide hydrolysis in PC12 cells.

Basic fibroblast growth factor (b FGF) was found to be equally potent mitogen as compared to alpha-thrombin to reinitiate DNA synthesis in quiescent PC12 cells. Whereas thrombin was found to be an activator of phospholipase C as judged by a rapid increase in the formation of inositol triphosphate, inositol biphosphate and a massive accumulation of inositol phosphate when 20 mM LiCl was present as an inhibitor of inositol mono phosphatases, basic FGF failed to induce the breakdown of the polyphosphoinositides in quiescent PC12 cells to any appreciable levels, however, a simultaneous increase in the level of diacylglycerol was observed. b FGF also failed to stimulate protein kinase C which is believed to be activated by diacylglycerol. It is therefore concluded that bFGF receptor mediated 'signalling is not via phospholipase C activation and bFGF's early mitogenic responses and DNA synthesis are initiated independent of the inositol lipids and protein kinase C activation. Thus bFGF must have its own unique signal transducing mechanism independent of inositol pathways.