Bauldry S A, Wykle R L, Bass D A
Department of Medicine, Bowman Gray School of Medicine, Wake Forest University, Winston-Salem, North Carolina 27103.
J Biol Chem. 1988 Nov 15;263(32):16787-95.
Both 1,2-diacyl- and 1-O-alkyl-2-acylglycerols are formed during stimulation of human neutrophils (PMN), and both can prime respiratory burst responses for stimulation by the chemotactic peptide, N-formyl-Met-Leu-Phe (fMLP); however, mechanisms of priming are unknown. Arachidonic acid (AA) release through phospholipase A2 activation and metabolism by 5-lipoxygenase are important activities of PMN during inflammation and could be involved in the process of primed stimulation. Therefore, we have examined the ability of diacyl- and alkylacylglycerols to act as priming agents for AA release and metabolism in human neutrophils. After prelabeling PMN phospholipids with [3H]AA, priming was tested by incubating human PMN with the diacylglycerol, 1-oleoyl-2-acetylglycerol (OAG), or its alkylacyl analog, 1-O-delta 9-octadecenyl-2-acetylglycerol (EAG) before stimulating with fMLP. fMLP (1 microM), OAG (20 microM), or EAG (20 microM) individually caused little or no release of labeled AA. However, after priming PMN with the same concentrations of either OAG or EAG, stimulation with 1 microM fMLP caused rapid (peak after 1 min) release of 6-8% of [3H]AA from cellular phospholipids; total release was similar with either diglyceride. Priming cells with OAG also enhanced conversion of released AA to leukotriene B4 (LTB4) and 5-hydroxyeicosatetraenoic acid (5-HETE) upon subsequent fMLP stimulation, but AA metabolites were not increased in EAG-primed PMN. If fMLP was replaced with the calcium ionophore A23187 (which directly causes release of AA and production of LTB4 and 5-HETE), priming by both diglycerides again enhanced release of [3H]AA, but only OAG priming increased lipoxygenase activity. Indeed, EAG pretreatment markedly reduced LTB4 and 5-HETE production. Thus, both diglycerides prime release of AA from membrane phospholipids but have opposite actions on the subsequent metabolism of AA.
1,2 - 二酰基甘油和1 - O - 烷基 - 2 - 酰基甘油在人中性粒细胞（PMN）受刺激过程中均会形成，并且二者均可引发对趋化肽N - 甲酰 - 蛋氨酸 - 亮氨酸 - 苯丙氨酸（fMLP）刺激的呼吸爆发反应；然而，引发机制尚不清楚。通过磷脂酶A2激活释放花生四烯酸（AA）以及5 - 脂氧合酶对其进行代谢，是PMN在炎症过程中的重要活动，可能参与引发刺激过程。因此，我们研究了二酰基甘油和烷基酰基甘油作为人中性粒细胞中AA释放和代谢引发剂的能力。在用[³H]AA对PMN磷脂进行预标记后，通过在fMLP刺激前将人PMN与二酰基甘油1 - 油酰基 - 2 - 乙酰甘油（OAG）或其烷基酰基类似物1 - O - δ9 - 十八碳烯基 - 2 - 乙酰甘油（EAG）孵育来测试引发作用。单独的fMLP（1微摩尔）、OAG（20微摩尔）或EAG（20微摩尔）几乎不会引起或根本不会引起标记AA的释放。然而，在用相同浓度的OAG或EAG引发PMN后，用1微摩尔fMLP刺激会导致细胞磷脂中6 - 8%的[³H]AA迅速释放（1分钟后达到峰值）；两种甘油二酯的总释放量相似。用OAG引发细胞还会增强随后fMLP刺激时释放的AA向白三烯B4（LTB4）和5 - 羟基二十碳四烯酸（5 - HETE）的转化，但在EAG引发的PMN中AA代谢产物并未增加。如果用钙离子载体A23187（它直接导致AA释放以及LTB4和5 - HETE的产生）替代fMLP，两种甘油二酯引发作用再次增强了[³H]AA的释放，但只有OAG引发增加了脂氧合酶活性。实际上，EAG预处理显著降低了LTB4和5 - HETE的产生。因此，两种甘油二酯均引发膜磷脂中AA的释放，但对AA随后的代谢具有相反作用。
