Surette M E, Palmantier R, Gosselin J, Borgeat P
Centre de Recherche en Inflammation, Immunologie et Rhumatologie, Centre de Recherche du CHUL, Ste. Foy, Québec, Canada.
J Exp Med. 1993 Oct 1;178(4):1347-55. doi: 10.1084/jem.178.4.1347.
Stimulation of heparinized blood with 1 microM formyl-methionyl-leucyl-phenylalanine (FMLP) resulted in the formation of < 30 pmol/ml plasma of 5-lipoxygenase (5-LO) products. The preincubation of blood with 1 microgram/ml of lipopolysaccharide (LPS) (Escherichia coli 0111-B4) for 30 min before stimulation with FMLP resulted in the accumulation of 250-300 pmol of 5-LO products per ml plasma. The major products detected were leukotriene B4 and (5S)-hydroxy-6,8,11,14-eicosatetraenoic acid which were produced in equivalent amounts. The priming activity was detectable with as little as 1-10 ng LPS per ml blood and was optimal using 1-10 micrograms LPS/ml blood. The priming for 5-LO product synthesis was optimal after 20-30 min of preincubation with LPS and declined at preincubation times > 30 min. The priming effect of LPS was also observed using the complement fragment C5a or interleukin 8 as agonists. Polymorphonuclear leukocytes (PMN) and peripheral blood mononuclear cells accounted for 80 and 20% of the synthesis of 5-LO products, respectively. The ability of LPS to prime isolated PMN was dependent on the presence of plasma and was inhibited by the anti-CD14 antibody IOM2, indicating a CD14-dependent priming mechanism. The priming of whole blood with tumor necrosis factor alpha (TNF-alpha) and LPS was additive and the presence of mononuclear cells did not enhance the ability of LPS to prime PMN, indicating that the priming activity of LPS is independent of LPS-induced TNF-alpha synthesis. The mechanism by which LPS enhance 5-LO product synthesis in PMN was investigated. Treatment of PMN with LPS strongly enhanced the release of arachidonic acid after stimulation with FMLP. The release of arachidonic acid was optimal 2-3 min after stimulation with FMLP, attaining levels 5-15-fold greater than those observed in unprimed cells stimulated with FMLP. These results demonstrate that LPS dramatically increases the ability of blood to generate 5-LO products, and support the putative role of leukotrienes in pathological states involving LPS.
用1微摩尔/升的甲酰甲硫氨酰亮氨酰苯丙氨酸（FMLP）刺激肝素化血液，导致每毫升血浆中5-脂氧合酶（5-LO）产物的生成量<30皮摩尔。在用FMLP刺激前，将血液与1微克/毫升的脂多糖（LPS）（大肠杆菌0111-B4）预孵育30分钟，导致每毫升血浆中积累250 - 300皮摩尔的5-LO产物。检测到的主要产物是白三烯B4和（5S）-羟基-6,8,11,14-二十碳四烯酸，它们的生成量相当。每毫升血液中低至1 - 10纳克的LPS就能检测到启动活性，使用1 - 10微克/毫升血液时启动活性最佳。用LPS预孵育20 - 30分钟后，5-LO产物合成的启动效果最佳，预孵育时间>30分钟时启动效果下降。使用补体片段C5a或白细胞介素8作为激动剂时，也观察到了LPS的启动作用。多形核白细胞（PMN）和外周血单核细胞分别占5-LO产物合成的80%和20%。LPS启动分离的PMN细胞的能力依赖于血浆的存在，并被抗CD14抗体IOM₂抑制，表明存在一种依赖CD14的启动机制。用肿瘤坏死因子α（TNF-α）和LPS对全血进行启动具有累加效应，单核细胞的存在并未增强LPS启动PMN的能力，这表明LPS的启动活性独立于LPS诱导的TNF-α合成。研究了LPS增强PMN中5-LO产物合成的机制。用LPS处理PMN后，在用FMLP刺激时花生四烯酸的释放显著增强。在用FMLP刺激后2 - 3分钟，花生四烯酸的释放达到最佳水平，比未启动的FMLP刺激细胞中观察到的水平高5 - 15倍。这些结果表明，LPS显著增加了血液生成5-LO产物的能力，并支持了白三烯在涉及LPS的病理状态中的假定作用。
