Billah M M, Bryant R W, Siegel M I
J Biol Chem. 1985 Jun 10;260(11):6899-906.
When human neutrophils, previously labeled in their phospholipids with [14C]arachidonate, were stimulated with the Ca2+-ionophore, A23187, plus Ca2+ in the presence of [3H]acetate, these cells released [14C]arachidonate from membrane phospholipids, produced 5-hydroxy-6,8,11,14-[14C]eicosatetraenoic acid (5-HETE) and 14C-labeled 5S,12R-dihydroxy-6-cis,8,10-trans, 14-cis-eicosatetraenoic acid ([14C]leukotriene B4), and incorporated [3H]acetate into platelet-activating factor (PAF, 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine). Ionophore A23187-induced formation of these radiolabeled products was greatly augmented by submicromolar concentrations of exogenous 5-hydroperoxy-6,8,11,14-eicosatetraenoic acid (5-HPETE), 5-HETE, and leukotriene B4. In the absence of ionophore A23187, these arachidonic acid metabolites were virtually ineffective. Nordihydroguaiaretic acid (NDGA) and several other lipoxygenase/cyclooxygenase inhibitors (butylated hydroxyanisole, 3-amino-1-(3-trifluoromethylphenyl)-2-pyrazoline and 1-phenyl-2-pyrazolidinone) caused parallel inhibition of [14C]arachidonate release and [3H]PAF formation in a dose-dependent manner. Specific cyclooxygenase inhibitors, such as indomethacin and naproxen, did not inhibit but rather slightly augmented the formation of these products. Furthermore, addition of 5-HPETE, 5-HETE, or leukotriene B4 (but not 8-HETE or 15-HETE) to neutrophils caused substantial relief of NDGA inhibition of [3H]PAF formation and [14C]arachidonate release. As opposed to [3H]acetate incorporation into PAF, [3H]lyso-PAF incorporation into PAF by activated neutrophils was little affected by NDGA. In addition, NDGA had no effect on lyso-PAF:acetyl-CoA acetyltransferase as measured in neutrophil homogenate preparations. It is concluded that in activated human neutrophils 5-lipoxygenase products can modulate PAF formation by enhancing the expression of phospholipase A2.
当先前用[14C]花生四烯酸标记磷脂的人中性粒细胞在[3H]乙酸存在的情况下用钙离子载体A23187加钙离子刺激时,这些细胞从膜磷脂中释放[14C]花生四烯酸,产生5-羟基-6,8,11,14-[14C]二十碳四烯酸(5-HETE)和14C标记的5S,12R-二羟基-6-顺式,8,10-反式,14-顺式二十碳四烯酸([14C]白三烯B4),并将[3H]乙酸掺入血小板活化因子(PAF,1-O-烷基-2-乙酰基-sn-甘油-3-磷酸胆碱)中。亚微摩尔浓度的外源性5-氢过氧-6,8,11,14-二十碳四烯酸(5-HPETE)、5-HETE和白三烯B4极大地增强了离子载体A23187诱导的这些放射性标记产物的形成。在没有离子载体A23187的情况下,这些花生四烯酸代谢产物几乎没有作用。去甲二氢愈创木酸(NDGA)和其他几种脂氧合酶/环氧化酶抑制剂(丁基化羟基茴香醚、3-氨基-1-(3-三氟甲基苯基)-2-吡唑啉和1-苯基-2-吡唑烷酮)以剂量依赖的方式平行抑制[14C]花生四烯酸的释放和[3H]PAF的形成。特异性环氧化酶抑制剂,如吲哚美辛和萘普生,并不抑制而是略微增强这些产物的形成。此外,向中性粒细胞中添加5-HPETE、5-HETE或白三烯B4(但不是8-HETE或15-HETE)可显著减轻NDGA对[3H]PAF形成和[14C]花生四烯酸释放的抑制作用。与[3H]乙酸掺入PAF不同,活化的中性粒细胞将[3H]溶血PAF掺入PAF的过程几乎不受NDGA的影响。此外,在中性粒细胞匀浆制剂中检测到,NDGA对溶血PAF:乙酰辅酶A乙酰转移酶没有影响。结论是,在活化的人中性粒细胞中,5-脂氧合酶产物可通过增强磷脂酶A2的表达来调节PAF的形成。