Fonteh A N, Bass D A, Marshall L A, Seeds M, Samet J M, Chilton F H
Department of Internal Medicine, Bowman Gray School of Medicine, Winston-Salem, NC 27157.
J Immunol. 1994 Jun 1;152(11):5438-46.
This study investigated the role of secretory PLA2 (sPLA2) in arachidonic acid (AA) release and lipid mediator generation in mouse bone marrow-derived mast cells. Initial studies indicated that mast cells contain multiple PLA2 activities and secreted PLA2 activity upon Ag stimulation. The secreted PLA2 activity is blocked by DTT and by a group II PLA2-neutralizing Ab. Mast cells incubated with groups I or II PLA2 selectively release polyunsaturated fatty acids, such as AA, into supernatant fluids. The bulk of AA released by sPLA2 is derived from phosphatidylethanolamine. The fatty acids released by extracellular PLA2 mimic those found in supernatant fluids after Ag stimulation of mast cells. Incubation of mast cells with PLA2 generates cyclooxygenase (CO) products but no 5-lipoxygenase (5-LO) products. Ag, but not PLA2, induces the translocation of 5-LO to cellular membranes and the formation of 5-LO products. The addition of sPLA2 to Ag-stimulated mast cells increases the synthesis of 5-LO products. Octadeuterated AA (2H8AA) added to the outside of cells to trace extracellular AA metabolism is rapidly converted to CO products. Addition of sPLA2, or Ag in combination with 2H8AA reduces the quantity of 2H8AA converted to deuterated CO products, when compared with adding 2H8AA alone. The reduction of deuterated CO products can be accounted for with increases in nondeuterated CO products. 2H8AA is only converted to 5-LO produces when mast cells are activated with Ag. The amount of 2H8AA converted to deuterated 5-LO products is reduced by the addition of PLA2 to Ag-stimulated cells. The competitive formation of deuterated and nondeuterated products observed when mast cells are incubated with 2H8AA, indicates that extracellular AA released by sPLA2 is used for both CO and 5-LO product formation. Taken together, these data reveal a role for sPLA2 in the release of AA and demonstrate that AA released by this mechanism is used for eicosanoid generation.
本研究调查了分泌型磷脂酶A2（sPLA2）在小鼠骨髓来源肥大细胞中花生四烯酸（AA）释放及脂质介质生成过程中的作用。初步研究表明，肥大细胞含有多种磷脂酶A2活性，且在抗原刺激后会分泌磷脂酶A2活性。分泌的磷脂酶A2活性可被二硫苏糖醇（DTT）和一种II型磷脂酶A2中和抗体所阻断。用I型或II型磷脂酶A2孵育肥大细胞可选择性地将多不饱和脂肪酸，如AA，释放到上清液中。sPLA2释放的大部分AA来源于磷脂酰乙醇胺。细胞外磷脂酶A2释放出的脂肪酸与肥大细胞经抗原刺激后的上清液中发现的脂肪酸相似。用磷脂酶A2孵育肥大细胞会生成环氧化酶（CO）产物，但不会生成5-脂氧合酶（5-LO）产物。抗原而非磷脂酶A2可诱导5-LO转位至细胞膜并形成5-LO产物。将sPLA2添加到经抗原刺激的肥大细胞中可增加5-LO产物的合成。添加到细胞外以追踪细胞外AA代谢的十八氘代AA（2H8AA）会迅速转化为CO产物。与单独添加2H8AA相比，添加sPLA2或抗原与2H8AA组合会减少转化为氘代CO产物的2H8AA量。氘代CO产物的减少可通过非氘代CO产物的增加来解释。只有当肥大细胞用抗原激活时，2H8AA才会转化为5-LO产物。向经抗原刺激的细胞中添加磷脂酶A2会减少转化为氘代5-LO产物的2H8AA量。当肥大细胞与2H8AA孵育时观察到的氘代和非氘代产物的竞争性形成表明，sPLA2释放的细胞外AA可用于CO和5-LO产物的形成。综上所述，这些数据揭示了sPLA2在AA释放中的作用，并证明通过该机制释放的AA可用于类花生酸的生成。
