Downey G P, Butler J R, Tapper H, Fialkow L, Saltiel A R, Rubin B B, Grinstein S
Toronto Hospital, and Department of Medicine, University of Toronto, Ontario, Canada.
J Immunol. 1998 Jan 1;160(1):434-43.
Exposure of neutrophils to inflammatory stimuli such as the chemoattractant FMLP leads to activation of responses including cell motility, the oxidative burst, and secretion of proteolytic enzymes. A signaling cascade involving sequential activation of Raf-1, mitogen-activated protein kinase (MEK), and extracellular signal regulated kinase (ERK) is also rapidly activated after agonist exposure. The temporal relationship between these events suggests that the kinases may be involved in triggering the effector functions, but direct evidence of a causal relationship is lacking. To assess the role of the MEK/ERK pathway in the activation of neutrophil responses, we studied the effects of PD098059, a potent and selective inhibitor of MEK. Preincubation of human neutrophils with 50 microM PD098059 almost completely (>90%) inhibited the FMLP-induced activation of MEK-1 and MEK-2, the isoforms expressed by neutrophils. This dose of PD098059 virtually abrogated chemoattractant-induced tyrosine phosphorylation and activation of ERK-1 and ERK-2, implying that MEKs are the predominant upstream activators of these mitogen-activated protein kinases. Pretreatment of neutrophils with the MEK antagonist inhibited the oxidative burst substantially and phagocytosis only moderately. In addition, PD098059 antagonized the delay of apoptosis induced by exposure to granulocyte-macrophage CSF. However, the effects of PD098059 were selective, as it failed to inhibit other responses, including chemoattractant-induced exocytosis of primary and secondary granules, polymerization of F-actin, chemotaxis, or activation of phospholipase A2. We conclude that MEK and ERK contribute to the activation of the oxidative burst and phagocytosis, and participate in cytokine regulation of apoptosis.
中性粒细胞暴露于诸如趋化因子FMLP等炎性刺激下会引发包括细胞运动、氧化爆发以及蛋白水解酶分泌等反应的激活。在激动剂暴露后，一条涉及Raf-1、丝裂原活化蛋白激酶（MEK）和细胞外信号调节激酶（ERK）顺序激活的信号级联反应也会迅速被激活。这些事件之间的时间关系表明，这些激酶可能参与触发效应器功能，但缺乏因果关系的直接证据。为了评估MEK/ERK途径在中性粒细胞反应激活中的作用，我们研究了MEK的强效选择性抑制剂PD098059的作用。用50微摩尔的PD098059预孵育人中性粒细胞几乎完全（>90%）抑制了FMLP诱导的MEK-1和MEK-2的激活，MEK-1和MEK-2是中性粒细胞表达的亚型。这个剂量的PD098059实际上消除了趋化因子诱导的酪氨酸磷酸化以及ERK-1和ERK-2的激活，这意味着MEK是这些丝裂原活化蛋白激酶的主要上游激活剂。用MEK拮抗剂预处理中性粒细胞可显著抑制氧化爆发，对吞噬作用的抑制则较弱。此外，PD098059拮抗了暴露于粒细胞-巨噬细胞集落刺激因子诱导的凋亡延迟。然而，PD098059的作用具有选择性，因为它未能抑制其他反应，包括趋化因子诱导的初级和次级颗粒胞吐、F-肌动蛋白聚合、趋化作用或磷脂酶A2的激活。我们得出结论，MEK和ERK有助于氧化爆发和吞噬作用的激活，并参与细胞因子对凋亡的调节。
