Stolk J, Hiltermann T J, Dijkman J H, Verhoeven A J
Department of Pulmonology, University Hospital Leiden, The Netherlands.
Am J Respir Cell Mol Biol. 1994 Jul;11(1):95-102. doi: 10.1165/ajrcmb.11.1.8018341.
Phagocytes are able to generate reactive oxygen species by an activatable NADPH oxidase system. We investigated the inhibition of NADPH oxidase activation by a methoxy-substituted catechol, apocynin. Oxygen uptake by neutrophils incubated with 300 microM apocynin was completely inhibited at 7 min after addition of serum-treated zymosan (STZ), with a lagtime of inhibition of 2 to 3 min. The lagtime of effect of apocynin in neutrophils relatively deficient of myeloperoxidase was about 50% longer when compared with normal cells. Inhibition of the STZ-induced respiratory burst by apocynin was also observed in human eosinophils but not in human alveolar macrophages. Immunoblots of neutrophil membranes, isolated at 2 and 7 min after STZ stimulation of neutrophils, demonstrated translocation of the cytosolic oxidase components p47-phox and p67-phox to the membrane fraction. Translocation at 7 min after STZ stimulation was markedly reduced when the neutrophils had been incubated with 300 microM apocynin, but translocation was normal after 2 min of stimulation. These properties suggest that apocynin is an intracellular inhibitor of the assembly of NADPH oxidase in neutrophils and eosinophils and that apocynin requires conversion by peroxidases to exert its inhibitory effect. The capacity of neutrophils for intracellular killing of Staphylococcus aureus was not affected by apocynin. The potential therapeutic value of apocynin was demonstrated in vitro by its ability to protect secretory leukocyte proteinase inhibitor from oxidative inactivation by neutrophils.
吞噬细胞能够通过可激活的NADPH氧化酶系统产生活性氧物质。我们研究了一种甲氧基取代的儿茶酚——夹竹桃麻素对NADPH氧化酶激活的抑制作用。用300微摩尔夹竹桃麻素孵育的中性粒细胞在加入血清处理的酵母聚糖（STZ）后7分钟，其氧气摄取被完全抑制，抑制延迟时间为2至3分钟。与正常细胞相比，髓过氧化物酶相对缺乏的中性粒细胞中夹竹桃麻素作用的延迟时间长约50%。在人嗜酸性粒细胞中也观察到夹竹桃麻素对STZ诱导的呼吸爆发的抑制作用，但在人肺泡巨噬细胞中未观察到。在STZ刺激中性粒细胞后2分钟和7分钟分离的中性粒细胞膜的免疫印迹显示，胞质氧化酶成分p47 - phox和p67 - phox向膜部分易位。当用300微摩尔夹竹桃麻素孵育中性粒细胞时，STZ刺激后7分钟的易位明显减少，但刺激2分钟后易位正常。这些特性表明夹竹桃麻素是中性粒细胞和嗜酸性粒细胞中NADPH氧化酶组装的细胞内抑制剂，并且夹竹桃麻素需要过氧化物酶转化才能发挥其抑制作用。夹竹桃麻素不影响中性粒细胞对金黄色葡萄球菌的细胞内杀伤能力。夹竹桃麻素在体外的潜在治疗价值通过其保护分泌型白细胞蛋白酶抑制剂免受中性粒细胞氧化失活的能力得以证明。
