Naids F L, Rest R F
Department of Microbiology and Immunology, Hahnemann University School of Medicine, Philadelphia, Pennsylvania 19102-1192.
Infect Immun. 1991 Dec;59(12):4383-90. doi: 10.1128/iai.59.12.4383-4390.1991.
Nonopsonized gonococci possessing opacity-associated (Opa; previously PII) outer membrane proteins stimulate neutrophils to undergo a vigorous oxidative response when measured by luminol-dependent chemiluminescence (LDCL). In these studies, we characterized the mechanism of this stimulation. No gonococci that we tested induced measurable release of neutrophil superoxide anion (O2-) or hydrogen peroxide (H2O2) as measured by reduction of cytochrome c or the oxidation of scopoletin, respectively. Neutrophils pretreated with gonococci and then exposed to phorbol myristate acetate, the chemotactic peptide formylmethionylleucylphenylalanine, or opsonized zymosan released levels of neutrophil O2- and H2O2 comparable to controls, indicating that gonococci were not preventing or inhibiting neutrophil O2- or H2O2 release. To ascertain a possible explanation for these seemingly contradictory observations (i.e., induction of LDCL, but no release of O2- or H2O2), we further characterized the ability of Opa+ gonococci to stimulate LDCL. By using 1 mM azide and 4 U of horseradish peroxidase to monitor extracellular LDCL selectively and 2,000 U of catalase to monitor intracellular LDCL selectively, we determined that greater than 80% of total gonococcus-induced neutrophil LDCL occurred intracellularly. In addition, neutrophils stimulated with Opa+ gonococci showed a marked increase in O2 uptake and hexose monophosphate shunt activity. We conclude that Neisseria gonorrhoeae induces neutrophil oxidative metabolism without causing release of detectable amounts of reactive oxygen intermediates into the surrounding milieu. The gonococcus apparently directs oxidase assembly and activity to the phagolysosomal membrane. This could be a mechanism by which extracellular gonococci persist for extended periods in vivo in the presence of high concentrations of neutrophils.
当通过鲁米诺依赖的化学发光法（LDCL）检测时，具有不透明相关（Opa；以前称为PII）外膜蛋白的非调理淋病奈瑟菌会刺激中性粒细胞发生强烈的氧化反应。在这些研究中，我们对这种刺激机制进行了表征。我们测试的淋病奈瑟菌均未诱导中性粒细胞超氧阴离子（O2-）或过氧化氢（H2O2）的可测量释放，分别通过细胞色素c的还原或 scopoletin 的氧化来测量。用淋病奈瑟菌预处理的中性粒细胞，然后暴露于佛波酯肉豆蔻酸酯乙酸盐、趋化肽甲酰甲硫氨酰亮氨酰苯丙氨酸或调理酵母聚糖，释放的中性粒细胞O2-和H2O2水平与对照组相当，表明淋病奈瑟菌并未阻止或抑制中性粒细胞O2-或H2O2的释放。为了确定对这些看似矛盾的观察结果（即诱导LDCL，但不释放O2-或H2O2）的可能解释，我们进一步表征了Opa+淋病奈瑟菌刺激LDCL的能力。通过使用1 mM叠氮化物和4 U辣根过氧化物酶选择性监测细胞外LDCL，以及2000 U过氧化氢酶选择性监测细胞内LDCL，我们确定超过80%的总淋病奈瑟菌诱导的中性粒细胞LDCL发生在细胞内。此外，用Opa+淋病奈瑟菌刺激的中性粒细胞显示O2摄取和己糖磷酸旁路活性显著增加。我们得出结论，淋病奈瑟菌诱导中性粒细胞氧化代谢，而不会导致可检测量的活性氧中间体释放到周围环境中。淋病奈瑟菌显然将氧化酶组装和活性导向吞噬溶酶体膜。这可能是细胞外淋病奈瑟菌在高浓度中性粒细胞存在下在体内长期持续存在的一种机制。
