Cardiovascular Pharmacology Unit, Central Drug Research Institute (Council of Scientific and Industrial Research), Lucknow, India.
Nitric Oxide. 2010 Apr 1;22(3):226-34. doi: 10.1016/j.niox.2010.01.001. Epub 2010 Jan 11.
High availability of NO, oxidative stress and neutrophil extracellular trap (NETs) contents are often noticed at the site of inflammation/infection. Studies from this lab and others have reported NO mediated free radical generation from neutrophils; role of NO in NETs formation however remains undefined so far. The present study was therefore undertaken to explore the effect of NO donors on NET release from human neutrophils (PMNs), using confocal/scanning microscopy, measuring the extracellular DNA content and NET-bound elastase activity. Addition of NO donors (SNAP and SNP) to adhered PMNs led to a time and concentration dependent NETs release, which was blocked by N-acetyl cysteine, suggesting involvement of free radicals in NETs formation. Free radical formation by NO donors was assessed by using DCF-DA, DMPO-nitrone antibody and by p47 phox migration to the neutrophils membrane. NO mediated formation of free radicals and NETs was significantly reduced by the pretreatment of neutrophils with diphenyleneiodonium (DPI), a NADPH-oxidase inhibitor and 4-aminobenzoic acid hydrazide (ABAH), a myeloperoxidase inhibitor, suggesting role of enzymatic free radical generation by NO donors. We thus demonstrate that NO by augmenting free radical formation in human neutrophils mediates NETs release.
高浓度的一氧化氮（NO）、氧化应激和中性粒细胞胞外诱捕网（NETs）内容物通常在炎症/感染部位被观察到。本实验室和其他实验室的研究报告称，NO 可从中性粒细胞中产生自由基；然而，NO 在 NETs 形成中的作用目前尚未确定。因此，本研究旨在通过共聚焦/扫描显微镜、测量细胞外 DNA 含量和 NET 结合弹性蛋白酶活性，来探索 NO 供体对人中性粒细胞（PMN）中 NET 释放的影响。将 NO 供体（SNAP 和 SNP）添加到附着的 PMN 中，导致 NETs 释放呈时间和浓度依赖性，这种释放被 N-乙酰半胱氨酸阻断，表明自由基参与了 NETs 的形成。通过使用 DCF-DA、DMPO-硝酮抗体和 p47 phox 向中性粒细胞膜迁移来评估 NO 供体产生的自由基。NO 介导的自由基和 NETs 的形成通过用 NADPH 氧化酶抑制剂二苯乙烯碘（DPI）和髓过氧化物酶抑制剂 4-氨基苯甲酰肼（ABAH）预处理中性粒细胞而显著减少，表明 NO 供体通过酶促自由基生成介导了 NETs 的形成。因此，我们证明了 NO 通过增强人中性粒细胞中的自由基形成来介导 NETs 的释放。
