Department for Cellular Microbiology, Max Planck Institute for Infection Biology, Berlin, Germany.
J Innate Immun. 2009;1(3):181-93. doi: 10.1159/000205281. Epub 2009 Feb 26.
Neutrophil extracellular traps (NETs) play an important role in innate immunity to microbial infections. NETs have been described in several species, but the molecular details of NET formation and their role in infection has not been addressed, partly because we lack optimal experimental models. Here we describe tools to investigate NET formation in neutrophils isolated from mice. Upon in vitro stimulation of wild-type mouse neutrophils with PMA, we analyzed 3 important steps in the process of NET formation: reactive oxygen species (ROS) production, NET cell death and NET release. As expected, neutrophils from NADPH oxidase-deficient mice failed to produce ROS and did not die nor release NETs upon stimulation. We found that neutrophils from several mouse strains produced NETs with different efficiency and that NET formation correlated with the amount of ROS produced. Activation with Candida albicans also resulted in ROS production and NET cell death. The hyphal form of this fungus induced NETs more effectively than the yeast form. With this work, we provide tools to study in vitro NET assembly in the mouse system.
中性粒细胞胞外诱捕网（NETs）在先天免疫微生物感染中起着重要作用。在几种物种中已经描述了 NETs，但其形成的分子细节及其在感染中的作用尚未得到解决，部分原因是我们缺乏最佳的实验模型。在这里，我们描述了从小鼠中分离的中性粒细胞中研究 NET 形成的工具。在用 PMA 体外刺激野生型小鼠中性粒细胞后，我们分析了 NET 形成过程中的 3 个重要步骤：活性氧（ROS）的产生、NET 细胞死亡和 NET 的释放。正如预期的那样，NADPH 氧化酶缺陷型小鼠的中性粒细胞在刺激时不能产生 ROS，也不会死亡或释放 NETs。我们发现，几种小鼠品系的中性粒细胞产生 NET 的效率不同，并且 NET 的形成与产生的 ROS 量相关。用白色念珠菌激活也会导致 ROS 的产生和 NET 细胞的死亡。这种真菌的菌丝形式比酵母形式更有效地诱导 NETs。通过这项工作，我们提供了在小鼠系统中研究体外 NET 组装的工具。
