Neutrophils are a part of the innate immune system and produce reactive oxygen species (ROS) to extinguish pathogens. The major source of ROS in neutrophils is NADPH oxidase, which is fueled by NADPH generated via the pentose phosphate pathway; however, it is unclear how other accessory glucose metabolism pathways and mitochondrial activity influence the respiratory burst. We examined the temporal dynamics of the respiratory burst and delineated how metabolism changes over time after neutrophil activation. Bone marrow-derived neutrophils were stimulated with phorbol 12-myristate 13-acetate (PMA), and the respiratory burst was measured via extracellular flux analysis. Metabolomics experiments utilizing 13C6-glucose highlighted the activation of glycolysis as well as ancillary pathways of glucose metabolism in activated neutrophils. PMA stimulation acutely increased 13C enrichment into glycerol 3-phosphate (G3P) and citrate, whereas increases in 13C enrichment in the glycogen intermediate, UDP-hexose, and end products of the hexosamine and serine biosynthetic pathways occurred only during the late phase of the oxidative burst. Targeted inhibition of the G3P shuttle, glycogenolysis, serine biosynthesis, and mitochondrial respiration demonstrated that the G3P shuttle contributes to the general magnitude of ROS production; that glycogen contributes solely to the early respiratory burst; and that the serine biosynthetic pathway activity and Complex III-driven mitochondrial activity influence respiratory burst duration. Collectively, these results show that the neutrophil oxidative burst is highly dynamic, with coordinated changes in metabolism that control the initiation, magnitude, and duration of ROS production.

中文翻译：

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中性粒细胞氧化爆发的相位分配由葡萄糖代谢和线粒体活性的辅助途径协调。


中性粒细胞是先天免疫系统的一部分，可产生活性氧 （ROS） 来消灭病原体。中性粒细胞中 ROS 的主要来源是 NADPH 氧化酶，它由通过磷酸戊糖途径产生的 NADPH 提供动力;然而，目前尚不清楚其他辅助葡萄糖代谢途径和线粒体活性如何影响呼吸爆发。我们检查了呼吸爆发的时间动力学，并描述了中性粒细胞激活后新陈代谢如何随时间变化。用佛波醇 12-肉豆蔻酸酯 13-乙酸酯 （PMA） 刺激骨髓来源的中性粒细胞，并通过细胞外通量分析测量呼吸爆发。利用 13C6-葡萄糖的代谢组学实验强调了活化的中性粒细胞中糖酵解的激活以及葡萄糖代谢的辅助途径。PMA 刺激急剧增加了 13C 富集到 3-磷酸甘油 （G3P） 和柠檬酸盐中，而糖原中间体、UDP-己糖以及己糖胺和丝氨酸生物合成途径的终末产物中 13C 富集的增加仅发生在氧化爆发的后期。G3P 穿梭、糖原分解、丝氨酸生物合成和线粒体呼吸的靶向抑制表明，G3P 穿梭有助于 ROS 产生的一般量级;糖原仅有助于早期呼吸爆发;丝氨酸生物合成途径活性和复合物 III 驱动的线粒体活性影响呼吸爆发持续时间。总的来说，这些结果表明，中性粒细胞氧化爆发是高度动态的，代谢的协调变化控制着 ROS 产生的开始、幅度和持续时间。