Clinical Laboratory, The Second Affiliated Hospital of Harbin Medical University, Harbin, China.
Department of Anesthesiology, The First Affiliated Hospital of Harbin Medical University, Harbin, China.
Cell Biochem Funct. 2019 Mar;37(2):56-61. doi: 10.1002/cbf.3363. Epub 2019 Jan 30.
Neutrophils participate in the regulation of pathogens by phagocytosis as well as by generating neutrophil extracellular traps (NETs). Antiphospholipid antibodies, particularly those targeting beta-2-glycoprotein I (β2GPI), stimulate monocytes, platelets, and endothelial cells with prothrombotic participation. This study aimed to explore NET generation in response to anti-β2GPI/β2GPI. A series of experiments involving the separation of primary human leukocytes, NETosis quantification using propidium iodide, exploration of NETosis by fluorescence microscopy, western blotting, examination of free Zn using FluoZin-3, and reactive oxygen species (ROS) examination with dihydrorhodamine 123 were performed in this study. We found that anti-β2GPI/β2GPI triggered NETosis, resembling phorbol 12-myristate 13-acetate (PMA)-induced NETosis in magnitude and morphology. The anti-β GPI/β GPI complex in isolation stimulated NETs without relying on p38, protein kinase B (AKT), extracellular signal-related kinase (ERK) 1/2, and zinc signals. NET generation was unaffected by the NADPH oxidase suppressor DP1. The anti-β GPI/β GPI complex stimulated ROS generation without relying on NADPH oxidase, which may participate in NET generation triggered via the anti-β GPI/β GPI complex. In summary, our results indicate that the anti-β GPI/β GPI complex reinforced NET generation by relying on ROS. THE SIGNIFICANCE OF THE PAPER IN THE CONTEXT OF CURRENT KNOWLEDGE: Neutrophils as one of the first lines of defence and essential in the response to pathogen invasion. They eradicate bacteria via phagocytosis or by releasing antimicrobial proteins in degranulation. In this study, we explored the capability of anti-β GPI/β GPI to stimulate NETosis, demonstrating that anti-β GPI/β GPI is a promising method for triggering NET. Anti-β GPI/β GPI induced ROS generation without relying on NADPH oxidase, which contributes to NETosis independently of ERK1/2, Zn , or AKT. Our results showed that anti-β2GPI/β2GPI triggered NETosis, resembling PMA-induced NETosis in magnitude as well as morphology. The anti-β GPI/β GPI complex in isolation stimulated NETs without relying on p38, AKT, ERK1/2, or zinc signals. The anti-β GPI/β GPI complex stimulated ROS generation without relying on NADPH oxidase, which may participate in NET generation triggered via the anti-β GPI/β GPI complex.
中性粒细胞通过吞噬作用以及产生中性粒细胞细胞外陷阱 (NETs) 来参与病原体的调节。抗磷脂抗体，特别是针对β-2-糖蛋白 I (β2GPI) 的抗体，会刺激单核细胞、血小板和内皮细胞产生促血栓形成作用。本研究旨在探索针对抗-β2GPI/β2GPI 的 NET 生成。本研究进行了一系列实验，包括分离原代人白细胞、使用碘化丙啶定量 NETosis、通过荧光显微镜探索 NETosis、进行 Western blot 分析、使用 FluoZin-3 检测游离锌以及使用二氢罗丹明 123 检测活性氧 (ROS)。我们发现抗-β2GPI/β2GPI 触发了 NETosis，其规模和形态类似于佛波醇 12-肉豆蔻酸 13-乙酸酯 (PMA) 诱导的 NETosis。单独的抗-β GPI/β GPI 复合物在不依赖 p38、蛋白激酶 B (AKT)、细胞外信号调节激酶 (ERK) 1/2 和锌信号的情况下刺激 NET。NET 生成不受 NADPH 氧化酶抑制剂 DP1 的影响。抗-β GPI/β GPI 复合物刺激 ROS 生成而不依赖 NADPH 氧化酶，这可能参与了通过抗-β GPI/β GPI 复合物触发的 NET 生成。总之，我们的结果表明，抗-β GPI/β GPI 复合物通过依赖 ROS 来增强 NET 生成。
中性粒细胞作为第一道防线之一，在应对病原体入侵时至关重要。它们通过吞噬作用或脱颗粒释放抗菌蛋白来消灭细菌。在这项研究中，我们探索了抗-β GPI/β GPI 刺激 NETosis 的能力，表明抗-β GPI/β GPI 是触发 NET 的一种有前途的方法。抗-β GPI/β GPI 诱导 ROS 生成而不依赖 NADPH 氧化酶，这有助于 NETosis 的独立于 ERK1/2、Zn 或 AKT。我们的结果表明，抗-β2GPI/β2GPI 触发了 NETosis，其规模和形态与 PMA 诱导的 NETosis 相似。单独的抗-β GPI/β GPI 复合物刺激 NET 生成而不依赖于 p38、AKT、ERK1/2 或锌信号。抗-β GPI/β GPI 复合物刺激 ROS 生成而不依赖 NADPH 氧化酶，这可能参与了通过抗-β GPI/β GPI 复合物触发的 NET 生成。
