Neutrophils migrating towards chemoattractant gradients amplify their recruitment range by releasing the secondary chemoattractant leukotriene B₄ (LTB₄) refs. ^1,2. We previously demonstrated that LTB₄ and its synthesizing enzymes, 5-lipoxygenase (5-LO), 5-LO activating protein (FLAP) and leukotriene A₄ hydrolase, are packaged and released in exosomes³. Here we report that the biogenesis of the LTB₄-containing exosomes originates at the nuclear envelope (NE) of activated neutrophils. We show that the neutral sphingomyelinase 1 (nSMase1)-mediated generation of ceramide-enriched lipid-ordered microdomains initiates the clustering of the LTB₄-synthesizing enzymes on the NE. We isolated and analysed exosomes from activated neutrophils and established that the FLAP/5-LO-positive exosome population is distinct from that of the CD63-positive exosome population. Furthermore, we observed a strong co-localization between ALIX and FLAP at the periphery of nuclei and within cytosolic vesicles. We propose that the initiation of NE curvature and bud formation is mediated by nSMase1-dependent ceramide generation, which leads to FLAP and ALIX recruitment. Together, these observations elucidate the mechanism for LTB₄ secretion and identify a non-conventional pathway for exosome generation.

向趋化梯度迁移的中性粒细胞通过释放次级趋化因子白三烯 B ₄ (LTB ₄ ) 参考文献扩大了它们的募集范围。^1,2。我们之前证明，LTB ₄及其合成酶、5-脂氧合酶 (5-LO)、5-LO 活化蛋白 (FLAP) 和白三烯 A ₄水解酶在外泌体中被包装和释放³。在这里，我们报告了含有 LTB ₄的外泌体的生物合成起源于活化的嗜中性粒细胞的核膜 (NE)。我们表明，中性鞘磷脂酶 1 (nSMase1) 介导的富含神经酰胺的脂质有序微域的生成启动了 LTB _4的聚类-NE 上的合成酶。我们从活化的中性粒细胞中分离并分析了外泌体，并确定 FLAP/5-LO 阳性外泌体群体不同于 CD63 阳性外泌体群体。此外，我们观察到 ALIX 和 FLAP 在细胞核周围和胞质囊泡内的强烈共定位。我们提出 NE 曲率和芽形成的启动是由 nSMase1 依赖性神经酰胺生成介导的，这导致 FLAP 和 ALIX 募集。_{总之，这些观察结果阐明了 LTB 4}分泌的机制，并确定了外泌体生成的非常规途径。