Extracellular traps (ETs) have been found to be an important strategy of mammals to immobilize and kill invading microorganisms. In the present study, we observed the formation of ETs in the hemocytes of marine mollusks Ruditapes philippinarum in response to challenge from bacteria Vibrio anguillarum, and examined the potential factors and signaling pathways underling this process. We detected an increase of reactive oxygen species (ROS) and myeloperoxidase (MPO) production during ETosis, accompanied by significantly up-regulated expression of ROS-related and MPO genes. The suppression of ETs structures by nicotinamide adenine dinucleotide phosphate (NADPH) oxidase inhibitor (diphenyleneiodonium chloride, DPI) and MPO inhibitor (aminobenzoic acid hydrazide, ABAH) further confirmed the essential roles ROS and MPO played in ETosis. Furthermore, ET production could be inhibited by phosphotidylinsitol-3-kinase (PI3K) inhibitor (LY294002) and extracellular regulated protein kinase (ERK) inhibitor (U0126), suggesting the idea that both the PI3K and ERK pathways were suggested to function during ETosis. In addition, the ETosis process was accompanied by enhancement of glycolysis-related enzymatic activities, e.g., pyruvate kinase (PK) and hexokinase (HK), and over-expression of the glycolysis-related genes, e.g., PK, HK and glucose transport protein (GLUT), indicating high involvement of glycolysis in the ETosis process. Furthermore, our scanning electron microscopy (SEM) observation and antibacterial activities test successfully showed the patterns how clam ETs entrapped and killed the invading V. anguillarum. Taken together, our results revealed that ETosis with bactericidal effect increased ROS, MPO and glycolysis level and carried out in a ROS-, MPO-, PI3K-ERK-dependent manner.

细胞外诱捕器 (ETs) 已被发现是哺乳动物固定和杀死入侵微生物的重要策略。在本研究中，我们观察到海洋软体动物Ruditapes philippinarum的血细胞中 ET 的形成，以应对细菌弧菌的挑战。，并检查了这一过程的潜在因素和信号通路。我们检测到 ETosis 期间活性氧 (ROS) 和髓过氧化物酶 (MPO) 的产生增加，伴随着 ROS 相关基因和 MPO 基因的显着上调。烟酰胺腺嘌呤二核苷酸磷酸 (NADPH) 氧化酶抑制剂 (diphenyleneiodonium chloride, DPI) 和 MPO 抑制剂 (氨基苯甲酸酰肼, ABAH) 对 ETs 结构的抑制进一步证实了 ROS 和 MPO 在 ETosis 中的重要作用。此外，磷脂酰肌醇-3-激酶 (PI3K) 抑制剂 (LY294002) 和细胞外调节蛋白激酶 (ERK) 抑制剂 (U0126) 可以抑制 ET 的产生，这表明 PI3K 和 ERK 途径都被认为在 ETosis 期间起作用。此外，ETosis 过程伴随着糖酵解相关酶活性的增强，例如丙酮酸激酶 (PK) 和己糖激酶 (HK)，以及糖酵解相关基因的过度表达，例如 PK、HK 和葡萄糖转运蛋白 (GLUT) ，表明糖酵解高度参与 ETosis 过程。此外，我们的扫描电子显微镜 (SEM) 观察和抗菌活性测试成功地展示了蛤 ET 如何捕获和杀死入侵者的模式五、鳗鲡。总之，我们的结果表明具有杀菌作用的 ETosis 增加了 ROS、MPO 和糖酵解水平，并以 ROS-、MPO-、PI3K-ERK 依赖性方式进行。