Department of Obstetrics and Gynaecology, Shandong Provincial Hospital, Shandong University, Jinan, Shandong, China.
Key Laboratory of Birth Regulation and Control Technology of National Health Commission of China, Shandong Provincial Maternal and Child Health Care Hospital, Jinan, Shandong, China.
Am J Reprod Immunol. 2023 Mar;89(3):e13656. doi: 10.1111/aji.13656. Epub 2022 Dec 1.
Premature birth is a common obstetric complication but its pathogenesis is unclear. Inflammation at the maternal-fetal interface in preterm labor leads to the infiltration of neutrophils, which promotes inflammatory responses and induces the degradation of extracellular matrix and cell apoptosis, thus contributing to preterm labor. It is unclear whether neutrophil extracellular traps (NETs), a functional form of neutrophils, are involved in preterm labor.
After collecting amniotic membranes from research objects, we localized NETs by immunofluorescence and evaluated the expression of matrix metalloproteinase (MMP)-9 and MMP-2 by western blotting. Primary human amniotic epithelial cells (hAECs) subjected to treatment with NETs, 5-ethynyl-20-deoxyuridine cell proliferation assay, lactate dehydrogenase (LDH) assay, western blotting, and flow cytometry apoptosis assay were used to determine the effects of NETs on hAECs. We also elucidated possible mechanisms underlying the effects.
Compared with normal term women, NETs infiltration and MMP-9 expression in the amniotic membrane from preterm women had increased. Thereafter, NETs might suppress the proliferation and promote the apoptosis of hAECs. Furthermore, after NETs treatment, the mitochondrial membrane potential was significantly decreased, ERK1/2 phosphorylation expression was upregulated and reactive oxygen species (ROS) production was increased in hAECs. Changes in cell proliferation, LDH release, and cell apoptosis level due to NETs could be reversed by ROS inhibitor or ERK phosphorylation inhibitors.
NETs can promote the apoptosis of hAECs via ERK1/2 pathways dependent on ROS release.
早产是一种常见的产科并发症，但发病机制尚不清楚。在早产时的母体-胎儿界面的炎症导致中性粒细胞的浸润，从而促进炎症反应，并诱导细胞外基质的降解和细胞凋亡，从而导致早产。中性粒细胞胞外诱捕网（NETs）是否参与早产尚不清楚。
研究对象的羊膜膜收集后，我们通过免疫荧光定位 NETs，并通过蛋白质印迹评估基质金属蛋白酶（MMP）-9 和 MMP-2 的表达。NETs 处理的原代人羊膜上皮细胞（hAECs）、5-乙炔基-20-脱氧尿苷细胞增殖测定、乳酸脱氢酶（LDH）测定、蛋白质印迹和流式细胞术凋亡测定用于确定 NETs 对 hAECs 的影响。我们还阐明了其作用的可能机制。
与正常足月分娩的妇女相比，早产妇女的羊膜膜中 NETs 浸润和 MMP-9 表达增加。此后，NETs 可能抑制 hAECs 的增殖并促进其凋亡。此外，NETs 处理后，hAECs 中的线粒体膜电位显著降低，ERK1/2 磷酸化表达上调，活性氧（ROS）生成增加。ROS 抑制剂或 ERK 磷酸化抑制剂可逆转 NETs 引起的细胞增殖、LDH 释放和细胞凋亡水平的变化。
NETs 可以通过依赖 ROS 释放的 ERK1/2 途径促进 hAECs 的凋亡。
