Gestational diabetes mellitus (GDM) is a metabolic syndrome occurring in pregnant women and increases the risk of placental dysplasia. Neutrophil extracellular traps (NETs) may play a critical role in placental dysplasia. NETosis (neutrophil cell death by NET release) depends on NADPH/ROS pathway. In view of the adiponectin which is widely believed to be reduced in GDM patients suppresses NADPH oxidase and ROS generation of neutrophil. We speculate that increased NET release is associated with hypoadiponectinemia. Trophoblast apoptosis is significantly increased in GDM patients, but it is not clear whether NETs promotes cell apoptosis. This study aims to reveal the mechanism of Neutrophil Extracellular Traps generation and their role in trophoblast apoptosis in Gestational Diabetes Mellitus. We investigated the generation of NETs by cell-free DNA (cf-DNA) quantification, live-cell imaging, and reactive oxygen species (ROS) measurement. ERK1/2 and p38 MAPK signalling pathway proteins were detected by western blotting. The Cell Counting Kit-8 (CCK-8) assay, flow cytometry, and western blotting were performed to explore the effects of NETs on trophoblast apoptosis. We found that adiponectin inhibited NET release by suppressing ROS production, and p38 MAPK and ERK1/2 proteins were involved in the process. Further, NETs promoted trophoblast apoptosis by activating the ROS-dependent mitochondrial pathway, which is mediated by ERK1/2 signalling. The current study demonstrated that hypoadiponectinemia is the cause of NETs formation and NETs promoting trophoblast apoptosis.

Despite the widespread use of antiplatelets and anticoagulants, women with antiphospholipid syndrome (APS) may face pregnancy complications associated with placental dysplasia. Neutrophil extracellular traps (NETs) are involved in the pathogenesis of many autoimmune diseases, including vascular APS; however, their role in obstetric APS is unclear. Herein, we investigated the role of NETs by quantifying cell-free DNA and NET marker levels. Live-cell imaging was used to visualize NET formation, and MAPK signalling pathway proteins were analysed. Cell migration, invasion and tube formation assays were performed to observe the effects of NETs on trophoblasts and human umbilical vein endothelial cells (HUVECs). The concentrations of cell-free DNA and NETs in sera of pregnant patients with APS were elevated compared with that of healthy controls (HCs) matched to gestational week. APS neutrophils were predisposed to spontaneous NET release and IgG purified from the patients (APS-IgG) induced neutrophils from HCs to release NETs. Additionally, APS-IgG NET induction was abolished with inhibitors of reactive oxygen species, AKT, p38 MAPK and ERK1/2. Moreover, NETs were detrimental to trophoblasts and HUVECs. In summary, APS-IgG-induced NET formation deserves further investigation as a potential novel therapeutic target in obstetrical APS.

Obstetric antiphospholipid syndrome (OAPS) is mediated by antiphospholipid antibodies (aPLs, and anti-β2 glycoprotein I antibody is the main pathogenic antibody), and recurrent abortion, preeclampsia, foetal growth restriction and other placental diseases are the main clinical characteristics of placental pathological pregnancy. It is a disease that seriously threatens the health of pregnant women. Hydroxychloroquine (HCQ) was originally used as an anti-malaria drug and has now shown benefit in refractory OAPS where conventional treatment has failed, with the expectation of providing protective clinical benefits for both the mother and foetus. However, its efficacy and mechanism of action are still unclear. After clinical data were collected to determine the therapeutic effect, human trophoblast cells in early pregnancy were prepared and treated with aPL. After the addition of HCQ, the proliferation, invasion, migration and tubule formation of the trophoblast cells were observed so that the therapeutic mechanism of HCQ on trophoblast cells could be determined. By establishing an obstetric APS mouse model similar to the clinical situation, we were able to detect the therapeutic effect of HCQ on pathological pregnancy. The normal function of trophoblast cells is affected by aPL. Antibodies reduce the ability of trophoblast cells to invade and migrate and can impair tubule formation, which are closely related to placental insufficiency. HCQ can partially reverse these side effects. In the OAPS mouse model, we found that HCQ prevented foetal death and reduced the incidence of pathological pregnancy. Therefore, HCQ can improve pregnancy outcomes and reverse the aPL inhibition of trophoblast disease. In OAPS, the use of HCQ needs to be seriously considered.

Chorioamnionitis (CAM), as a common intrauterine infectious disease, is the leading cause of premature birth, stillbirth, neonatal infection and sepsis. The formyl peptide receptor 2 (FPR2) is a member of GPCRs widely distributed in a variety of tissues and is associated with many inflammatory diseases. With the discovery of FPR2 in human placenta, the possibility of exploring the function of FPR2 in obstetrics is evolving. The Resolvin D1 (RvD1) plays an important role in the resolution of inflammation by combining with FPR2. In this study, we evaluated the role of FPR2 and RvD1 in CAM, not only in the human placenta but also in mouse models. The expression of FPR2 increased in the placenta of CAM patients and the downstream PPARγ/NF-κB signalling changed accordingly. Moreover, Fpr2-/- mice were highly susceptible to LPS, displaying a worse CAM symptom, compared with WT mice. By establishing a model of trophoblast inflammation in vitro, it was confirmed that RvD1 rescued the effect of LPS on inflammation by combining with FPR2 and its downstream PPARγ/NF-κB pathway. Otherwise, RvD1 improved the preterm labour in a mouse model of CAM induced by LPS. Altogether, these findings show that RvD1 alleviated the inflammation of trophoblast in vivo and in vitro through FPR2/PPARγ/NF-κB pathway, suggesting RvD1/FPR2 might be a novel therapeutic strategy to alleviate CAM.

Background and aims: Preeclampsia (PE) is the leading cause of maternal and fetal morbidity and mortality worldwide. Apoptosis of trophoblast cells induced by oxidative stress is a principal reason of placental injury in PE. 6-Gingerol, an antioxidant from ginger, plays an important role in many disease models, but its effect on obstetric diseases has not been elucidated. In this study, we investigated the protective effect of 6-gingerol against placental injury. Methods: In vitro hypoxia/reoxygenation (H/R) model of HTR8/Svneo cells and preeclamptic mice model were established to simulate PE. The effects of 6-Gingerol on PE were evaluated by morphological detection, biochemical analysis, and Western blot. Results: We found that H/R treatment induced cell apoptosis, increased the production of reactive oxygen species, malondialdehyde and lactate dehydrogenase, and decreased superoxide dismutase in trophoblast. In addition, the polarization of mitochondrial membrane potential and the cellular calcium flux were also destroyed under H/R condition, which also activated BCL2-interacting protein 3 (BNIP3) and provoked excessive mitophagy. Importantly, 6-Gingerol reversed these corrosive effects. Furthermore, the placenta damage in PE-like mouse caused by the cell apoptosis, oxidative stress and mitophagy was mitigated by 6-Gingerol. Conclusion: These findings suggest that 6-Gingerol exerts a protective effect against placental injury in PE by reducing oxidative stress and inhibiting excessive mitophagy caused by mitochondrial dysfunction.