The aim of this study was to evaluate the role of angiotensin-converting enzyme 1 (ACE1) in H2O2-induced trophoblast cell injury and the potential molecular mechanisms. Oxidative stress was modeled by exposing HTR-8/SVneo cells to 200 µM H2O2. Western blot and real-time quantitative PCR methods were used to detect protein and mRNA expression level of ACE1 in chorionic villus tissue and trophoblast HTR-8/SVneo cell. Inhibition of ACE1 expression was achieved by transfection with small interfering RNA. Then flow cytometry, Cell Counting Kit-8, and Transwell assay was used to assess apoptosis, viability, and migration ability of the cells. Reactive oxygen species (ROS) were detected by fluorescent probes, and malondialdehyde (MDA), superoxide dismutase (SOD), and reduced glutathione (GSH) activities were determined by corresponding detection kits. Angiotensin-converting enzyme 1 expression was upregulated in chorionic villus tissue of patients with missed abortion (MA) compared with individuals with normal early pregnancy abortion. H2O2 induced elevated ACE1 expression in HTR-8/SVneo cells, promoted apoptosis, and inhibited cell viability and migration. Knockdown of ACE1 expression inhibited H2O2-induced effects to enhance cell viability and migration and suppress apoptosis. Additionally, H2O2 stimulation caused increased levels of ROS and MDA and decreased SOD and GSH activity in the cells, whereas knockdown of ACE1 expression led to opposite changes of these oxidative stress indicators. Moreover, knockdown of ACE1 attenuated the inhibitory effect of H2O2 on the Nrf2/HO-1 pathway. Angiotensin-converting enzyme 1 was associated with MA, and it promoted H2O2-induced injury of trophoblast cells through inhibiting the Nrf2 pathway. Therefore, ACE1 may serve as a potential therapeutic target for MA.
Graphical Abstract
In this study, we collected villous tissue from missed miscarriage patients (missed abortion (MA) group) and normal early pregnancy miscarriage patients (normal group). Simultaneously in vitro, angiotensin-converting enzyme 1 (ACE1) was transfected into HTR-8/SVneo cells and exposed to 200 µ M H2O2. Use RT qPCR to detect the expression level of ACE1. Western blot is used to detect the protein expression levels of Nrf2 (nucleus), ACE1, Keap1, and HO-1. Flow cytometry and Transwell were used to detect apoptosis level and migration ability of cells, respectively. Fluorescent probes were used to detect reactive oxygen species (ROS) activity. The level of malondialdehyde (MDA) and the activity of superoxide dismutase (SOD) and glutathione (GSH) in HTR-8/SVneo cells were detected by biochemical kit. Our results showed that ACE1 was upregulated in the villous tissue of MA patients compared with normal early miscarriage patients. Knocking down ACE1 expression can inhibit H2O2-induced cell apoptosis, reduce ROS and MDA levels, promote SOD and GSH activity, and enhance cell viability and metastasis ability. In addition, knocking down ACE1 can weaken the inhibitory effect of H2O2 on the Nrf2/HO-1 pathway in cells.
