CX43 inhibit ferroptosis in villous and decidua tissues of patients with unexplained recurrent spontaneousabortion through the Nrf2/GPX4 signaling pathway

Purpose Approximately 50% of RSA cases are classified as unexplained recurrent spontaneous abortion (URSA), while effective treatment options are still limited.Despite its growing recognition in various diseases, the specific molecular mechanisms of ferroptosis in URSA patients warrant further investigation. Ferroptosis, a novel form of regulated cell death characterized by the accumulation of reactive oxygen species and lipid peroxidation, has shown heightened susceptibility in trophoblast cells.we will validate the occurrence of ferroptosis in the chorionic and decidual tissues of URSA patients and further explore the molecular mechanisms of ferroptosis in URSA through in vitro, this research seeks to provide important theoretical insights into the pathological mechanisms and treatment strategies for URSA. Material and methods Transmission electron microscopy was used to observe the mitochondrial morphology in the chorionic tissues of URSA patients, Western blotting and immunohistochemistry were employed to assess the expression of relevant genes as well as indicators of ferroptosis and lipid peroxidation. An erastin-induced HTR-8/SVneo cell model was established to evaluate the regulatory role of CX43 in oxidative stress and ferroptosis. Results In URSA patients, there is a significant increase in iron deposition in chorionic tissue, accompanied by alterations in mitochondrial morphology, including increased density, reduced cristae, and membrane damage. Protein expression of CX43, FPN1, GPX4, Nrf2, and SCL7A11 is decreased in the chorionic and decidual tissues of URSA patients, while TFRC protein expression is elevated, In the ferroptosis cell model, treatment with Erastin led to a concentration-dependent decrease in the viability of HTR-8/SVneo cells, with 5 μM chosen for subsequent mechanistic studies. Overexpression of CX43 significantly enhanced cell viability in the Erastin treatment group, reducing intracellular ROS and MDA levels while increasing GSH levels. Expression of Nrf2, GPX4, and SCL7A11 was significantly decreased in the Erastin treatment group, whereas overexpression of CX43 restored the expression of these markers. Additionally, CX43 overexpression facilitated the nuclear translocation of Nrf2. Conversely, CX43 interference led to decreased cell viability and reduced expression of antioxidant molecules; however, the Nrf2 agonist TBHQ partially restored the oxidative damage in these cells. Conclusions In the URSA patient group, ferroptosis and changes in the levels of its biomarkers were observed in the chorionic and decidual tissues. Overexpression of CX43 improved oxidative stress and ferroptosis,whereas silencing CX43 exacerbated ferroptosis in the in vitro model. CX43 ameliorates oxidative stress and ferroptosis by modulating the Nrf2/GPX4 pathway.