Homocysteine induces endometrial ferroptosis via MAPK pathway in recurrent pregnancy loss

Recurrent pregnancy loss (RPL) with complex etiology and elevated homocysteinemia (HCY) has been recognized one of the risk factors, however the mechanism of HCY participation in RPL are not fully elucidated.

Samples from RPL_HHCY, RPL_NHCY and controls were used to metabolomics and proteomic analysis. Cell counting kit-8 assay, EdU assay kit, wound healing assay and induced decidualization were performed to observe the HCY induced dysfunction of human endometrial stromal cells (hESCs). Intracellular ROS, lipid peroxidation, MDA, GSH and Fe²⁺ were examined. Western blotting was used to measure protein expression.

We found differential metabolites were enriched in glutathione metabolism, and differentially protein expression were enriched in the ferroptosis. In vitro, ferrostatin-1 (Fer-1) could improve the decrease of HCY induced cell viability, proliferation, migration and decidualization of hESCs, and reverse ROS, lipid peroxidation, MDA, GSH and Fe²⁺ levels. Also, Fer-1 enhanced GPX4 and SLC3A2, lightened ACSL4 protein expression. Gene Set Variation Analysis (GSVA) found MAPK is an important pathway for ferroptosis, and inhibition MAPK signaling pathway reversed the phosphor-ERK (p-ERK), p-JNK and p-P38 amplified by HCY.

Our findings implicate that HCY disturbs the function of hESCs by activation of the MAPK signaling pathway induced ferroptosis and may contribute to RPL. This provides a theoretical basis for the relationship between high HCY and RPL.