Untargeted metabolomics and transcriptomics study reveals an activated ferroptosis metabolic spectrum in recurrent spontaneous abortion.

Recurrent spontaneous abortion (RSA) poses a significant obstetric challenge and its underlying molecular mechanisms remain largely unknown. Ferroptosis, an iron-dependent cell death, is characterized by iron overload and widespread lipid peroxidation. In this study, we discovered a characteristic metabolic spectrum of ferroptosis in RSA by untargeted UPLC-MS/MS metabolomics analysis. The metabolic profiles of the villous of RSA patients could be effectively differentiated from these of controls in OPLS-DA and random forest analysis, indicating distinct metabolic signatures in RSA villous. These signatures mainly included pathways of Glutamate Metabolism, Methionine, Cysteine, SAM and Taurine Metabolism, Polyamine Metabolism, and Lipid Peroxidation Metabolism. Notably, we noticed that the metabolic alterations in RSA villous showed strong ferroptosis phenotypes, including high levels of lipid peroxidation and lower levels of cysteine, serine, methionine, glutamate, and glutamine, etc. Furthermore, these strong ferroptosis-related metabolic changes in the villous of RSA patients were supported by the transcriptomics study results. RNA-seq results revealed that the expressions of 89 ferroptosis-related genes were altered in the RSA villous. GSEA, GO and KEGG analysis of the RNA-seq data also found significantly enriched ferroptosis-related pathways. Elevated mRNA expression of ACSL4, FTH1, TFRC, PTGS2 and decreased expression of GPX4 and SLC7A11 in the RSA villous were further confirmed by RT-PCR. The findings in this study underscore the pivotal role that ferroptosis plays in RSA and offers perspectives of developing new strategies for RSA diagnosis and therapy.