Yeli Sun ^1,2,3,4 Guohua Li ^1,3,4,5 Yuan Tan ^1,6* Mengwen Kong ^1,2,3,4* Junyuan Li ^1,2,3,4* Shuyun Wang ^1,6*

• ¹ School of Medicine, Tongji University, Shanghai, China
• ² Shanghai First Maternity and Infant Hospital, Shanghai, China
• ³ Shanghai Institute of Maternal-Fetal Medicine and Gynecologic Oncology, Shanghai, China
• ⁴ Shanghai key laboratory of Maternal Fetal Medicine, Shanghai, China
• ⁵ Department of Reproductive Immunology, Shanghai First Maternity and Infant Hospital, Shanghai, China
• ⁶ Department of Integrated Traditional Chinese Medicine (TCM) & Western Medicine, Shanghai First Maternity and Infant Hospital, Shanghai, China

Recurrent spontaneous abortion (RSA) represents a significant clinical challenge, with its underlying mechanisms yet to be fully elucidated. Despite advances in understanding, the precise pathophysiology driving RSA remains unclear. Angelica sinensis, a traditional herbal remedy, is frequently used as an adjunctive treatment for miscarriage. However, it remains uncertain whether its primary active component, Angelica sinensis polysaccharide (ASP), plays a definitive role in its therapeutic effects. The specific function and mechanism of ASP in the context of RSA require further investigation. In this study, we sought to evaluate autophagy levels at the maternal-fetal interface in RSA patients and in an RSA mouse model treated with ASP, complemented by a comprehensive metabolomic analysis. Autophagy flux in the decidua was compared between eight RSA patients and eight healthy pregnant women.Additionally, changes in autophagy flux were assessed in an RSA mouse model following ASP treatment, with embryos and placental tissues collected for subsequent metabolomic profiling. Our results revealed a significant reduction in Beclin 1 protein levels in the decidua of RSA patients compared to the normal pregnancy group. Conversely, ASP treatment in the RSA mouse model restored autophagy-related protein expression, including ATG7, ATG16L, and Beclin 1, to levels higher than those observed in the untreated RSA group. Metabolomic analyses further identified significant changes in phosphatidylethanolamine levels between ASP-treated and control groups, with differential metabolites enriched in pathways related to glycolysis/gluconeogenesis, glycerolipid metabolism, and glycine, serine, and threonine metabolism. Functional assays revealed that ASP enhances trophoblast cell proliferation, migration, and invasion. In summary, our findings demonstrate diminished autophagy activity in RSA patients, while ASP appears to restore autophagy and regulate key metabolic pathways, including glycolysis/gluconeogenesis. These results provide new insights into the protective mechanisms of ASP in RSA, suggesting its potential as a therapeutic intervention for this condition.