Integrating network pharmacology and metabolomics to elucidate the mechanism of action of Yangluan formula for treating of diminished ovarian reserve.

Objectives: The Yangluan Formula (YLF) influences the outcomes of in vitro fertilization-embryo transfer (IVF-ET) in infertile women with diminished ovarian reserve (DOR); however, the potential mechanisms by which YLF ameliorates DOR have not yet been elucidated. The aim of this study was to examine the effects of YLF on IVF-ET outcomes in infertile women with DOR, and to elucidate the potential mechanisms by which YLF addresses DOR.

Methods: Non-targeted metabolomics studies were conducted on follicular fluid specimens procured from individuals with DOR treated with or without YLF, and from patients with normal ovarian reserve who underwent IVF-ET treatment. Distinct metabolites were identified using untargeted metabolomics, and MetaboAnalyst was used to examine metabolic pathways. After applying network pharmacology (NP), the target of YLF acting on DOR was determined. Cytoscape software was used to develop compound-reaction-enzyme-gene networks, and molecular docking (MD) simulations were conducted to confirm the link between YLF and crucial targets.

Results: Patients with DOR showed a notable reduction in the number of oocytes retrieved, incidence of 2PN fertilization, and number of cleaved embryos (P < 0.001). Additionally, the DOR cohort exhibited a markedly reduced quantity of high-quality embryos on day 3 compared to the CON cohort (P < 0.005). In contrast to the DOR cohort, the YLF cohort exhibited notably superior outcomes in terms of 2PN fertilization rates, cleavage-stage embryo development, and the number of high-grade embryos on day 3 (P < 0.05). The proportion of 2PN fertilization observed in YLF subjects substantially exceeded that in DOR individuals (81.2 % vs. 64.3 %, P < 0.05). Combined analysis of metabolomics and NP, focusing on five key targets for the action of YLF (monoamine oxidase A, monoamine oxidase B, myeloperoxidase, xanthine dehydrogenase, and phosphodiesterase 3A), four key metabolites (pelargonic acid, 1-(5-Phospho-D-ribosyl)-5-amino-4-imidazolecarboxylate, isokobusone, 5-O-(1-Carboxyvinyl)-3-phosphoshikimate), and two related pathways (glycine, serine, alanine, and threonine metabolism).

Conclusion: We elucidated the mode of action of YLF in DOR treatment by integrating metabolomics and NP. YLF can effectively improve IVF outcomes in patients with DOR. This study provides new perspectives on the mechanism by which YLF improves ovarian function.