CONCLUSION

This study demonstrated that mouse endometrial epithelial cells undergo substantial proteomic remodeling during the transition from Day 1 to Day 4 of pregnancy, corresponding to the establishment of uterine receptivity. Comparative proteomic analysis identified 80 differentially expressed protein spots, of which 52 proteins were successfully characterized. Functional classification revealed coordinated regulation of proteins associated with cellular metabolism, antioxidant defense, ER stress responses, cytoskeletal organization, protein folding, and reproductive system development. Notably, Gstm2 exhibited consistent upregulation at both the protein and transcript levels during the receptive phase, whereas vimentin expression remained relatively stable, suggesting distinct functional roles in epithelial adaptation and structural maintenance during early pregnancy.

The findings provide important insights into the molecular events occurring at the embryo–maternal interface and highlight the potential contribution of redox homeostasis and stress adaptation pathways to successful implantation. The coordinated modulation of antioxidant-related proteins (Gstm2, Gstm7, Prdx2, and Cat) together with ER-associated proteins (PDIA3 and HSPA5) suggests that the maintenance of oxidative balance and protein-folding capacity may be integral components of epithelial preparation for embryo attachment. These observations contribute to the growing body of evidence that uterine receptivity is governed by interconnected molecular networks rather than individual regulatory factors.

A major strength of this study is the use of isolated endometrial epithelial cells, which enabled cell type–specific characterization of proteomic changes that may be obscured in whole-tissue analyses. Furthermore, integration of proteomic profiling with immunofluorescence and quantitative real-time PCR validation increased confidence in the biological relevance of the identified candidates. Nevertheless, several limitations should be considered, including the study's exploratory nature, the use of pooled samples, the limited proteome coverage associated with gel-based proteomics, the absence of embryo verification via uterine flushing, and the lack of functional validation experiments to establish causal relationships between candidate proteins and implantation outcomes.

Future investigations should employ high-resolution quantitative proteomic platforms, improved epithelial cell purification strategies, and functional approaches such as gene knockdown, overexpression, or conditional knockout models to clarify the mechanistic roles of candidate proteins during implantation. Validation in additional mouse strains and human endometrial samples, particularly from women experiencing RIF, will be essential to determine the translational relevance of these findings.

Overall, the present study provides a comprehensive epithelial-specific proteomic profile of the mouse endometrium during early pregnancy and identifies coordinated changes in antioxidant, stress-response, metabolic, and structural pathways associated with uterine receptivity. These findings establish a valuable molecular framework for future studies to elucidate implantation mechanisms and identify biomarkers or therapeutic targets relevant to reproductive success.