Characterization of Ovarian Small Round Cells Reveals Distinct Molecular and Cellular Features Under Spermatogonial Stem Cell‐Supportive Conditions

The mammalian ovary contains diverse cell populations with distinct molecular and functional characteristics. Rare ovarian-derived cell populations remain incompletely characterized. Spermatogonial stem cells (SSCs) provide a well-established model for studying cellular features associated with pluripotency, niche interaction, and germline-associated programs. Here, we investigated whether ovarian-derived cells exhibit selected characteristics associated with these features under defined culture conditions. Using mouse ovarian tissue, we isolated and characterized ovarian-derived small round cells (OSRCs) following culture under SSC-supportive conditions. Using Oct4-GFP transgenic mice, we identified two ovarian-derived populations: ovarian epithelial-like cells (OELCs) and OSRCs. These populations were compared using morphology, alkaline phosphatase activity, marker expression analysis, molecular comparison with SSC populations, and transplantation into an SSC-supportive testicular environment. OELCs showed characteristics of ovarian somatic cells, whereas OSRCs exhibited increased expression of selected pluripotency-associated markers and variable expression of germline-associated markers. OSRCs differed from undifferentiated SSCs, showing reduced or heterogeneous expression of markers such as Ddx4 and Dazl. Following transplantation, OSRCs localized within seminiferous tubules but showed no evidence of germline differentiation. OSRCs represent a distinct ovarian-derived cell population with partial overlap with pluripotency-associated and germline-associated molecular features. These findings provide a basis for further investigation of ovarian cellular heterogeneity.