Differential Gene Expression of Mesenchymal Stromal Cell and Granulosa Cell Markers in Equine Follicular Aspirates

Equine follicular aspirates obtained during ovum pick-up contain a heterogeneous mixture of mesenchymal stromal cells (MSC) and granulosa cells (GC). This study evaluates how selective isolation and expansion of MSC decrease GC populations. Follicular aspirates-derived cells were cultured up to passage four (P4) following a previously validated protocol. While freshly isolated follicular aspirates cells (FAC) displayed heterogeneous morphologies, early passages (P3-P4) showed a uniform, spindle-shaped phenotype compatible with MSC. Gene expression analysis by RT-qPCR revealed a marked upregulation of mesenchymal and stemness-associated genes (FN1, THY1, FGF2 and CD105) in P4-MSC compared with FAC, confirming enrichment of the MSC population. Conversely, GC-specific genes (FSHR, FOXL2 and CYP11A1) were significantly upregulated in FAC. These findings suggest that early passages efficiently promote selective MSC expansion while progressively eliminating GC.