Evidence for existence of molecular stemness markers in porcine ovarian follicular granulosa cells

Abstract Granulosa cells (GCs) are important component of the follicle, a principal functional unit of the ovary. They undergo highly dynamic changes during folliculogenesis and play a vital role in oocyte’s maturation. Recently, it has been shown that GCs also exhibit stem cell properties, since they express OCT-4, Nanog, Sox-2, which are markers of pluripotency, as well as several mesenchymal stem cell markers, such as CD29, CD44, CD90, CD105, CD117 or CD166. In addition, GCs are able to differentiate towards neurogenic, chondrogenic and osteogenic lineages. Since the use of embryonic stem cells in regenerative medicine is burdened with ethical concerns and the risk of immune rejection or teratoma formation, adult stem cells are emerging as a promising alternative. GCs especially seem to provide a promising source of stem cells, since they are easily obtainable during assisted reproduction techniques. In order to better understand the genetic changes taking place in proliferating granulosa cells cultured in vitro, we isolated GCs from 40 prepubertal gilts and cultured them in vitro for 168 h. After 24, 48, 72, 96, 120, 144 and 168 h of cultivation the total RNA was extracted, reverse transcription was conducted and RT-qPCR reaction was performed. We observed that CD44, CD90 and IGF1 were upregulated after the cultivation, whereas CD105 and LIF were downregulated. Collectively, our results confirm stemness potential of porcine GCs and provide an insight into the transcriptome changes during in vitro cultivation. Running title: Molecular stemness markers in porcine granulosa cells