A Novel in vitro Co-culture Systems on Differentiation of Embryonic Stem Cells into Oocyte-like Cells in an in vivo Manner

Differentiation of Embryonic Stem Cells into Oocyte-like cells in vitro is challenging. Successful derivation of oocyte from stem cells can provide an alternative source for curing ovogenesis problems. The current study aims to demonstrate a new protocol with two different types of media for differentiating embryonic stem cells (ESCs) into oocyte-like cells (OLCs). After culturing mouse ESCs, embryoid bodies (EBs) were generated from ESCs by hanging drop (HD) method. To final differentiation of oocyte-like cells (OLCs), the EBs were cultured in two different types of media for 12 days (first 7 days EBs were cultured in in vitro maturation diluted in Granulose Cell- Conditioned Medium and Follicular Fluid [1:1:1] followed by 5 days of culture in in vitro maturation diluted in uterine condition medium [1:1] ). According to the MTT test, the viability rate increased in the experimental group compared to the control EBs cultured alone. Expression of Oct4, as a pluripotency marker, decreased during the differentiation process of EBs in the experimental group. Co-culturing of EBs with our mentioned protocol increased germ cell markers (Stella and Mvh) and increased Oocyte-specific markers (ZP1, Figα and GDF9). Our study introduces a promising in vitro protocol for achieving successful oogenesis through creating interactions of EBs with granulosa cells and uterine condition medium.