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

Abstract

<strong>Background:</strong>Differentiation of Embryonic Stem Cells into Oocyte-like cells <em>in vitro</em> 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).<br /> <strong>Methods:</strong> 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 <em>in vitro</em> 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] ).<br /> <strong>Results:</strong>According tothe MTT test, the viability rate increased in the experimental group compared to the control EBs cultured alone. Expression of <em>Oct4</em>, 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 (<em>Stella</em> and <em>Mvh</em>) and increased Oocyte-specific markers (ZP1,<em> Fig</em>α and <em>GDF9</em>).<br /> <strong>Conclusion:</strong>Our study introduces a promising <em>in vitro</em> protocol for achieving successful oogenesis through creating interactions of EBs with granulosa cells and uterine condition medium.