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Differentiating mouse ESC towards gonadal progenitors cells

ABSTRACT: We developed a gradual differentiation protocol to differentiate mESC towards the EpiSC lineage, followed by early mesoderm cells. These were later differentiated to intermediate mesoderm and we found a condition (IM3) that best generates early gonadal progenitor cells which are normally present at the bipotential gonad at E11.5. Forced expression of 2 transcription factors: Nr5A1 (SF1) and Dmrt1 induced a specific-Sertoli marker (TESCO-CFP) and the cells started to self-aggregate and form tubule-like structures, highly resembling the testis cords. There was proper induction of relevant markers at the correct time-points as shown by both qPCR, immuno-staining and RNA-Seq that includes comparison to in-vivo gonads.

ORGANISM(S): Mus musculus

PROVIDER: GSE165133 | GEO | 2021/10/15

REPOSITORIES: GEO

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Project description:The nuclear receptor subfamily 5 group A member 1 (NR5A1), encoding steroidogenic factor 1 (SF-1), has been identified as a critical factor in gonadal development in animal studies. A previous study of ours suggested that upregulation of NR5A1 during early gonadal differentiation in male (46,XY) human pluripotent stem cells steers the cells into a more mature gonadal cell type. However, the detailed role of NR5A1 in female gonadal differentiation has yet to be determined. In this study, by combining the processes of gonadal differentiation and conditional gene activation, we show that NR5A1 induction predominantly upregulates the female gonadal marker inhibin subunit α (INHA) and steroidogenic markers steroidogenic acute regulatory protein (STAR), cytochrome P450 family 11 subfamily A member 1 (CYP11A1), cytochrome P450 family 17 subfamily A member 1 (CYP17A1), hydroxy-delta-5-steroid dehydrogenase (HSD3B2) and hydroxysteroid 17-beta dehydrogenase 1 (HSD17B1). In contrast, NR5A1 induction did not seem to affect the bipotential gonadal markers gata binding protein 4 (GATA4) and Wilms tumour suppressor 1 (WT1) nor the female gonadal markers r-spondin 1 (RSPO1) and wnt family member 4 (WNT4). Differentially expressed genes were highly associated with adrenal and ovarian steroidogenesis pathways. Moreover, time-series analysis revealed different dynamic changes between male and female -induced samples, where continuously upregulated genes in female gonadal differentiation were mostly associated with adrenal steroidogenesis. Thus, in contrast to male gonadal differentiation, NR5A1 is necessary but not sufficient to steer human embryonic stem cell (hESC)-derived bipotential gonadal-like cells towards a more mature somatic, female cell fate. Instead, it seems to direct bipotential gonadal-like cells more towards a steroidogenic-like cell population. The information obtained in this study helps in elucidating the role of NR5A1 in gonadal differentiation of a female stem cell line.

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Differentiating mouse ESC towards gonadal progenitors cells

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