Abstract
In the human fetal testis, fetal germ cells (FGCs) are progressively surrounded by supporting Sertoli cells inside seminiferous cords. During the second trimester, the FGCs develop asynchronously and can be observed in several stages of development. However, the mechanism that regulates the transition between the different developmental stages as well as the formation of spermatogonia is currently not well understood. For this, it is necessary to develop suitable isolation protocols and a platform for in vitro culture of FGCs of different stages. Here, we report a method to isolate distinct populations of FGCs and Sertoli cells from second trimester human testis using a panel of conjugated antibodies for THY1, PDPN, ALPL, KIT, and SUSD2 for fluorescence-activated cell sorting (FACS) followed by in vitro culture up to 7 days. This platform provides the base for cellular and molecular characterization of the different testicular cell populations to investigate the transition between FGCs and spermatogonia and shed some light on crucial processes of early human gametogenesis unknown until now.
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Acknowledgments
We would like to thank the generous donors and the helpful staff of the Vrelingshuis in Utrecht for the human fetal material used in this study. We also thank S. Hillenius, T. van der Helm, and S. Czukiewska for processing human fetal material for cryopreservation and the LUMC Flowcytometry Core Facility and the LUMC Microscopy Core Facility for technical support. This work was supported by the Dutch Research Council (VICI-2018-91819642 to AWO, YWC, CMR, and SMCSL), the Dutch organization ZonMw (ZonMw PSIDER 10250022120001 to SMCSL), and the Novo Nordisk Foundationgrant (renew NNF21CC0073729 to AWO, MB, and SMCSL).
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Roelse, C.M., Overeem, A.W., Chang, Y.W., Boubakri, M., Chuva de Sousa Lopes, S.M. (2024). Isolation and In Vitro Culture of Germ Cells and Sertoli Cells from Human Fetal Testis. In: Barchi, M., De Felici, M. (eds) Germ Cell Development. Methods in Molecular Biology, vol 2770. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3698-5_6
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DOI: https://doi.org/10.1007/978-1-0716-3698-5_6
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