Modeling Klinefelter Syndrome Using Induced Pluripotent Stem Cells Reveals Impaired Germ Cell Differentiation

Olivier Botman; Youssef Hibaoui; Maria G Giudice; Jérôme Ambroise; Catherine Creppe; Anis Feki; Christine Wyns

doi:10.3389/fcell.2020.567454

Modeling Klinefelter Syndrome Using Induced Pluripotent Stem Cells Reveals Impaired Germ Cell Differentiation

Front Cell Dev Biol. 2020 Oct 7:8:567454. doi: 10.3389/fcell.2020.567454. eCollection 2020.

Authors

Olivier Botman^{1

2}, Youssef Hibaoui^{3

4}, Maria G Giudice^{1

2}, Jérôme Ambroise⁵, Catherine Creppe⁶, Anis Feki^{3

4}, Christine Wyns^{1

2}

Affiliations

¹ Gynecology Unit, Institut de Recherche Expérimentale et Clinique (IREC), Université catholique de Louvain, Brussels, Belgium.
² Department of Gynecology-Andrology, Cliniques Universitaires Saint-Luc, Brussels, Belgium.
³ Stem Cell Research Laboratory, Department of Obstetrics and Gynecology, Geneva University Hospitals, Geneva, Switzerland.
⁴ Department of Obstetrics and Gynecology, Hôpital Fribourgeois (HFR) Fribourg, Hôpital Cantonal, Fribourg, Switzerland.
⁵ Center for Applied Molecular Technologies (CTMA), Institut de Recherche Expérimentale et Clinique (IREC), Université catholique de Louvain, Brussels, Belgium.
⁶ Groupe Interdisciplinaire de Génoprotéomique Appliquée (GIGA)-Signal Transduction, C.H.U. Sart Tilman, University of Liège, Liège, Belgium.

Abstract

Klinefelter syndrome (KS), with an incidence between 1/600 and 1/1,000, is the main genetic cause of male infertility. Due to the lack of an accurate study model, the detailed pathogenic mechanisms by which this X chromosome aneuploidy leads to KS features remain unknown. Here, we report the generation and characterization of induced pluripotent stem cells (iPSCs) derived from a patient with KS: 47XXY-iPSCs. In order to compare the potentials of both 47XXY-iPSCs and 46XY-iPSCs to differentiate into the germ cell lineage, we developed a directed differentiation protocol by testing different combinations of factors including bone morphogenetic protein 4 (BMP4), glial-derived neurotrophic factor (GDNF), retinoic acid (RA) and stem cell factor (SCF) for 42 days. Importantly, we found a reduced ability of 47XXY-iPSCs to differentiate into germ cells when compared to 46XY-iPSCs. In particular, upon germ cell differentiation of 47XXY-iPSCs, we found a reduced proportion of cells positive for BOLL, a protein required for germ cell development and spermatogenesis, as well as a reduced proportion of cells positive for MAGEA4, a spermatogonia marker. This reduced ability to generate germ cells was not associated with a decrease of proliferation of 47XXY-iPSC-derived cells but rather with an increase of cell death upon germ cell differentiation as revealed by an increase of LDH release and of capase-3 expression in 47XXY-iPSC-derived cells. Our study supports the idea that 47XXY-iPSCs provides an excellent in vitro model to unravel the pathophysiology and to design potential treatments for KS patients.

Keywords: Klinefelter syndrome; Klinefelter syndrome iPSCs; germ cell differentiation; induced pluripotent stem cells; post-meiotic cells; primordial germ cells.