SENA YILDIZ1, JACOBIE STEENBERGEN1, ANKE MCLUSKEY-DANKBAR1, HANA KUBO2,3, MONICA M. LARONDA2-4, JENNY A. VISSER1
1 Department of Internal Medicine, Erasmus MC, University Medical Center Rotterdam, Rotterdam, the Netherlands
2 Stanley Manne Children’s Research Institute, Ann & Robert H. Lurie Children’s Hospital of Chicago, Chicago, IL, USA
3 Department of Pediatrics, Feinberg School of Medicine, Northwestern University, Chicago, IL, USA
4 Department of Obstetrics and Gynecology, Feinberg School of Medicine, Northwestern University, Chicago, IL, USA
Introduction
Polycystic ovary syndrome (PCOS) is a common reproductive and metabolic disorder in women. In PCOS, Antil-Müllerian hormone (AMH) expression is increased and also its window of expression is prolonged. Yet, the mechanisms regulating AMH expression and the downstream consequences of this altered AMH expression remain unknown. Therefore, we aim to study the transcriptional regulation of AMH as well as downstream effects of AMH signaling in human induced pluripotent stem cells (iPSCs)-derived granulosa-like cells.
Methods
To differentiate iPSC into granulosa-like cells, several ligand-induced differentiation protocols were analyzed in combination with two types of base medium. Assessment for correct lineage commitment and GC markers was performed by gene expression analysis.
Results
Our results indicate that all protocol combinations suppress expression of pluripotency markers. However, protocol-dependent differences were observed in expression of early and late stage GC markers. Expression of the late-follicular stage marker CYP19A1 increased ~100 fold, whereas AMH expression increased 3-5 fold but remained low. Strikingly, a 200-300 fold increase in AMHR2 expression was observed when iPSCs were differentiated in RPMI medium. Combining different differentiation strategies further enhanced GC marker expression (AMHR2, FSHR), however AMH expression remained low. Preliminary results suggest that AMH stimulation initiates downstream signaling.
Conclusion
Our findings demonstrate that iPSCs can be differentiated into granulosa-like cells of distinct follicular stages depending on the differentiation protocol used. Studies are ongoing to further enhance AMH expression to unravel its transcriptional regulation and to assess AMHR2 responsiveness in iPSC-derived granulosa cells obtained from normo-ovulatory women and patients with PCOS.