ESPE Abstracts

Human sex development relies on differentiation of the gonads, in which Sertoli and granulosa cells play a key role for men and women respectively. Many variations of sex development (VSD) are due to alteration of these two cell types. The study of the mechanisms underlying these conditions is crucial for optimal clinical management of VSDs, as for more than 50% [1] of all VSD patients, the molecular cause of their condition remains unknown. The collection of primary cells is invasive and cumbersome for the patients and their in vitro culture extremely difficult due to short lifespan and the loss of the unique characteristics. Additionally, the available cell models do not reproduce the patient-specific mechanisms of the disease. An attractive alternative are patients-derived cells which would allow a “personalized“ clarification of the pathophysiology of disease. Eventually, the analysis of several patients would identify common pathways and shed more light on the origin of these disorders. Human induced-pluripotent stem cells (iPSCs) are developing as exciting cell sources for applications in regenerative medicine and drug discovery, primarily based on their extensive similarities to their human embryonic stem cell counterparts and shared properties of self-renewal and multilineage differentiation capabilities. iPSCs can be derived from somatic cells like fibroblasts, peripheral blood mononuclear cells (PBMCs) or urinary progenitors cells (UPCs) via ectopic expression of transcription factors SOX2, OCT4, KLF4, NANOG, C-MYC and LIN28 [2]. UPCs are a powerful source with great potential for reprogramming into iPSCs, as the collection is simple, inexpensive, and non-invasive. In addition, UPCs have properties such as clonogenicity, expansion and self-renewal capacity [3, 4]. In this study the focus lies on the generation and characterization of the UPCs and iPSCs, as it is crucial to know the sources before differentiating into Sertoli or granulosa cells. The reprograming from UPCs to iPSCs was performed with either nucleofection or magnetofection, two viral vector-free methods. The characterization was done qualitative with antibody staining and quantitative with qPCR. For the analysis the following markers were chosen: CD73, CD44, CD146, OCT4, NANOG, SSEA-1, SOX2, and TRA 1-81. UPCs expressed CD44, CD73, CD146 and stem cell markers such as OCT4 and NANOG, but not SOX2 and TRA-1-81. The iPSCs expressed all stem cell markers examined here. UPCs-derived iPSCs may become an alternative source for generating human cell models ideally directly from the patients and could be of great importance for basic research in sex development and reproductive medicine.