Sex is determined by the onset of SRY expression around week 6 in the human XY embryo. Differences of Sex Development (DSD) are a spectrum of conditions in which gonadal, chromosomal or anatomical sex is atypical 1. SRY mutations account for 15% of 46,XY DSDs, and rare mutations in about 30 other genes are involved, but the underlying molecular basis in many forms of DSD remains unknown, due in part to the lack of manipulatable models that recapitulate human sex determination.
Mouse models using knockout/transgenic approaches have been useful in studying the function of critical sex determination genes. Differences in gene expression thresholds and genetic robustness between humans and mice are becoming apparent2. Limitations include functional redundancy, gene dosage or genetic buffering, such that mutations causing DSD in humans produces no phenotype consequence in mice3.
An in vitro model that can be used to model Sertoli cell function in the testis is NT2/D1, a human pluripotent clonal cell line derived from a testicular tumour4. NT2/D1 cells can be used to model a variety of human developmental processes: undifferentiated they can model human Sertoli cell development, and when differentiated they can model neuronal development5 or smooth muscle development6.
We have established an NT2/D1-cas9 cell line, and characterised these via a suite of cell phenotyping assays including xCELLigence® RTCA and germ cell maturation assays. This model can be used in conjunction with siRNA knockdown investigations of genes of interest, to assess their individual contributions to characteristics such as cell adhesion, proliferation and capacity to nurture germ cells.