Inhibition of WNT/β-catenin signalling during sex-specific gonadal differentiation is essential for normal human fetal testis development

Abstract Sex-specific gonadal differentiation is directed by complex signalling promoting development in either male or female direction, while simultaneously inhibiting the opposite pathway. In mice, the WNT/β-catenin pathway promotes ovarian development and the importance of actively inhibiting this pathway to ensure normal testis development has been recognised. However, the implications of alterations in the tightly regulated WNT/β-catenin signalling during human fetal gonad development has not yet been examined in detail. Thus, the aim of this study was to examine the consequences of dysregulating the WNT/β-catenin signalling pathway in the supporting cell lineage during sex-specific human fetal gonad development using an established and extensively validated ex vivo culture model. Inhibition of WNT/β-catenin signalling in human fetal ovary cultures resulted in only minor effects, including reduced secretion of RSPO1 and reduced cell proliferation although this was not consistently found in all treatment groups. In contrast, promotion of WNT/β-catenin signalling in testes severely affected development and function. This included disrupted seminiferous cord structures, reduced cell proliferation, reduced expression of SOX9/AMH, reduced secretion of Inhibin B and AMH as well as loss of the germ cell population. Additionally, Leydig cell function was markedly impaired with reduced secretion of testosterone, androstenedione and INSL3. Together, this study suggests that dysregulated WNT/β-catenin signalling during human fetal gonad development severely impairs testicular development and function. Importantly, our study highlights the notion that sufficient inhibition of the opposite pathway during sex-specific gonadal differentiation is essential to ensure normal development and function also applies to human fetal gonads.