Spatio-temporal distribution of RSPO1/DKK1 machinery in testis embryonic development and its role in testicular angiogenesis

Testicular vasculogenesis is one of the key processes regulating male gonad morphogenesis. The knowledge of the molecular cues underlining this phenomenon could represent a central element in the understanding of the onset of testicular morphogenetic disorders. RSpondin-1 (RSPO1) has been clearly established as a candidate for ovary determination both in mice and humans. On the contrary, very few data are available on the expression and role of RSPO1 during testicular development. Tis study aims to clarify the distribution pattern of RSPO1 as well as of other partners of its machinery during the entire period of testicular morphogenesis and to indicate the role of this complex system in testicular development. To this end we frst studied the distribution pattern of RSPO1, its antagonist Dickkopf-1 (DKK1), and the signaling efector β-catenin during testis development, from 11.5 to 18.5 dies post coitum (dpc). Our results clearly demonstrate that RSPO1 and DKK1 distribution pattern partially overlaps but, intriguingly, RSPO1 is always detectable in the testicular coelomic portion, the region that organizes testicular vasculature, whilst DKK1 is never detectable in this area. Moreover, performing organ culture experiments of embryonic testes, we demonstrated that DKK1 acted as an inhibitor of testis recruitment of endothelial mesonephric cells. Interestingly, this inhibition was rescued by the co-administration of RSPO1. In addition, RSPO1 alone has been able to enhance, in culture, testicular vasculogenesis. Tese results let us to conclude that RSPO1 machinery is involved in the control of the correct testicular vascular compartment organization.