Hsa_circ_0000417 downregulation promotes hypospadiac formation by modulating AR-PTEN-AKT axis

Background: Congenital hypospadias is a common congenital malformation of the urinary system in male children. To explore the mechanism of hypospadias is particularly important,because it could provide a potential guide for clinical treatment. Dysregulated apoptosis of penile mesenchymal cells has been previously demonstrated to underly hypospadiac urethral closure failure.we found hsa_circ_0000417 significantly downregulated in hypospadias preputial specimens,act as a ceRNA for AR via sequestering hsa_miR-6756-5p, and that the biological functions of hsa_circ_0000417 may significantly involve the PI3K/AKT pathway in previous reports.However, the regulatory mechanisms remain poorly unknown. Methods: : We performed study on primary culture fibroblasts from normal and hypospadias tissues in vitro.The cell apoptosis was measured by fow cytometry using Apoptosis Detection Kit,and the protein level of AR,bax, caspase 9 and AKT signaling pathway were measured by Western Blot (WB). we examined the role of hsa_circ_0000417 in vivo using the mouse model of hypospadias. Results: : The effects of hsa_circ_0000417 in foreskin fibroblasts were studied both in vitro and vivo. Functionally, knockdown of hsa_circ_0000417 influence the mesenchymal cell apoptosis whereas overexpression had the reverse effects. Mechanistically, primary culture fibroblasts and mice models were detected by flow cytometry and Western Blot (WB)，androgen receptor (AR)、cell apoptosis proteins of AKT、BAX、Caspase9 were significantly decreased expression compared with normal tissues, attenuated mesenchymal cell apoptosis. Further investigation revealed that hsa_circ_0000417 directly interacted with miR-6756. miR-6756 inhibitors had a rescue effect on the proliferation and apoptosis capacities by knockdown of hsa_circ_0000417.It suggested that hsa_circ_0000417 may regulate AR-PTEN-AKT pathway influence the apoptosis of fibroblasts. Conclusions: : Taken together,hsa_circ_0000417 functioned as a molecular sponge for miR-6756 promotes the occurrence of hypospadias through the AR-PTEN-AKT axis and may serve as a novel diagnostic marker and target for treatment of hypospadiac patients.