Binding of microRNA-135a (miR-135a) to homeobox protein A10 (HOXA10) mRNA in a high-progesterone environment modulates the embryonic implantation factors beta3-integrin (ITGβ3) and empty spiracles homeobox-2 (EMX2)

BACKGROUND: Patients with elevated circulating progesterone concentrations on the day of the human chorionic gonadotropin (hCG) trigger had relatively low implantation rates during assisted reproductive treatments. In this study, we assess the hypothesis that different concentrations of progesterone regulate the expression of homeobox protein A10 (HOXA10) and its downstream genes through miRNA-135a. METHODS: MicroRNA-135a (miR-135a), HOXA10, beta3-integrin (ITGβ3), and empty spiracles homeobox-2 (EMX2) expression levels in endometrial tissues from patients with elevated progesterone were measured. To determine the threshold of progesterone level which can impair implantation, Ishikawa cells were used to determine the expression of the aforementioned 4 genes after exposure to 5 graded concentrations of progesterone. The dual-luciferase reporter assay was used to verify whether miR-135a regulated the expression of HOXA10. Furthermore, the effects of HOXA10 on the expression of key endometrial receptivity genes ITGβ3 and EMX2 were confirmed. RESULTS: High progesterone levels promoted miR-135a expression in vivo, and miR-135a bound to the 3'-untranslated region (3'-UTR) of HOXA10 mRNA to inhibit HOXA10 expression. Reduction of HOXA10 promoted EMX2 expression and inhibited ITG-3 production. Progesterone promoted the expression of HOXA10 in vitro at low concentrations. However, when the concentration was greater than 10(−7) ng/mL, progesterone inhibited HOXA10 by promoting miR-135a expression, thereby altering the expression of related genes and affecting endometrial receptivity. CONCLUSIONS: In vitro, the trend in miR-135a expression (which first decreased and then increased) was in direct contrast to that of HOXA10 expression (which first increased and then decreased) as progesterone levels increased. The key factors regulating endometrial receptivity included ITGβ3 and EMX2, which were confirmed to be regulated by HOXA10. High progesterone levels affected miR-135a expression, and miR-135a inhibited HOXA10 expression, thereby affecting endometrial receptivity.