GnRH antagonist impairs the process of embryo implantation by inhibiting motility of endometrial stromal cells through reducing c-kit expression

Background: It has been recognized that the gonadotropin-releasing hormone antagonist (GnRH-ant) protocol has a detrimental effect on clinical outcomes compared to the GnRH agonist (GnRH-a) protocol during in vitro fertilization-fresh embryo transfer (IVF-ET) cycles. However, the related mechanisms were unclear. Methods: A total of 18,561 patients, who underwent fresh IVF-ET cycles in the Center for Assisted Reproduction of Jiangxi Maternal and Child Health Hospital from January 2014 to September 2021, were retrospectively analyzed. The propensity score matching (PSM) technique was used to control for confounding factors between the GnRH-ant and GnRH-a groups. Human endometrial stromal cells (hESCs) were collected for primary culture and treated with relevant receptor antagonists and activators. RT-PCR, Western Blot, immunofluorescence staining, cell migration and adhesion assays, and animal experiments were employed to elucidate the molecular mechanism by which GnRH antagonist affects the migration and adhesion ability of hESCs. Results: There was no statistical difference between the two groups in terms of baseline characteristics after matching basal status by propensity score matching. The result showed that the endometrial thickness (10.4 +/- 2.35 vs. 11.03 +/- 2.61 mm, p < .001) on trigger day was significantly lower in the GnRH-ant group. Compared with the GnRH-a protocol, the implantation rate (39.71% vs. 50.36%, p < .001), biochemical pregnancy rate (64.26% vs. 72.7%, p < .001), clinical pregnancy rate (56.39% vs. 65.24%, p < .001), live birth rate (45.25% vs. 56.1%, p < .001) in the GnRH-ant group were significantly decreased. Contrarily, the rate of early miscarriage in the GnRH-ant group (13.95% vs. 9.04%, p < .001) was higher than in the GnRH-a group. Furthermore, after treating with GnRH-ant, hESCs showed a reduced expression of HOXA10 and MMP-9 proteins, and a weakened migration ability. Subsequently, by establishing the co-culture system of hESCs and JAR trophoblast spheroids, we found that GnRH-ant inhibited the adhesion and invasion ability of trophoblast cells. Moreover, we also found a decreased expression and phosphorylation of c-kit receptor in decidualized hESCs after treating with GnRH-ant. Similar results as observed above were also confirmed when inhibiting the activation of c-kit receptor by imatinib. Conclusions; GnRH-ant could reduce the motility of hESCs by inhibiting the expression and activation of the C-kit receptor, which impaired the process of embryo implantation.