Expression and significance of miR-30d-5p and SOCS1 in patients with recurrent implantation failure during implantation window

Abstract

Background: Poor endometrial receptivity is a major factor that leads to recurrent implantation failure. However, the traditional method cannot accurately evaluate endometrial receptivity. Various studies have indicated that microRNAs (miRNAs) are involved in multiple processes of embryo implantation, but the role of miRNAs in endometrial receptivity in patients with recurrent implantation failure (RIF) remains elusive. In the present study, we investigated the presence of pinopodes and the roles of miR-30d-5p, suppressor of cytokine signalling 1 (SOCS1) and the leukaemia inhibitory factor (LIF) pathway in women with a history of RIF during the implantation window.

Methods: Endometrial tissue samples were collected between January 2018 to June 2019 from two groups of women who underwent in vitro fertilisation and embryo transfer (IVF-ET) or frozen ET. The RIF group included 20 women who underwent ≥ 3 ETs, including a total of ≥ 4 good-quality embryos, without pregnancy, whereas the control group included 10 women who had given birth at least once in the past year. An endometrial biopsy was performed during the implantation window (LH + 7). The development of pinopodes in the endometrial biopsy samples from all groups was evaluated using scanning electron microscopy (SEM). Quantitative reverse transcription-polymerase chain reaction and western blotting were used to investigate the expression levels of miR-30d-5p, SOCS1, and the LIF pathway.

Results: The presence of developed pinopodes decreased in patients with RIF on LH + 7. The expression level of miR-30d-5p decreased in the endometria during the implantation window of patients with RIF, whereas the mRNA and protein levels of SOCS1 were significantly higher in the RIF group than in the control group. Furthermore, a negative correlation was observed between the expression of miR-30d-5p and SOCS1 (r² = 0.8362). In addition, a significant decrease in LIF and p-STAT3 expression was observed during the implantation window in patients with RIF.

Conclusions: MiR-30d-5p and SOCS1 may be potential biomarkers for endometrial receptivity. Changes in pinopode development and abnormal expression of miR-30d-5p, SOCS1 and LIF pathway in the endometrium could be the reasons for implantation failure.

Keywords: Embryo transfer; Endometrial receptivity; In vitro fertilization; MiR-30d-5p; Recurrent implantation failure; SOCS1.

Fig. 1

Scanning electron micrographs (SEM) of uterine epithelial tissues from patients on LH + 7. A Absence of pinopodes. Secretory cells are covered by microvilli (white arrow). B Developing pinopodes. the small semi-spherical projection in the apical membrane is covered by short and sparse microvilli (white arrow). C Fully developed pinopodes. Microvilli have disappeared from the projections which appear to be fully maximally distended (white arrow). D Regressing pinopodes. The projections have a wrinkled surface (white arrow); small microvilli tips reappeared on the membranes. E Comparison of the proportion of the various coverage of pinopodes between the two groups. F SEM of pinopodes were graded semi-quantitatively. Control: receptive control (n = 10); RIF: recurrent implantation failure (n = 20)

Fig. 2

Schematic of the 3′-UTR of SOCS1 with the predicted binding site for has-miR-30d-5p

Fig. 3

Comparison of miR-30d-5p and SOCS1 expression in endometrium samples from the RIF and control groups. A, B The relative expression levels of miR-30d-5p and SOCS1 in the RIF group (n = 20) and the control group (n = 10), as detected via RT-qPCR; U6 was used as an internal control for miR-30d-5p; GAPDH was used as an internal control for SOCS1. C The relative expression level of SOCS1 protein in the RIF group (n = 20) and the control group (n = 10), as detected via western blotting; GAPDH was used as internal control. D Spearman correlation suggests a significant negative correlation between the expressions of miR-30d-5p and SOCS1. The Spearman correlation coefficient, P values, and sample numbers are indicated on the upper right of the plot. *P < 0.05

Fig. 4

Comparison of miR-30d-5p and SOCS1 expression in endometrium samples from the RIF and control groups. A The relative expression level of LIF mRNA in the RIF group (n = 20) and the control group (n = 10), as detected via RT-qPCR; GAPDH was used as an internal control for LIF. B The relative expression levels of LIF, p-STAT3 and STAT3 protein in the RIF group (n = 20) and the control group (n = 10), as detected via western blotting; GAPDH was used as internal control. C, D Data of expression of LIF and p-STAT3 protein were expressed as mean ± SD. *P < 0.05