[Construction of competitive endogenous RNA regulatory network and clinical verification for repeated implantation...

Objective: To construct a repetitive implantation failure (RIF)-related competitive endogenous RNA (ceRNA) regulatory network and validate with clinical samples. Methods: RIF-related long non-coding RNA (lncRNA), microRNA (miRNA) and messenger RNA (mRNA) from the high-throughput gene expression omnibus (GEO) database Expression profile data set were obtained to construct a ceRNA regulatory network of lncRNA-miRNA-mRNA. At the same time, weighted gene co-expression network analysis (WGCNA) was used to explore hub genes in the network. This retrospective study collected RIF patients and controls (at least one pregnancy history after assisted conception) who underwent in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) for assisted pregnancy from 2020 to 2021 at the Reproductive Medicine Center of the First Affiliated Hospital of Zhengzhou University. In the endometrial tissue of patients with 1 pregnancy history, real-time fluorescence quantitative polymerase chain reaction (qRT-PCR) was used to verify the mRNA expression levels of RIF-related hub genes, and Western blotting and immunohistochemistry were used to verify protein expression levels of vascular cell adhesion molecule-1 (VCAM1). Results: A RIF-related ceRNA regulatory network consisting of 32 lncRNAs, 31 miRNAs and 88 mRNAs was constructed, and 7 RIF-related hub genes were identified using WGCNA. By intersecting 88 mRNAs and hub genes in the ceRNA network, two RIF-related key genes were obtained, i.e., VCAM1 and interleukin-2 receptor α (interleukin-2 receptor α, IL-2RA). In clinical verification, the ages of the control group and RIF group [M (Q₁, Q₃)] were 26.50 (25.00, 34.00) and 30.50 (25.75, 35.25) years old, respectively (P>0.05). Compared with the control group, the mRNA [0.30 (0.15, 0.42) vs 0.99 (0.69, 1.34), P=0.001] and protein expression [0.44 (0.16, 1.27) vs 2.39 (1.58, 2.58), P<0.001] of VCAM1 in the endometrium of the RIF group were both reduced. Conclusions: This study uses bioinformatics analysis methods to construct a RIF-related ceRNA regulatory network, and it is confirmed through clinical samples that the expression level of VCAM1 in the endometrial tissue of RIF patients is significantly reduced.