Background:About 10% of individuals undergoing in vitro fertilization encounter recurrent implantation failure (RIF), which represents a worldwide social and economic concern. Nevertheless, the critical genes and genetic mechanisms underlying RIF are largely unknown.Methods:We first obtained three comprehensive microarray datasets “GSE58144, GSE103465 and GSE111974”. The differentially expressed genes (DEGs) evaluation, enrichment analysis, as well as efficient weighted gene co-expression network analysis (WGCNA), were employed for distinguishing RIF-linked hub genes, which were tested by RT-qPCR in our 30 independent samples. Next, we studied the topography of infiltration of 22 immune cell subpopulations and the association between hub genes and immune cells in RIF using the CIBERSORT algorithm. Finally, a novel ridge plot was utilized to exhibit the potential function of core genes.Results:The enrichment of GO/KEGG pathways reveals that Herpes simplex virus 1 infection and Salmonella infection may have an important role in RIF. After WGCNA, the intersected genes with the previous DEGs were obtained using both variance and association. Notably, the subsequent nine hub genes were finally selected: ACTL6A, BECN1, SNRPD1, POLR1B, GSK3B, PPP2CA, RBBP7, PLK4, and RFC4, based on the PPI network and three different algorithms, whose expression patterns were also verified by RT-qPCR. With in-depth analysis, we speculated that key genes mentioned above might be involved in the RIF through disturbing endometrial microflora homeostasis, impairing autophagy, and inhibiting the proliferation of endometrium. Furthermore, the current study revealed the aberrant immune infiltration patterns and emphasized that uterine NK cells (uNK) and CD4+ T cells were substantially altered in RIF endometrium. Finally, the ridge plot displayed a clear and crucial association between hub genes and other genes and key pathways.Conclusion:We first utilized WGCNA to identify the most potential nine hub genes which might be associated with RIF. Meanwhile, this study offers insights into the landscape of immune infiltration status to reveal the underlying immune pathogenesis of RIF. This may be a direction for the next study of RIF etiology. Further studies would be required to investigate the involved mechanisms.

中文翻译：

---

加权基因共表达网络分析和临床验证确定了复发性着床失败患者子宫内膜细胞中的枢纽基因和免疫细胞浸润

背景：大约 10% 的人正在经历体外受精遇到反复着床失败（RIF），这是一个世界范围的社会和经济问题。然而，RIF背后的关键基因和遗传机制在很大程度上是未知的。方法：我们首先获得了三个综合微阵列数据集“GSE58144、GSE103465和GSE111974”。采用差异表达基因 (DEG) 评估、富集分析以及有效加权基因共表达网络分析 (WGCNA) 来区分 RIF 连接的中心基因，并通过 RT-qPCR 在我们的 30 个独立样本中进行测试。接下来，我们使用 CIBERSORT 算法研究了 RIF 中 22 个免疫细胞亚群的浸润拓扑以及中枢基因和免疫细胞之间的关联。终于有小说了利用岭图展示核心基因的潜在功能。结果：GO/KEGG通路富集揭示单纯疱疹病毒1感染和沙门氏菌感染可能在 RIF 中起重要作用。 WGCNA 之后，使用方差和关联获得与先前 DEG 相交的基因。值得注意的是，最终选择了随后的九个枢纽基因：ACTL6A、BECN1、SNRPD1、POLR1B、GSK3B、PPP2CA、RBBP7、PLK4 和 RFC4，基于PPI网络和三种不同算法，其表达模式也通过RT-qPCR验证。通过深入分析，我们推测上述关键基因可能通过扰乱子宫内膜菌群稳态、损害自噬、抑制子宫内膜增殖等方式参与RIF。此外，当前的研究揭示了异常的免疫浸润模式，并强调子宫 NK 细胞 (uNK) 和 CD4+RIF 子宫内膜中的 T 细胞发生显着改变。最后，岭图显示了枢纽基因与其他基因和关键通路之间的清晰且关键的关联。结论：我们首先利用WGCNA来识别可能与RIF相关的最有潜力的九个枢纽基因。同时，本研究深入了解免疫浸润状态，揭示 RIF 的潜在免疫发病机制。这可能是下一步RIF病因学研究的一个方向。需要进一步研究来调查所涉及的机制。