Increased AIF-1-mediated p38 MAPK phosphorylation in the mid-secretory endometrium impairs endometrial receptivity in women with recurrent implantation failure

Abstract STUDY QUESTION How does allograft inflammatory factor-1 (AIF-1) affect endometrial receptivity in women with recurrent implantation failure (RIF)? SUMMARY ANSWER Significant upregulation of AIF-1 in the endometrial stromal cells of women with RIF inhibits cell proliferation and decidualization via the p38 mitogen-activated protein kinase (MAPK) phosphorylation, thereby reducing endometrial receptivity. WHAT IS KNOWN ALREADY RIF is a challenging clinical issue, with AIF-1, a cytokine-inducible protein linked to allograft rejection, potentially contributing to its pathogenesis. However, the precise mechanisms remain elusive. STUDY DESIGN, SIZE, DURATION This study analyzed endometrial tissue samples from women diagnosed with RIF and a control group of fertile women from December 2018 to December 2023. Single-cell RNA sequencing (scRNA-seq) data from public datasets (GSE111976, GSE250130, GSE183837) were integrated to characterize AIF-1 expression patterns in endometrium. Isolated human endometrial stromal cells (HESCs) from the human endometrium and an endometrial stromal cell line were used for in vitro analysis, and an in vivo mouse model with AIF-1 overexpression in the uterus was employed to evaluate implantation outcomes. PARTICIPANTS/MATERIALS, SETTING, METHODS Mid-secretory endometrial samples were collected from the 18 patients with RIF and 18 control patients; endometrial samples from another five different phases during the menstrual cycle were collected from 30 additional control patients. Quantitative PCR, western blot, immunohistochemical and immunofluorescence analyses, and RNA sequencing were conducted to determine the expression levels of AIF-1 and various markers. Cell proliferation, decidualization, and trophoblast outgrowth were measured. AIF-1 overexpression and gene silencing were achieved by plasmid injection and short hairpin RNA, respectively. For in vivo experiments, CD-1 mice with intrauterine injection of an AIF-1 plasmid were used. Phosphorylation of p38 was inhibited by PD169316. MAIN RESULTS AND THE ROLE OF CHANCE Based on scRNA-seq analysis and our own endometrial tissue detection, AIF-1 was significantly increased in HESCs in patients with RIF compared with their control group during the mid-secretory phase. AIF-1 overexpression resulted in reduced cell proliferation, inadequate cell decidualization, and diminished embryo outgrowth in in vitro experiments, and it reduced the number of embryo implantation sites in CD-1 mice; these effects were mitigated by PD169316, an inhibitor of p38 MAPK. LIMITATIONS, REASONS FOR CAUTION Although the study establishes a link between increased AIF-1 expression in endometrial stromal cells and reduced endometrial receptivity, the role of AIF-1 in endometrial macrophages during embryo implantation remains unclear. WIDER IMPLICATIONS OF THE FINDINGS The findings suggest that targeting the AIF-1 and p38 MAPK pathway could serve as a promising therapeutic strategy to improve endometrial receptivity in RIF patients. STUDY FUNDING/COMPETING INTEREST(S) This study was supported by the National Key R&D Program of China (No. 2022YFC2703800) and the National Natural Science Foundation of China [Nos. 82271703; 82371704; 82071596; 82071712; 82101800; 81701513]. The authors declare that they have no conflict of interests with the contents of this manuscript. TRIAL REGISTRATION NUMBER N/A.