Abstract STUDY QUESTION Does abnormal serotonin homeostasis contribute to impaired endometrial decidualization in patients with recurrent implantation failure (RIF)? SUMMARY ANSWER Abnormal serotonin homeostasis in patients with RIF, which is accompanied by decreased monoamine oxidase (MAO) expression, affects the decidualization of endometrial stromal cells and leads to embryo implantation failure. WHAT IS KNOWN ALREADY Previous studies have indicated that the expression of MAO, which metabolizes serotonin, is reduced in the endometrium of patients with RIF, and serotonin can induce disruption of implantation in rats. However, whether abnormal serotonin homeostasis leads to impaired decidualization in patients with RIF and, if so, the mechanism involved, remains unclear. STUDY DESIGN, SIZE, DURATION Endometrial samples from 25 patients with RIF and 25 fertile patients were used to investigate the expression levels of monoamine oxidase A (MAOA), monoamine oxidase B (MAOB) and serotonin. We isolated human endometrial stromal cells to investigate the role of MAOA, MAOB and serotonin in inducing decidualization in vitro and further explored the underlying mechanism using RNA sequencing (RNA-seq) and liquid chromatography-mass spectrometry (LC/MS) analyses. PARTICIPANTS/MATERIALS, SETTING, METHODS The levels of serotonin in the endometrium of patients with RIF were detected by enzyme-linked immunosorbent assay (ELISA) and immunohistofluorescence, and the key genes involved in abnormal serotonin metabolism were analysed via combination with single-cell sequencing data. The effects of MAOA or MAOB on the decidualization of stromal cells were investigated using an in vitro human endometrial stromal cell-induced decidualization model and a mouse artificially induced decidualization model. The potential mechanisms by which MAOA and MAOB regulate decidualization were explored by RNA-seq and LC/MS analysis. MAIN RESULTS AND THE ROLE OF CHANCE We found that women with RIF have abnormal serotonin metabolism in the endometrium and attenuated MAO in endometrial stromal cells. Endometrial decidualization was accompanied by increased MAO in vivo and in vitro. However attenuated MAO caused an increased local serotonin content in the endometrium, impairing stromal cell decidualization. RNA-seq and LC/MS analyses showed that abnormal lipid metabolism, especially phosphatidylcholine metabolism, was involved in the defective decidualization caused by MAO deficiency. Furthermore, decidualization defects were rescued by phosphatidylcholine supplementation. LARGE SCALE DATA RNA-seq information and raw data can be found at NCBI Bioproject number PRJNA892255. LIMITATIONS, REASONS FOR CAUTION This study revealed that impaired serotonin metabolic homeostasis and abnormally reduced MAO expression were among the reasons for RIF. However, the source and other potential functions of serotonin in the endometrium remain to be further explored. WIDER IMPLICATIONS OF THE FINDINGS This study provides new insights into the mechanisms of serotonin homeostasis in human endometrial decidualization and new biomarkers or targets for the treatment of patients with RIF. STUDY FUNDING/COMPETING INTEREST(S) X.Sheng. is supported by grants from the National Natural Science Foundation of China (82001629), the Youth Program of Natural Science Foundation of Jiangsu Province (BK20200116), and Jiangsu Province Postdoctoral Research Funding (2021K277B). H.Sun. is supported by grants from the National Natural Science Foundation of China (82030040). G.Yan. is supported by grants from the National Natural Science Foundation of China (82171653). The authors declare no conflicts of interest.

Dysregulation of endometrial stromal serotonin homeostasis leading to abnormal phosphatidylcholine metabolism impairs decidualization in patients with recurrent implantation failure