| Decidualization is a process in which uterine stromal cells proliferate and differentiate into decidual cells.It is essential for successful pregnancy in many female animals.Embryo implantation requires the coordination of the fetomaternal interface,and the maternal level requires endometrial decidualization to reach a tolerant state.Melatonin（MT）is an indole hormone that plays a dual role in the reproductive process.Studies have shown that melatonin can not only promote the development of follicles in female animals,but also cause anti-reproductive phenomena in animals.Long-term use of melatonin can cause ovulation disorders and infertility in women.Melatonin may play an important role in the process of decidualization in female animals.This study was conducted to investigate the effect of melatonin on the proliferation and differentiation of uterine stromal cells during decidualization and the regulatory mechanism of decidualization.The results of the study showed that the model of decidualization was successfully induced in vitro,and most of the uterine stromal cells remained in the G0/G1 phase during the decidualization process.In order to study the effect of melatonin on the differentiation of uterine stromal cells,the two differentiation marker genes insulin-like growth factor binding protein 1（IGFBP1）and prolactin（PRL）in uterine stromal cells were detected by fluorescence quantitative PCR changes in m RNA levels.The results found that the two were reduced significantly,indicating that melatonin can inhibit the differentiation process of uterine stromal cells.The effects of melatonin and MT2 receptor inhibitor 4-P-PDOT on the content of superoxide anion（O2-）in cells were detected by flow cytometry.It was found that melatonin can increase the level of O2-in cells,and 4-P-PDOT can inhibit the action of melatonin.Then detect the addition of melatonin,melatonin and 4-P-PDOT to detect the changes of glutathione peroxidase（GPX）,glutathione reductase（GR）,reduced glutathione（GSH）and malondialdehyde（MDA）in cells.The results showed that melatonin can reduce the levels of GPX,GR and GSH in cells,and increase the levels of O2-and MDA in cells,thereby affecting cell differentiation.Melatonin can also inhibit cell differentiation by inhibiting NOTCH1 in the NOTCH signaling pathway and the content of recombination signal binding protein for immunoglobulin kappa J region（RBPJ）m RNA,and adding NOTCH1 recombinant protein（r NOTCH1）can increase the levels of IGFBP1 and PRL m RNA.It is suggested that melatonin inhibits the differentiation process of uterine stromal cells by inhibiting NOTCH signaling pathway.In order to further study the downstream target genes that melatonin plays in the NOTCH signaling pathway,fluorescence quantification and dual luciferase experiments have proved that melatonin can reduce the level of Nuclear factor E2-related factor 2（NRF2）m RNA in cells and reduce the activity of antioxidant response element ARE,while r NOTCH1 can reduce the inhibitory effect of melatonin on NRF2,the NRF2 inhibitor ML385further enhanced the inhibition of melatonin differentiation.This effect can continue to affect the expression of Forkhead box O1（FOXO1）in uterine stromal cells,and ultimately affect cell decidualization.In summary,this experiment is based on the decidualization model to study the effect of melatonin on decidualization of uterine stromal cells and its regulatory mechanism.The results show that melatonin binds to the MT2 receptor.On the one hand,it increases the level of superoxide anion in the cell by inhibiting the production of antioxidants in the cell to inhibit cell differentiation.On the other hand,it inhibits the cleavage of NOTCH1,thereby inhibiting the nuclear transcription factors RBPJ and NRF2,thereby inhibiting the expression of FOXO1,and finally inhibiting the differentiation of uterine stromal cells,achieving an inhibitory effect on decidualization. |
