Aldosterone Regulation And Function On Human Decidualization

Successful pregnancy is a physiological process,including embryo implantation,decidualization,placentation and parturition.The epithelial differentiation of endometrial stromal cells is called decidualization,which is strictly regulated by many hormones and genes,and has functions on control of trophoblast invasion,protection of the conceptus from maternal immune rejection and contribution of nutrients for embryo development.Secretory glands in mid-secretory phase of menstrual cycle produce numerous cytokines or other substances to promote embryo development,embryo implantation and decidualization.Aldosterone,one important mineralocorticoid,is responsible for various physiological and pathological processes.This research examined the protein expression for RENIN,ACE,AT1 R and CYP11B2 in human endometrium of different phase of menstrual cycle by immunohistochemistry.In cultured human endometrial stromal cells,we next explored the function of aldosterone on decidualization by real time-PCR,Western blot,and immunofluorescence.Moreover,promoter prediction and luciferase reporter assay were used to examine the MR regulation on LKB1.Last,we used Co-immunoprecipitation to analyze the interaction between PDK4 and p-CREB.In this study,AT1 R and CYP11B2 were specifically expressed in endometrial glandular epithelium during mid-secretory phase,while decreased during late secretory phase.According to the immunohistochemistry analysis of MR protein in endometrium,we found that MR expression in stromal cells was elevated during mid-secretory phase compared with mid-proliferative and late-secretory phases,suggesting that aldosterone synthesized in secretory glands may have a function on decidualization through paracrine interaction.After analysis of decidualization markers in decidual cells treated with aldosterone,we found aldosterone can contribute to decidualization through activation of p-CREB/FOXO1.From the result of immunohistochemistry,PDK4 increased in both glands and stromal cells during mid-secretory phase while decreased in mid-proliferative and late-secretory phases.Because the tetramer form of PKM2 did not change in decidual cells,we speculated Warburg effect did not work in h ESCs undergoing decidualization.Therefore,the effect of PDK4 on decidualization may not be inactivation of PDHE1?,which can promote Warburg effect.Knockdown of PDK4 will significantly reduce markers of decidualization and p-CREB/FOXO1 signaling pathway.Due to result that silence of PDK4 did not promote decidualization,the impact of PDK4 on decidualization is independent on activation of PDHE1?.Based on co-immunoprecipitation,PDK4 can bind with p-CREB to prevent the ubiquitination of p-CREB.Thus,p-CREB can transport to nucleus to bind CBP/P300,promoting decidualization by up-regulation of FOXO1.The activation of AMPK can stimulate expression of PDK4 and its downstream p-CREB/FOXO1 signaling,and silence of AMPK will restrain PDK4/p-CREB/FOXO1 signaling,indicating PDK4 in decidualization was regulated by AMPK.From the analysis of LKB1 promoter,we found MR has a binding site in LKB1 promoter.The further verification by luciferase reporter assay showed that aldosterone-MR complex can directly regulate LKB1 in nucleus.The induction of LKB1 can activate PDK4/p-CREB/FOXO1 through phosphorylation of AMPK.Inhibition of 11?-HSD2 can prevent aldosterone-MR complex to enter nucleus,leading to blockade of LKB1/p-AMPK/PDK4/p-CREB/FOXO1.These data showed that aldosterone can activate MR to facilitate decidualization through LKB1/p-AMPK/PDK4/p-CREB/FOXO1 signaling pathway.In conclusion,aldosterone can be synthesized from endometrial glandular epithelium during mid-secretory phase and promote decidualization by activation of LKB1/p-AMPK/PDK4/p-CREB/FOXO1 signaling pathway.