Advanced Maternalage-associated SIRT1 Deficiency Compromises The Trophoblast Epithelial-mesenchymal Transition Through The Upregulation Of Vimentin Acetylation

Background:Advanced maternal age（AMA）is usually defined as maternal age≥35 years.With the rapid increase of AMA pregnancies,more and more evidence suggests that AMA is associated with various adverse pregnancy outcomes,such as preeclampsia（PE）,gestational diabetes mellitus（GDM）,preterm birth（PTB）and intrauterine fetal death.Many literatures have confirmed that advanced maternal age is an independent risk factor for adverse pregnancy outcomes but there is a lack of in-depth studies on the underlying etiological mechanisms.The pathological features of placental premature senescence such as poor vascular perfusion,chorionic villi maturation disorder and extensive vascular lesions strongly suggest that placental premature senescence and trophoblast dysfunction may be important reasons for the increased incidence of adverse pregnancy outcomes caused by AMA.SIRT1 is an NAD⁺-dependent deacetylase with well-known antiaging effects,but its connection with placental senescence is unreported.This hypothesis will be validated from the perspective of tissue,cell and individual using techniques of molecular biology,cell biology and pharmacology,the results of this study may elucidate the possibility of regulating trophoblast function by targeting trophoblast SIRT1,which could further provide new therapeutic strategy for AMA pregnant women.Methods:First,human term placenta and early pregnancy villi were collected from AMA women（≥40 years old）and the control group（<30years old）,and the protein expression levels of Sirtuins family and senescence related markers were detected by Western blot.Immunohistochemistry（IHC）and immunofluorescence（IF）were used to determine the expression and localization of SIRT1 in human villi and term placenta.Mouse AMA model was established by cross breeding young and aged male and female C57 mice.SIRT1 expression and activity in HTR8/SVneo cells were manipulated by sh RNA transfection or treatment with a SIRT1 activator or inhibitor.Trophoblast-specific Sirt1-KO mouse placentas were generated by mating Elf5-Cre and Sirt1^fl/flmice.Trophoblast cell mobility was assessed with transwell invasion and wound-healing assays.SIRT1-binding proteins in HTR8/SVneo cells and human placental tissue were identified by mass spectrometry.Results:We demonstrated that SIRT1 is the only differentially expressed sirtuin between AMA and normal placentas.SIRT1 is mainly located in cytotrophoblast（CTBs）and extravillous trophoblast（EVTs）.SIRT1 was also decreased in the placenta of aged female mice,and the placental junctional zone（Jz）was significantly narrowed.SIRT1 loss upregulates P53 acetylation and P21 and impairs trophoblast invasion and migration.Sirt1-KO mouse placentas exhibit senescence markers and morphological disruption,with decreased placental weight and fetal weight.In trophoblast,SIRT1 interact with vimentin to regulate its acetylation level and affect the epithelial mesenchymal transition（EMT）process.Conclusion:SIRT1 promotes trophoblast EMT to gain invasiveness by modulating vimentin acetylation.AMA placentas are associated with premature senescence during placentation due to SIRT1 loss.Therefore,SIRT1 may be a therapeutic target for improving placental development and perinatal outcomes in AMA pregnancies.