The Study On The Expression And Function Of DCAF13 In The Decidua Of Preeclampsia Patients

Preeclampsia（PE）is a syndrome occurring during pregnancy in which hypertension（≥140/90mm Hg）and/or proteinuria（≥0.3g/L）develops after 20 weeks of gestation in women with normal blood pressure,which affects more than eight million mother-infant pairs each year.PE is a specific disease in humans and severe PE can cause uterine and placental dysfunction,intrauterine growth restriction and premature birth.So far,the only and most reliable treatment for PE is delivery,but the pathogenesis of PE is still an enigma.About the pathogenesis,two-stage theory is now widely accepted:In stageⅠ,in the latent phase before the onset of symptoms during pregnancy,there is impaired remodelling of the spiral arteries,which inhibits the development of the placenta;In stageⅡ,ischemic-hypoxic metabolic disorders that allow for the release of large amounts of undesirable factors,thereby leading to an inflammatory response in the mother,which eventually results in PE.Eclampsia is defined when PE worsens and epilepsy occurs.To date,most previous studies on the pathogenesis of PE have been focused on placenta.However,decidualization is the foundation for placentation and growth.After pregnancy,as the blastocyst implants,the endometrium thickens,capillaries fill up,blood supply becomes abundant and the endometrial stromal cells become morphologically larger and rounder.This series of changes is known as the decidua reaction,at which point the endometrium is transformed into the decidua.The endometrium undergoes a radical change during decidualization,allowing the endometrium to adjust to an acceptable state,which provides a solid foundation for the successful implantation of the blastocyst.Dysfunction of the decidua has been reported in a variety of obstetric complications,including recurrent pregnancy failure and preterm labour.More and more studies have shown that abnormal decidualization is an important factor leading to PE.Thus,the formation of decidua is important in the initiation and maintenance of pregnancy.DCAF13（DDB1 and CUL4 associated factor 13）is a novel adaptin of the CRL4 E3ubiquitin ligase complex.Its function is to recruit SUV39H1 to the CRL4 E3 ubiquitin ligase,thereby allowing SUV39H1 to be ubiquitinated and degraded,leaving H3K9me3 at a low level and ultimately allowing transcription of r DNA and a subset of genes associated with germ layer differentiation to begin.In addition,DCAF13 is essential for the maintenance of follicles and the growth and development of oocytes.However,the role of DCAF13 in decidualization remains unclear.Importantly,after searching The Human Protein Atlas website,we found that DCAF13was expressed at high levels in the deicidua.In addition to this,pre-laboratoryRNA sequencing of decidua collected from PE and normal pregnant women revealed that Dcaf13 expression was significantly downregulated in deciduas of PE.The relationship between PE,decidualization and DCAF13 has led us to consider and explore the relationship between the three.In the present study,a combination of multiple approaches demonstrated that DCAF13plays an integral role in the decidualization process of endometrial stromal cells.Firstly,it was found that DCAF13 levels were significantly lower in the decidua of pregnant women with PE compared to normal decidua,both at the protein level and at the mRNA level.Secondly,after dosing induced decidualization of endometrial stromal cells,DCAF13 expression was found to be increased during the decidualization process.We thus came to the tentative conclusion that DCAF13 is most likely to be involved in decidualization.Therefore,further transfection of siRNA to interfere with Dcaf13 and manual induction of decidualization in endometrial stromal cells revealed that the cells did not appear decidualization:the cell morphology did not become larger and rounder,and PRL and IGFBP1 were significantly reduced,suggesting that reduced expression of DCAF13 would prevent decidualization.To further investigate the role of DCAF13 in decidualization,we used a mouse model to further explore the link between DCAF13 and decidualization.To begin with,we explored wild-type mouse with normal gestation and found that there was a significant trend towards higher expression of DCAF13 after the mouse underwent decidualization.Besides,we also obtained conditional knockout Dcaf13 mouse(PR Cre^+/-;Dcaf13^fl/fl,cKO)using PR Cre tool mouse and found no significant difference in the morphology and number of ovaries and oocytes in cKO mouse.In addition,we found no significant differences in Dcaf13 expression in the ovaries of cKO mouse,but in the uterus,Dcaf13 expression was significantly downregulated.After HE staining,the uterus of cKO mouse was found to have smaller structures in each part than the normal group,although there were longitudinal muscles,circular muscles,stroma and luminal epithelium.The cKO mouse was found to be sterile after being mated with wild-type mouse.After artificially inducing decidualization of the mouse uterus,two decidual regulators（Bmp2 and Wnt4）were detected by qPCR and found that the expression of both Bmp2 and Wnt4was significantly increased in the control uterus after induction of decidualization,but not significantly different in the cKO mouse.Moreover,the endometrial stromal cells of cKO mouse were found to be fibrous and tightly arranged by HE staining.We further examined Ki67,a marker molecule for cell proliferation,and found that this protein was barely expressed in the endometrium of cKO mouse.Thus it was confirmed that the uterus of cKO mouse did not undergo decidualization.In summary,DCAF 13 plays a pivotal role in the normal functioning of the decidualization.Abnormal expression of DCAF13 affects the process of decidualization,which in turn is involved in the development of PE.