| Background and Objective:Preeclampsia(Pre-eclampsia,PE)is a serious risk of perinatal maternal and child safety of pregnancy-specific disease,mainly characterized by hypertension,proteinuria.The etiology and pathogenesis is not clear,the treatment method is still lack of targeted.At present,the maternal immune dysfunction is an important part of the process of preeclampsia.Decidua-derived mesenchymal stem cells(DMSCs)are a class of immunodeficient adult stem cells with high immunity and low immunogenicity,which can be secreted by autocrine and paracrine Cytokines,in the body’s natural immune and acquired immunity to play a strong immune regulation.In the case of ischemia and hypoxia at the interface of the mother and child,DMSCs can undergo a series of adaptive changes,including changes in quantity and function,among which the most significant changes are changes in cell migration capacity.Chemokines CXCL12 is a chemokine that is secreted by interstitial cells and epithelial cells,and binding with its specific receptor CXCR4,plays an important role in embryonic formation,stem cell mobilization,tumor migration and metastasis by mediating downstream signaling pathway.However,it has not been reported in the pathogenesis of preeclampsia.In this study,we will study the changes of DMSCs migration behavior in patients with preeclampsia on CXCL12 / CXCR4 axis,and further clarify the pathogenesis of PE,and provide DMSCs for the treatment of PE The experimental basis.Methods:1.Isolation,culture and identification of decidual mesenchymal stem cellsTo collect the tissue samples(decidual tissue,umbilical cord tissue,placental tissue)and umbilical cord blood and peripheral blood samples of cesarean section in 14 cases of preeclampsia patients from the Daping Hospital of the Third Military Medical University,the same method to collect normal term cesarean section of the pregnant women of the above specimens,a total of 11 cases,the normal control group.The mesenchymal stem cells were extracted from decidual tissue by type Ⅱ collagenase digestion and adherent.The expressions of surface markers(such as CD90,CD105,CD44,CD73,CD45,CD34,CD11 b,CD19 and HLA-DR)were analyzed by flow cytometry,and the cells were induced by stem cell three-line differentiation medium.thereby identifying the extracted cells as mesenchymal stem cells derived from decidual tissue.2.Study on the changes of CXCL12 / CXCR4 in patients with preeclampsiaThe third generation DMSCs with good growth status were used to detect CXCR4 and HIF-1α expression in DMSCs and normal pregnant women by PCR and Western blot.The expression of CXCL12 in umbilical cord,placenta and decidua of PE patients and normal pregnant women was analyzed by immunohistochemistry.In addition,the use of ELISA method to detect PE patients and normal pregnant women umbilical cord blood and peripheral blood CXCL12.3.CXCL12 / CXCR4 signal axis-mediated migration of DMSCs in PEThe third generation DMSCs with good growth status were used to compare the migration ability of PE patients and normal pregnant women by transwell chamber migration experiment.The transwell cells were incubated in 21% O2 concentration and 1% O2 concentration in cell incubator 24 h,to explore the hypoxia on DMSCs migration behavior changes.4.Statistical methodsThe data were analyzed by SPSS19.0,Graph Pad Prism 6.0,Image-Pro plus6.0 software.The comparison between the two groups was based on the t test of independent samples.The comparison between multiple groups was analyzed by one-way ANOVA Completed,that P <0.05 was statistically significant.Results:1.Successful extraction of mesenchymal stem cells from decidual tissueThe cells extracted from decidual tissue express CD40(+),CD10(+),CD44(+),CD73(+),CD45(-),CD34(-),CD11b(-),CD19(-),HLA-DR(-),which is consistent with MSC surface marker expression.Transgenic mesenchymal stem cells showed that the cells were able to differentiate into osteoblasts,chondrocytes and adipocytes.Therefore,we believe that cells extracted from decidual tissue are mesenchymal stem cells and can be used for subsequent experiments.2.Study on the changes of CXCL12/CXCR4 in patients with preeclampsiaQuantitative PCR and Western Blot showed that CXCR4 and HIF-1α in DMSCs were significantly higher than those in normal control group(P<0.05).Immuno-histo chemical results showed that the expression of CXCL12 in PE tissues showed a gradient of decidua>placenta>umbilical cord,but showed umbilical cord>placenta>decidua in normal pregnant women.The levels of CXCL12 in decidual tissue of PE patients were significantly higher than those in the normal group(P<0.05),but there was no significant difference in the content of CXCL12 between umbilical cord and placenta.The levels of CXCL12 in umbilical cord blood of PE patients were significantly higher than those of peripheral blood(P<0.05).There was no significant difference in the levels of CXCL12 in umbilical cord blood and peripheral blood in the normal control group(P>0.05).The levels of CXCL12 in the peripheral blood of the normal control group were significantly higher than those in the peripheral blood of PE patients(P<0.05).There was no significant difference in the levels of CXCL12 in umbilical cord blood between the two groups.3.CXCL12/CXCR4 signal axis-mediated DMSCs migration behavior in PEThe migration of DMSCs was observed under the induction of CXCL12,while the migration ability of DMSCs was significantly decreased after AMD3100 was inhibited by CXCR4(P<0.05).The migration ability of DMSCs in CXCL12/CXCR4 signal axis was significantly higher than that in normal control group(P<0.05).In the hypoxic environment,the migration ability of DMSCs was significantly decreased(P<0.05)compared with that in normoxic environment.Conclusions:1.The expression of CXCR4 in decidual tissue and umbilical cord blood of PE patients was significantly higher than that of normal group,and the expression of CXCR4 in DMSCs was significantly higher than that in normal group,which may be related to the change of DMSCs migration ability.2.The concentration gradient of CXCL12 in the maternal-fetal interface of PE patients(decidua>placenta>umbilical cord,Umbilical cord blood >peripheral blood)was opposite to that of normal pregnant women,(umbilical cord> placenta> decidua).Which plays a role in the migration of DMSCs to the interface of maternal fetus,and provides some experimental basis for the application of DMSCs in the treatment of PE.3.The migration of DMSCs could be mediated by CXCL12,which was mediated by CXCL12 chemokine,from low concentration to high concentration.The PE patients in normoxia and hypoxia environment,the migration capacity were significantly higher than the normal control group DMSCs,indicating that the DMSCs of PE patients can migrate to the decidual layer with the concentration gradient of CXCL12,and play an immuno regulatory function,which may play a role in the occurrence and development of PE. |
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