| Objective: To detect the expression of WAVE2in placentas and maternalsera of preeclamptic and normal women. To evaluate the reactive oxygenspecies (ROS) levels in placentas of this two groups, and analyze thecorrelation between these two mediators. In addition, an in vitro model ofhypoxia/reoxygenation (H/R) was utilized to mimic oxidative stress introphoblasts to explore the relationship between oxidative stress andWAVE2, and to analyze the role of them in the pathology ofpreeclampsia.Methods:(1) Twenty women with preeclampsia and twenty three normalterm pregnancy, who delivered from August2011to February2012in thedepartment of Obstetrics at the First Affiliated Hospital of ChongqingMedical University, were recruited and divided into PE group and controlgroup. The immunohistochemical streptravidin-biotin peroxidase methodwas employed to study the localization of WAVE2protein in placentaltissues. Quantitative real-time polymerase chain reaction was employed to assay the levels of WAVE2mRNA, and western blotting was used toassess the levels of WAVE2protein. The concentrations of WAVE2inmaternal sera were detected by ELISA. Furthermore, tissue homogenateswas applied to determine the level of ROS. The correlation between ROSlevel and WAVE2protein were analyzed.(2) The HTR-8/SVneo cells(immortalized human first trimester extravillous trophoblast cells) wereexposed to H/R condition which was employed to mimicischemia/reperfusion insult of placental trophoblasts. The HTR-8/SVneocells were pre-incubated overnight before H/R intervention. Afterexposure to H/R and normoxic conditions for48hours respectively, flowcytometry was employed to analyze intracellular ROS level and cellapoptosis in HTR8/SVneo cells. Moreover, transwell migration andinvasion assays were utilized to analyze the migration and invasion ofHTR8/SVneo cells respectively. CCK8assay was performed to detect theproliferation of HTR8/SVneo cells. Furthermore, cellimmunofluorescence and western blotting were performed to determinethe location and expression of WAVE2in trophoblasts. The levels ofWAVE2in cell supernatant were evaluated by ELISA.Results:(1) WAVE2protein mostly located in cytoplasm of trophoblastsand vascular endothelial cells. Placenta tissues of preeclampsia groupshowed weak staining. Moreover, the relative levels of WAVE2mRNAwere significantly decreased in preeclampsia groups compared to the levels of WAVE2protein in preeclampsia group and control group were0.63±0.08and1.34±0.19respectively, with significant differencesbetween these two groups (P <0.05). Compared with the normalpregnancies, the levels of WAVE2protein in maternal sera weresignificantly reduced in the preeclamptic patients [(1.08±0.12) ng/ml vs(0.62±0.09) ng/ml, P <0.05]. The level of ROS in placenta in thepreeclampsia group was significantly higher than that in control group[(144.2±12.3) nmol.mg-1.prot-1vs (75.2±8.7) nmol.mg-1.prot-1, P <0.05]. There is a significant negative correlation between ROS level andWAVE2protein expression in preeclamptic placenta(r=-0.726, P=0.000).(2) H/R exposure induced significant high levels of ROS anddecreased WAVE2expression. Moreover, WAVE2protein in cellsupernatant got from H/R condition was also lower than control group[(0.29±0.05) ng/ml vs (0.68±0.08) ng/ml, P <0.05]. Furthermore, H/Rexposure could also lead to increased cell proliferation inhibition, highercell apoptosis and decreased cell migration and invasion.Conclusion:(1) Excessive oxidative stress in placentas of preeclampticwomen was involved in the development of PE and correlated with thedecreased expression of WAVE2.(2) The in vitro study demonstrated thatH/R-induced oxidative stress could lead to the decreased of WAVE2introphoblasts which providing a good in vitro model for further study that the upstream mechanism of the regulation of WAVE2expression, andalso providing the novel targets in the pathogenesis in PE. |
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