Complement C5a Mediated Placental Dysfunction Contributes To The Pathogenesis Of Preeclampsia

Background:Preeclampsia(PE),characterized by new-onset hypertension and proteinuria after 20 weeks of pregnancy,is a severe obstetric complication leading to maternal and fetal morbidity and mortality worldwide,especially in China.Recent studies have indicated that complement system plays a vital role in the process of normal pregnancy.However,the excess activation of complement system contributes to the onset of PE.Complement C5 a is one of the most important product of complement cascade.However,the underlying mechanisms of C5 a in the pathogenesis of PE remains unknown.Objective: Make it clear that whether C5 a was involved in the onset of PE and further investigate the detailed cellular and molecular mechanisms of complement C5 a in the onset of PE and to determine whether it is associated with maternal vascular functions.Method: 1)In our study,we enrolled 24 women with PE and 32 normal pregnant women,matched for maternal age and gestational age.The clinical,laboratory and vascular characteristics were recorded.Blood and placenta samples were also collected for furthur study.2)We examined the inflammatory factors in serum of two groups of women by Bio-Plex assay and detected the subphenotype of peripheral blood monocyte cells(PBMC)by flow cytometry analysis.3)We next assessed the levels of C5 a in serum,PBMCs and placentas by ELISA,real-time PCR(RT-PCR)or Western blot.The location of C5 a and C5 a receptor(C5a R)was detected by immunofluorescence staining.4)Correlations between maternal circulating C5 a and blood pressure and arterial stiffness were calculated with Pearson correlation coefficient.5)RT-PCR and immunofluorescent staining were used to assess the angiogenic factors in placentas of two groups.6)Human extravillous trophoblast(EVT)cell line HTR-8/SVneo cells were cultured to evaluate effect of C5a-C5aR axis on proliferation,migration and tube formation abilities of trophoblasts by CCK-8,transwell and matrigel assays.7)Western blot was furthur used to estimate the effect of C5 a on the protein expression of MAPKs and AKT signaling pathways.Results: 1)clinical characterists showed that women with PE are more likely to deliver smaller babies earlier and have pregnancy complications.2)Serum levels of inflammatory factors were increased in women with PE than in normal controls and M1-like proinflammatory PBMCs were simultaneously elevated in PE groups.3)Serum levels of C5 a were significantly enhanced in women with PE.However,no significnat differences were found in C5 m RNA levels of PBMCs between two groups.Furthermore,deposition of C5 a was significantly enhanced in preeclamptic placentas and colocalized with CD11 b positive macrophages,indicating that placental macrophage-derived C5 a contribute to the onset of PE.In addition,C5aR was expressed on placental trophoblasts.4)Pearson correlation analysis indicated a significant positive correlation between serum levels of C5 a and blood pressure and arterial stiffness.5)Preeclamptic placentas showed increased expression of pro-angiogenic factors and decreased expression of anti-angiogenic factors.6)In vitro study indicated that C5aR agonist dramatically reduced the migration and tube formation abilities of HTR-8/SVneo cells.In contrast,cells transfected with C5aR si RNA rescued the abilities aforementioned.7)Further study revealed that AKT signaling pathway was involved in the regulation of trophoblast function.Conclusion: 1)PE is associated with an systemic inflammatory state.2)Elevated serum levels of C5 a were positively correlated with blood pressure and arterial stiffness.3)Placental macrophages derived C5 a contribute to the onset of PE via C5aR.4)C5a-C5aR axis regulate the migration,tube formation and agiogensis of placental trophoblasts,contributing to the placental dysfunction.