Application of iTRAQ proteomics in identification of the differentially expressed proteins of placenta of pregnancy with preeclampsia.

OBJECTIVE: To explore the differential protein profile of preeclampsia and identify its potential biomarker. METHODS: Around 20 pregnant women with preeclampsia (preeclampsia group) and 20 normal-term pregnancy (normal group) were collected from 2017 to 2018 in the study. Total protein of placenta tissues was extracted, denaturized, deoxidized, and enzymolyzed. The sample was labeled with isobaric tags for relative and absolute quantitation (iTRAQ) and analyzed with mass spectrum to identify differentially expressed proteins. RESULTS: There were 37 proteins, which were differentially expressed with significance (P < 0.05). Among them, 17 proteins were upregulated and 20 proteins were downregulated with significance in the placenta of preeclampsia group compared with control group, those proteins may have an induction or protection function during the development of preeclampsia. CONCLUSION: iTRAQ technology can effectively screen the differentially expressed proteins in the placenta, which can effectively diagnose the preeclampsia during pregnancy.

miR-362-3p regulates cell proliferation, migration and invasion of trophoblastic cells under hypoxia through targeting Pax3.

Preeclampsia (PE), a common obstetrical disorder, is one of the leading causes of pregnancy associated death. PE is closely linked with impaired migration and invasion ability of trophoblastic cells. miR-362-3p recently received our particular attention due not only to its aberrant expression in the placentas of patients with PE, but also to its important roles in regulating migration and invasion of various cells. This study was thus conducted to investigate the roles of miR-362-3p in PE and the related mechanism. The expression of miR-362-3p and Pax3 was examined in placentas of patients with PE and in normal placentas. HTR8/SVneo cells were cultured under hypoxia and transfected with miR-362-3p mimics, miR-362-3p inhibitors or Pax3 over-expression vectors. Results showed up-regulation of miR-362-3p but down-regulation of Pax3 in placentas of preeclamptic pregnancies. Luciferase report assay confirmed that Pax3 is a direct target of miR-362-3p. Although Pax3 was predicted to be targeted by miR-30a-3p and miR-181a-5p as well, their expression either had no difference between placentas of PE patients and normal placentas or showed less increment in placentas of PE patients than miR-362-3p. Exposure to hypoxia inhibited cell viability, migration and invasion of HTR8/SVneo cells. Increasing miR-362-3p by the mimics conferred improved effects on the inhibition. However, deletion of miR-362-3p or overexpression of Pax3 abolished the inhibiton. These results suggest that miR-362-3p/Pax3 axis regulates cell viability, migration and invasion of HTR8/SVneo cells under hypoxia. The present study adds to the further understanding of the pathogenesis of PE.

Metformin ameliorates high uric acid-induced insulin resistance in skeletal muscle cells.

Hyperuricemia occurs together with abnormal glucose metabolism and insulin resistance. Skeletal muscle is an important organ of glucose uptake, disposal, and storage. Metformin activates adenosine monophosphate-activated protein kinase (AMPK) to regulate insulin signaling and promote the translocation of glucose transporter type 4 (GLUT4), thereby stimulating glucose uptake to maintain energy balance. Our previous study showed that high uric acid (HUA) induced insulin resistance in skeletal muscle tissue. However, the mechanism of metformin ameliorating UA-induced insulin resistance in muscle cells is unknown and we aimed to determine it. In this study, differentiated C2C12 cells were exposed to UA (15 mg/dl), then reactive oxygen species (ROS) was detected with DCFH-DA and glucose uptake with 2-NBDG. The levels of phospho-insulin receptor substrate 1 (IRS1; Ser307), phospho-AKT (Ser473) and membrane GLUT4 were examined by western blot analysis. The impact of metformin on UA-induced insulin resistance was monitored by adding Compound C, an AMPK inhibitor, and LY294002, a PI3K/AKT inhibitor. Our data indicate that UA can increase ROS production, inhibit IRS1-AKT signaling and insulin-stimulated glucose uptake, and induce insulin resistance in C2C12 cells. Metformin can reverse this process by increasing intracellular glucose uptake and ameliorating UA-induced insulin resistance.

Alpha-1-antitrypsin functions as a protective factor in preeclampsia through activating Smad2 and inhibitor of DNA binding 4.

Pre-eclampsia (PE) is one of the most common reason for high morbidity and mortality of maternal and prenatal infants. Production from oxidative stress results in maternal ROS system and anti-oxidation defense system imbalance to promote tissue ischemia and hypoxia, and ultimately impairs the maternal organs and placenta. Our previous study showed that exogenous Alpha-1-antitrypsin (AAT) and overexpression of AAT in umbilical vein cell (HUVEC) hypoxia-reoxygenation model could increase the activity of antioxidant enzymes, and played a protective role in preeclampsia animal model. In this study, we aim to investigate the underlying mechanism by which AAT prevents PE progress. Whole-exome sequencing was performed to screen the genes altered by AAT. We found that AAT knockdown altered the expression of Smad family and Id family genes, and further demonstrated that AAT positively regulated Id4 expression through activating Smad2. Reduced Id4 expression and Smad2 phosphorylation were observed in preeclampsia animal model, which was also confirmed in human placenta tissues. In addition, AAT protected HUVEC cells from hypoxia/reoxygenation injury and relieved preeclampsia symptoms through Smad2/Id4 axis. Our data illustrate AAT/Smad2/Id4 axis is an important mediator of placenta and vascular function during pregnancy. These findings provide insights into events governing pregnancy-associated disorders, such as preeclampsia.