Qiongjie Zhou

Changes of plasma and placental tissue factor pathway inhibitor-2 in women with preeclampsia and normal pregnancy

OBJECTIVES To investigate the maternal and fetal plasma changes of tissue factor pathway inibitor-2 (TFPI-2) and its placental expression in women with normal pregnancy and preeclampsia. MATERIAL AND METHODS We assessed the plasma TFPI-2 level in non-pregnant, normal pregnant and postpartum women, detected fetal plasma level and expression in placenta, and compared the changes in women with preeclampsia. Time-resolved fluoroimmunoassay and immunohistochemistry were used for plasma and placenta tissue detection, respectively. RESULTS Maternal plasma levels of TFPI-2 in normal pregnant women at 13weeks of gestation increased 9.3 times as compared with healthy non-pregnant women (149.3+/-17.1 versus 16.0+/-3.6ng/ml)reached a maximum level ( 282.6+/-17.1ng/ml) at 39weeks of gestation, and dramatically decreased to nearly a non-pregnant level on the first day of postpartum (32.3+/-7.1ng/ml); similar change was found in the placental expression. Fetal plasma TFPI-2 was significantly lower than the maternal level at delivery. The maternal plasma TFPI-2 in preeclampsia was significantly lower compared with that in normal pregnancy, coupled with significantly higher placental expression. CONCLUSIONS Placenta may be the main site of the high level of TFPI-2 production in maternal circulation, and the dramatic changes in preeclampsia provide a clue to elucidate its pathogenesis.

Identification of novel candidate maternal serum protein markers for Down syndrome by integrated proteomic and bioinformatic analysis

This study aimed to identify candidate protein biomarkers from maternal serum for Down syndrome (DS) by integrated proteomic and bioinformatics analysis.

14-3-3 tau (YWHAQ) gene promoter hypermethylation in human placenta of preeclampsia

INTRODUCTION Disruption of the 14-3-3 tau (YWHAQ) gene has been shown to be involved in preeclampsia (PE). The YWHAQ promoter could be differentially regulated by methylation in severe PE patients. METHODS Placental genomic DNA from patients with severe PE (n = 21) and controls who experienced a normal pregnancy (n = 16) was analyzed using dot-blot and immunohistochemistry. The placental methylation patterns of YWHAQ, expression of 14-3-3 tau and ten-eleven translocation (TET), were confirmed by bisulfite sequencing, immunohistochemistry, western blot and real-time PCR, respectively. RESULTS Genomic 5 hmC (P < 0.001), expression of 14-3-3 tau (P < 0.01) and TET (P < 0.05) were down-regulated, whereas 5 mC was up-regulated (P < 0.001) in preeclamptic placentas. Significant hypermethylation of the YWHAQ promoter was detected in PE placentas compared with control samples (19.1% vs. 9.4%, P = 0.0095). PE-specific hypermethylation of CpG2 - 4, CpG9, CpG17, CpG19 was identified in PE patients compared with controls (CpG2: 13.3% vs. 2.5%, P < 0.0001; CpG3: 14.8% vs. 3.1%, P < 0.0001; CpG4: 19.5% vs. 5.0%, P < 0.0001; CpG9: 15.7% vs. 5.0%, P = 0.0018; CpG17: 16.2% vs. 6.3%, P = 0.0003; and CpG19: 78.1% vs. 59.4%, P < 0.0001). DISCUSSION The observed participation of 14-3-3 tau in the regulation of the placental epigenome may participate in the molecular mechanisms that govern the pathological process of PE, although this requires further evaluation.

Roles of Apolipoprotein E (ApoE) and Inducible Nitric Oxide Synthase (iNOS) in Inflammation and Apoptosis in Preeclampsia Pathogenesis and Progression

Objectives To investigate potential roles of inducible nitric oxide synthase (iNOS) and apolipoprotein (apoE) in inflammation and apoptosis promoting pathological changes in preeclampsia in pregnant mice with apoE and/or iNOS knock out. Methods B6.129 mice were crossed to produce WT, apoE−/−, apoE+/−, iNOS−/−, iNOS+/− and apoE−/−iNOS−/− groups. Variants were confirmed by PCR. Serum lipid parameters (triglycerides, TG; total cholesterol, TC; high density lipoprotein, HDL; and low density lipoprotein, LDL), NO levels and placental electronic microscopic ultrastructures were evaluated, and blood pressure (BP), 24-hour urine protein and pregnancy outcomes were recorded for pregnant F1 generation mice. Placental expressions of inflammatory (tumor necrosis factor-α, TNF-α; interleukin-6, IL-6; nuclear factor-κB, NF-κb) and apoptotic markers (Bcl-2 associated X protein, Bax, B-cell lymphoma/leukemia-2, Bcl-2, and Caspase-3) were evaluated via Western blot. Results Serum lipids, BP and 24-hour urine protein levels were shown to be significantly higher and parturition and placenta weights were lower in apoE−/− and apoE−/−iNOS−/− groups (p<0.05). NO levels were lower in the apoE−/−iNOS−/− group. In addition, inflammatory/apoptosis parameters, including TNF-α, IL-6, NF-κb, Bax, Bcl-2 and Caspase-3 in the apoE−/−iNOS−/− group (p<0.01), as well as in the apoE−/− group (p<0.05), and NF-κB, Bax in iNOS−/− group (p<0.05) were higher compared with WT group. However, most of the inflammatory/apoptosis parameters in the iNOS+/− and the apoE+/− groups (p>0.05) showed no differences. In addition, placenta vascular endothelial and trophoblast cell morphological changes were demonstrated in both the apoE−/−iNOS−/− and apoE−/− groups. Conclusion Elevated lipid metabolism and inflammatory/apoptosis parameters suggest a potentially significant role of apoE in preeclampsia pathology, as well as a relationship between iNOS and preeclampsia progression.

Effects of tissue factor pathway inhibitor-2 expression on biological behavior of BeWo and JEG-3 cell lines.

Objectives: To investigate the effect of tissue factor pathway inhibitor-2 (TFPI-2) expression on biological behavior of BeWo and JEG-3 cell lines. Material and Methods: The expression of TFPI-2 in BeWo and JEG-3 cells was upregulated by pEGFP-N3-TFPI-2 and downregulated by small interference RNA transfection, confirmed by Western blotting assay and real-time polymerase chain reaction (RT-PCR). Boyden chamber, Cell Counting Kit-8 (CCK-8), and Hoechst 33258/terminal deoxynucleotidyltransferase-mediated UTP end labeling (TUNEL) assays were used for migration, invasion, and proliferation/apoptosis analysis, respectively. Results: In Western blotting and RT-PCR assay, protein and messenger RNA (mRNA) expression of TFPI-2 in transfected BeWo and JEG-3 cells were confirmed. Expression of TFPI-2 inhibited BeWo and downregulated JEG-3 cell migration, invasion, proliferation, and induced apoptosis (P < .05) in Boyden chamber, CCK-8, Hoechst 33258, and TUNEL detection, respectively. Conclusions: TFPI-2 expression caused invasion and proliferation impair and induced apoptosis in TFPI-2 regulated BeWo and JEG-3 cells. It provides a clue for potential role of TFPI-2 in trophoblast.

Risk factors for preterm premature rupture of membranes in Chinese women from urban cities

To investigate the prevalence of preterm premature rupture of membranes (PPROM) in urban areas in China and examine the associated risk factors.

Effects of magnesium sulfate on heart rate, blood pressure variability and baroreflex sensitivity in preeclamptic rats treated with L-NAME.

Objective: Magnesium sulfate is used for the treatment of preeclampsia. This study aimed to evaluate effects of magnesium sulfate on heart rate variability (HRV), blood pressure variability (BPV) and baroreflex sensitivity (BRS) in pregnant rats treated with NG-nitro-.-arginine-methyl ester (L-NAME). Methods: Sprague–Dawley rats were randomly divided into four groups: non-pregnancy and three pregnant groups. From gestational day 2–12, normal pregnancy group (NOR) received sterile water through intravenous injection, LN was injected with L-NAME [25 mg/day] to induce preeclampsia, while LNM group was also treated with magnesium sulfate at 0.3 g/kg at gestational day 13. HRV, BPV and BRS were monitored, and endothelial functions were detected at gestational day 14. Results: Rats with treatment of L-NAME showed significantly increased blood pressure and endothelin-1 concentration and decreased plasma NO concentration; concomitant treatment with magnesium sulfate suppressed blood pressure. Also, rats treated with L-NAME and magnesium sulfate underwent lower heart rate variability (lower absolute and normalized LF) and higher blood pressure variability (lower normalized HF and LF, higher normalized VLF) compared with non-pregnant or normal pregnant rats, whereas normalized LF of HRV in rats co-administered magnesium sulfate returned to normal level. Additionally, rats treated with L-NAME underwent decreased BRS-SP, BRS-NE, BRS LF and HF, and rats with concomitant magnesium sulfate partially reversed the decline (p > 0.05) and had significantly lower BRS-SP, BRS-NE. Conclusion: Sympathetic and parasympathetic impairment exist in preeclamptic rats treated with L-NAME. Magnesium sulfate may ameliorate alterations in the autonomic nervous system noted in preeclampsia.

Quantitative Proteomic Analysis of Serum from Pregnant Women Carrying a Fetus with Conotruncal Heart Defect Using Isobaric Tags for Relative and Absolute Quantitation (iTRAQ) Labeling

Objective To identify differentially expressed proteins from serum of pregnant women carrying a conotruncal heart defects (CTD) fetus, using proteomic analysis. Methods The study was conducted using a nested case-control design. The 5473 maternal serum samples were collected at 14–18 weeks of gestation. The serum from 9 pregnant women carrying a CTD fetus, 10 with another CHD (ACHD) fetus, and 11 with a normal fetus were selected from the above samples, and analyzed by using isobaric tags for relative and absolute quantitation (iTRAQ) coupled with two-dimensional liquid chromatography-tandem mass spectrometry(2D LC-MS/MS). The differentially expressed proteins identified by iTRAQ were further validated with Western blot. Results A total of 105 unique proteins present in the three groups were identified, and relative expression data were obtained for 92 of them with high confidence by employing the iTRAQ-based experiments. The downregulation of gelsolin in maternal serum of fetus with CTD was further verified by Western blot. Conclusions The identification of differentially expressed protein gelsolin in the serum of the pregnant women carrying a CTD fetus by using proteomic technology may be able to serve as a foundation to further explore the biomarker for detection of CTD fetus from the maternal serum.

microRNA expression profiling of heart tissue during fetal development

microRNAs (miRNAs) are important both in early cardiogenesis and in the process of heart maturation. The aim of this study was to determine the stage-specific expression of miRNAs in human fetal heart in order to identify valuable targets for further study of heart defects. Affymetrix microarrays were used to obtain miRNA expression profiles from human fetal heart tissue at 5, 7, 9 and 23 weeks of gestation. To identify differentially expressed miRNAs at each time-point, linear regression analysis by the R limma algorithm was employed. Hierarchical clustering analysis was conducted with Cluster 3.0 software. Gene Ontology analysis was carried out for miRNAs from different clusters. Commonalities in miRNA families and genomic localization were identified, and the differential expression of selected miRNAs from different clusters was verified by quantitative polymerase chain reaction (qPCR). A total of 703 miRNAs were expressed in human fetal heart. Of these, 288 differentially expressed miRNAs represented 5 clusters with different expression trends. Several clustered miRNAs also shared classification within miRNA families or proximal genomic localization. qPCR confirmed the expression patterns of selected miRNAs. miRNAs within the 5 clusters were predicted to target genes vital for heart development and to be involved in cellular signaling pathways that affect heart structure formation and heart-associated cellular events. In conclusion, to the best of our knowledge, this is the first miRNA expression profiling study of human fetal heart tissue. The stage-specific expression of specific miRNAs suggests potential roles at distinct time-points during fetal heart development.

Predicting lymph node metastasis in endometrial cancer using serum CA125 combined with immunohistochemical markers PR and Ki67, and a comparison with other prediction models

We aimed to evaluate the value of immunohistochemical markers and serum CA125 in predicting the risk of lymph node metastasis (LNM) in women with endometrial cancer and to identify a low-risk group of LNM. The medical records of 370 patients with endometrial endometrioid adenocarcinoma who underwent surgical staging in the Obstetrics & Gynecology Hospital of Fudan University were collected and retrospectively reviewed. Immunohistochemical markers were screened. A model using serum cancer antigen 125 (CA125) level, the immunohistochemical markers progesterone receptor (PR) and Ki67 was created for prediction of LNM. A predicted probability of 4% among these patients was defined as low risk. The developed model was externally validated in 200 patients from Shanghai Cancer Center. The efficiency of the model was compared with three other reported prediction models. Patients with serum CA125 < 30.0 IU/mL, either or both of positive PR staining > 50% and Ki67 < 40% in cancer lesion were defined as low risk for LNM. The model showed good discrimination with an area under the receiver operating characteristic curve of 0.82. The model classified 61.9% (229/370) of patients as being at low risk for LNM. Among these 229 patients, 6 patients (2.6%) had LNM and the negative predictive value was 97.4% (223/229). The sensitivity and specificity of the model were 84.6% and 67.4% respectively. In the validation cohort, the model classified 59.5% (119/200) of patients as low-risk, 3 out of these 119 patients (2.5%) has LNM. Our model showed a predictive power similar to those of two previously reported prediction models. The prediction model using serum CA125 and the immunohistochemical markers PR and Ki67 is useful to predict patients with a low risk of LNM and has the potential to provide valuable guidance to clinicians in the treatment of patients with endometrioid endometrial cancer.