Environmental Enrichment Protects Offspring of a Rat Model of Preeclampsia from Cognitive Decline.

Preeclampsia affects 5-7% of all pregnancies and contributes to adverse pregnancy and birth outcomes. In addition to the short-term effects of preeclampsia, preeclampsia can exert long-term adverse effects on offspring. Numerous studies have demonstrated that offspring of preeclamptic women exhibit cognitive deficits from childhood to old age. However, effective ways to improve the cognitive abilities of these offspring remain to be investigated. The aim of this study was to explore whether environmental enrichment in early life could restore the cognitive ability of the offspring of a rat model of preeclampsia and to investigate the cellular and molecular mechanisms by which EE improves cognitive ability. L-NAME was used to establish a rat model of preeclampsia. The spatial learning and memory abilities and recognition memory of 56-day-old offspring were evaluated by the Morris water maze and Novel object recognition (NOR) task. Immunofluorescence was performed to evaluate cell proliferation and apoptosis in the DG region of the hippocampus. qRT-PCR was performed to examine the expression levels of neurogenesis-associated genes, pre- and postsynaptic proteins and inflammatory cytokines. An enzyme-linked immune absorbent assay was performed to evaluate the concentration of vascular endothelial growth factor (VEGF) and inflammatory cytokines in the hippocampus. The administration of L-NAME led to increased systolic blood pressure and urine protein levels in pregnant rats. Offspring in the L-NAME group exhibited impaired spatial learning ability and memory as well as NOR memory. Hippocampal neurogenesis and synaptic plasticity were impaired in offspring from the L-NAME group. Furthermore, cell apoptosis in the hippocampus was increased in the L-NAME group. The hippocampus was skewed to a proinflammatory profile, as shown by increased inflammatory cytokine levels. EE improved the cognitive ability of offspring in the L-NAME group and resulted in increased hippocampal neurogenesis and synaptic protein expression levels and decreased apoptosis and inflammatory cytokine levels. Environmental enrichment resolves cognitive impairment in the offspring of a rat model of preeclampsia by improving hippocampal neurogenesis and synaptic plasticity and normalizing the apoptosis level and the inflammatory balance.

Perfluorooctane sulfonate exposure induces preeclampsia-like syndromes by damaging trophoblast mitochondria in pregnant mice.

Preeclampsia, defined as a hypertensive disorder during pregnancy, is a major cause of maternal and fetal mortality. Observational studies have shown that the exposure of per- and polyfluoroalkyl substances, such as perfluorooctane sulfonate (PFOS), is emerging as a significant environmental factor associated with preeclampsia risk. However, epidemiologic evidence is of correlative in nature, and unable to establish a causal relationship. Here, we established an animal model of PFOS-induced preeclampsia to explore the molecular mechanism of PFOS in placental trophoblast. In the mouse model, PFOS exposure by gavage at a dose of 10 mg/kg/d from embryonic day 7.5-16.5 was sufficient to induce preeclampsia-like symptoms such as hypertension, proteinuria, and renal glomerular endotheliosis, accompanied with placental abnormal stromal collagen deposition. In-vitro experiments of JEG-3 cells, PFOS exposure impaired trophoblast motility including the compromised abilities of migration, invasion and vascularization. Mechanistically, these pathological effects on cells resulted from SLC25A5-mediated mitochondrial damages, characterized by excessive ROS generation, decreased ATP production and mitochondrial membrane potential loss, and accompanied by the activation of p38 MAPK and JNK signaling pathways. This pioneering study provided biological plausibility to the causality verified by the animal model and the in vitro experiments, which indicates that PFOS exposure may cause preeclampsia during pregnancy via impairing trophoblast mitochondria.

Changing trends of adverse pregnancy outcomes with maternal age in primipara with singleton birth: A join point analysis of a multicenter historical cohort study in China in 2011-2012.

INTRODUCTION: Adverse pregnancy outcomes are related to two opposite maternal factors, youth and aging. However, the change in trend of specific outcomes with childbearing age is unknown. The aim of this study was to investigate the changing trend of various maternal and neonatal outcomes with maternal age from 17 to 44 years. MATERIAL AND METHODS: Data were extracted from the medical records from 2011-2012 of 39 public hospitals of 14 cities in China. The eligibility criteria were primiparity and singleton birth. Join point regression analysis was used to estimate the percent change per year of age (PCA) to explore the trends of adverse pregnancy outcomes with regard to maternal age and to identify the join point of maternal age when the trend was changed. RESULTS: A total of 89 171 women were eligible for analysis. There were four categories of trend styles. Continuously increasing trends were linear for placenta previa (PCA, 0.1%), placenta implantation (PCA, 0.09%) and postpartum hemorrhage (PCA, 0.22%), and nonlinear for gestational diabetes mellitus with one join point of 23 years (PCA, 0.17% and 0.71%) and cesarean section with four join points of 25, 28, 31 and 36 years (PCA, 1.39%, 0.34%, 1.51%, 3.49% and 0.94%). Continuously decreasing trends were linear for intrahepatic cholestasis (PCA, -0.02%) and nonlinear for anemia with one join point of 28 years (PCA, -0.49% and -0.04%). The bidirective trends were downward to upward for preterm birth (PCA, -2.93%, -0.36% and 0.38%), hypertension in pregnancy (PCA, -0.09%, 0.47%), low birthweight (PCA 0.51% and 0.38%), low 1-minute Apgar score (PCA, -0.28% and 0.07%), low 5-minute Apgar score at (PCA, -0.10% and 0.06%) and neonatal intensive care unit admission (PCA, -1.92%, -0.07% and 0.12%) with a nadir age of 28 years. The bidirective trend was upward to downward for macrosomia (PCA, 0.39% and -0.11%), with a peak age of 30 years. CONCLUSIONS: The changes in the trend of specific outcomes were different with maternal age, which means that youth, aging or both could affect the outcomes.

A hidden proteome encoded by circRNAs in human placentas: Implications for uncovering preeclampsia pathogenesis.

BACKGROUND: CircRNA-encoded proteins (CEPs) are emerging as new players in health and disease, and function as baits for the common partners of their cognate linear-spliced RNA encoded proteins (LEPs). However, their prevalence across human tissues and biological roles remain largely unexplored. The placenta is an ideal model for identifying CEPs due to its considerable protein diversity that is required to sustain fetal development during pregnancy. The aim of this study was to evaluate circRNA translation in the human placenta, and the potential roles of the CEPs in placental development and dysfunction. METHODS: Multiomics approaches, including RNA sequencing, ribosome profiling, and LC-MS/MS analysis, were utilised to identify novel translational events of circRNAs in human placentas. Bioinformatics methods and the protein bait hypothesis were employed to evaluate the roles of these newly discovered CEPs in placentation and associated disorders. The pathogenic role of a recently identified CEP circPRKCB119aa in preeclampsia was investigated through qRT-PCR, Western blotting, immunofluorescence imaging and phenotypic analyses. RESULTS: We found that 528 placental circRNAs bound to ribosomes with active translational elongation, and 139 were translated to proteins. The CEPs showed considerable structural homology with their cognate LEPs, but are more stable, hydrophobic and have a lower molecular-weight than the latter, all of which are conducive to their function as baits. On this basis, CEPs are deduced to be closely involved in placental function. Furthermore, we focused on a novel CEP circPRKCB119aa, and illuminated its pathogenic role in preeclampsia; it enhanced trophoblast autophagy by acting as a bait to inhibit phosphorylation of the cognate linear isoform PKCß. CONCLUSIONS: We discovered a hidden circRNA-encoded proteome in the human placenta, which offers new insights into the mechanisms underlying placental development, as well as placental disorders such as preeclampsia. Key points A hidden circRNA-encoded proteome in the human placenta was extensively identified and systematically characterised. The circRNA-encoded proteins (CEPs) are potentially related to placental development and associated disorders. A novel conserved CEP circPRKCB119aa enhanced trophoblast autophagy by inhibiting phosphorylation of its cognate linear-spliced isoform protein kinase C (PKC) ß in preeclampsia.

Trophoblast Exosomal UCA1 Induces Endothelial Injury through the PFN1-RhoA/ROCK Pathway in Preeclampsia: A Human-Specific Adaptive Pathogenic Mechanism.

Preeclampsia is regarded as an evolution-related disease that has only been observed in humans and our closest relatives, and the important factor contributing to its pathogenesis is endothelial dysregulation secondary to a stressed placenta. Hypoxia-inducible factor 1 subunit alpha (HIF1α), a highly conserved molecule in virtually all mammals, is regarded as a crucial regulator of the hypoxia adaptation and evolution. Persistent high expression of HIF1α in the placenta is one of the pathogenic mechanisms of preeclampsia. Therefore, human-specific molecules should link increased HIF1α to preeclampsia. We reported that urothelial cancer associated 1 (UCA1) is a potential mediator because it is a human-specific long noncoding RNA (lncRNA) that is upregulated in placental tissues and maternal serum from women with preeclampsia and is regulated by HIF1α. The cellular HIF1α-UCA1 pathway promoted the adaptation of trophoblasts to hypoxia by inducing vascular endothelial growth factor (VEGF) secretion and changes in the levels of key enzymes in glycolysis. On the other hand, circulating exosomal UCA1 secreted from stressed trophoblasts induced vascular endothelial dysfunction, especially excess ROS production, as measured by exosome extraction and a coculture system. At the molecular level, UCA1 physically bound to ubiquitin-specific peptidase 14 (USP14), which is a deubiquitinating enzyme, and UCA1 functioned as a scaffold to recruit USP14 to profilin 1 (PFN1), an actin-binding protein contributing to endothelial abnormalities and vascular diseases. This ternary complex inhibited the ubiquitination-dependent degradation of PFN1 and prolonged its half-life, further activating the RhoA/Rho-kinase (ROCK) pathway to induce ROS production in endothelial cells. Taken together, these observations suggest a role for the evolution-related UCA1 in the HIF1α-induced adaptive pathogenic mechanism of preeclampsia, promoting the survival of hypoxic trophoblasts and injuring maternal endothelial cells.

Differentially expressed circular RNAs and the competing endogenous RNA network associated with preeclampsia.

INTRODUCTION: Circular RNAs (circRNAs) are non-coding RNAs that are implicated in preeclampsia (PE) pathogenesis; however, their expression and functions in PE remain unclear. In this study, we aimed to investigate the expression of circRNAs in PE and construct a competing endogenous RNA (ceRNA) network, and analyze the associated pathways in PE pathogenesis. METHODS: We performed circRNA sequencing to identify the differential expression profile of circRNAs in PE as compared to normal pregnancy. The circRNA candidates were validated using quantitative reverse transcription polymerase chain reaction (qRT-PCR). Subsequently, we used datasets from the GEO database to generate the interaction network between circRNAs, microRNAs (miRNAs), and mRNAs. GO and KEGG enrichment analyses were performed to understand the functional significance of the differentially expressed circRNAs in PE. RESULTS: We identified 361 differentially expressed circRNAs (252 upregulated and 109 downregulated) in preeclamptic placentas. Within the selected 31 circRNAs, 6 of them were verified by qRT-PCR. GO and KEGG analyses revealed the potential pathways affected by these circRNAs, e.g., T cell receptor signaling and MAP kinase pathways. A total of 134 miRNAs and 199 mRNAs were revealed to be differentially expressed in PE by analyzing datasets from the GEO database. The circRNA-miRNA-mRNA network comprised 206 circRNAs, 50 miRNAs, and 38 mRNAs. KEGG analysis of the 38 mRNAs included pathways involved in AMPK and PI3K-Akt signaling. DISCUSSION: Our results reported the differential expression profile of circRNAs and the circRNA-miRNA-mRNA network in PE, which provides potential therapeutic targets for this disease.

Therapeutic Effects of Modified Gengnianchun Formula on Stress-Induced Diminished Ovarian Reserve Based on Experimental Approaches and Network Pharmacology.

AIM: To verify the effects of modified Gengnianchun formula (MGNC), a traditional Chinese medicine, on a stressed diminished ovarian reserve (DOR) animal model and predict the underlying mechanisms through network pharmacology strategies. METHODS: Sexually mature female C57BL/6 mice were allocated to five groups, abbreviated as the control (C) group, stress manipulated model (M) group, stress with normal saline gavage (N) group, stress with low-dose MGNC gavage (L) group, and stress with high-dose MGNC gavage (H) group. Body weight and the estrous cycle were monitored during the stress and gavage process. Serum stress hormones and reproductive hormones were evaluated by ELISA. Ovarian follicle counts were calculated, and ovarian follicle-stimulating hormone receptor (FSHR) and anti-Müllerian hormone (AMH) expression were assessed by Western blotting and immunohistochemistry. Network pharmacology strategies included active compound screening, drug and disease target analysis, gene ontology analysis, pathway analysis, and visualization of results. RESULTS: MGNC treatment significantly decreased serum corticosterone (CORT) and follicle-stimulating hormone (FSH) levels and increased testosterone (T) levels in the H group compared with the M and N groups. Primordial and preantral follicle counts and ovarian AMH and FSHR expression were significantly increased in the H group compared to those in the M and N groups. Through pharmacokinetic screening, we found 244 active compounds in MGNC. A total of 186 candidate intersection target genes of disease and MGNC were further screened to construct the interaction network. Gene ontology and KEGG pathway enrichment analysis ultimately unveiled a series of key targets that mainly mediated the effects of MGNC on DOR induced by chronic stress. The PI3K-Akt pathway may serve as the critical pathway underlying this therapeutic mechanism. CONCLUSION: MGNC is a promising formula to treat DOR induced by chronic stress, and the PI3K-Akt pathway may play an essential role in this effect.

Exposure to elevated per- and polyfluoroalkyl substances in early pregnancy is related to increased risk of gestational diabetes mellitus: A nested case-control study in Shanghai, China.

BACKGROUND: Long-chain per- and polyfluoroalkyl substances (PFASs) and their short-chain alternatives have been produced and used extensively in China. However, it is unclear whether these compounds contribute to the risk of gestational diabetes mellitus (GDM) in women residing in contaminated areas. OBJECTIVE: The study was performed to explore the association between PFASs varying in chain length and the risk of developing GDM. METHOD: A nested case-control study was conducted in a prospective cohort of 2,460 pregnant women between July 1, 2017, and January 31, 2019 in Shanghai, China. Twelve PFASs of interest were measured using ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC-Q/TOF MS) in the sera of pregnant women at 16-20 weeks. GDM was diagnosed by an oral glucose tolerance test administered over 24-28 gestational weeks. The cases and controls were matched by maternal age. The relationship between maternal serum PFAS level and GDM risk was determined by conditional logistic and linear regression analyses. RESULTS: A total of 165 GDM cases and 330 controls were enrolled in the study cohort. The frequencies of detection of PFHpA, PFDS, and PFOSA were all ≤80%. Hence, they were excluded from any further risk analysis. Perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA) were detected at relatively high serum levels (medians 6.57 ng/mL and 8.07 ng/mL, respectively). The serum levels of perfluorobutanesulfonic acid (PFBS) and perfluorododecanoic acid (PFDoA) were significantly higher in the GDM group than they were in the control group (P = 0.02 and P < 0.01, respectively) according to a nonparametric Wilcoxon rank sum test. A quartile analysis showed that the odds ratio of GDM would significantly increase at the highest PFBS and PFDoA levels. In the core model, the adjusted ORs were 2.02 (95% CI = 1.04-3.79) and 13.00 (95% CI = 4.74-24.59), respectively, after adjusting for maternal age, sampling time, parity and body mass index [BMI]). CONCLUSION: Elevated maternal serum PFBS and PFDoA levels in early pregnancy may be associated with a substantially higher GDM risk.

The Inhibition of Protein Kinase C ß Contributes to the Pathogenesis of Preeclampsia by Activating Autophagy.

BACKGROUND: Preeclampsia is a devastating hypertensive disorder of pregnancy with unknown mechanism. Recent studies have considered abnormal autophagy as a new cellular mechanism for this disorder, while little is known about how autophagy is specifically involved and what factors are implicated. Here, we report a previously unrecognized preeclampsia-associated autophagic regulator, PKCß, that is involved in placental angiogenesis. METHODS: PKCß levels were evaluated by quantitative real-time PCR, western blotting, immunofluorescence and by the analysis of public data. The autophagy-regulating role of PKCß inhibition in preeclampsia pathogenesis was studied in a mouse model, and in human umbilical vein endothelial cells (HUVECs) and human choriocarcinoma cells (JEG-3). FINDINGS: PKCß was significantly downregulated in human preeclamptic placentas. In a mouse model, the selective inhibition of PKCß by Ruboxistaurin was sufficient to induce preeclampsia-like symptoms, accompanied by excessive autophagic flux and a disruption in the balance of pro- and anti-angiogenic factors in mouse placentas. In contrast, autophagic inhibition by 3-methyladenine partially normalized hypertension, proteinuria and placental angiogenic imbalance in PKCß-inhibited mice. Our in vitro experiments demonstrated that PKCß inhibition activated autophagy, thus blocking VEGFA-induced HUVEC tube formation and resulting in the significant upregulation of sFLT1 and downregulation of VEGFA in JEG-3 cells. INTERPRETATION: These data support a novel model in which autophagic activation due to PKCß inhibition leads to the impairment of angiogenesis and eventually results in preeclampsia. FUNDING: Shanghai Key Program of Clinical Science and Technology Innovation, National Natural Science Foundation of China and Shanghai Medical Center of Key Programs for Female Reproductive Diseases.

Bisphenol A affects estradiol metabolism by targeting CYP1A1 and CYP19A1 in human placental JEG-3 cells.

Estradiol, in some way or another, plays a critically important physiologic role in the establishment and maintenance of pregnancy. This study was designed to investigate whether BPA affects the estradiol level of human placental JEG-3 cells, which may contribute to insights into the reproductive toxicity and endocrine disruption of BPA. The JEG-3 cells were treated with increasing concentrations of BPA (0.1 to 50 µM). We observed that BPA significantly reduced estradiol level of JEG-3 cells in a dose-dependent manner, which was accompanied by an increase in CYP1A1 protein level and an inhibition of CYP19A1 protein level. Additionally, by lentiviral transduction, we determined that estradiol level of JEG-3 cells over-expressing CYP1A1 gene was notably decreased and the decrease was of 84.9% compared to the control. Meanwhile, estradiol was almost undetectable in CYP19A1 knockdown group. On the contrary, the group with over-expression of CYP19A1 gene increased estradiol level by 8.6 fold while the CYP1A1 knockdown group increased by 5.6 fold. In summary, our research clearly showed that BPA alters JEG-3 estradiol synthesis and catabolism due to its action on CYP1A1 and CYP19A1 protein levels and may interfere with the normal process of placenta formation and embryonic development during early pregnancy.

Electroacupuncture Versus Pelvic Floor Muscle Training Plus Solifenacin for Women With Mixed Urinary Incontinence: A Randomized Noninferiority Trial.

OBJECTIVE: To evaluate the efficacy and safety of electroacupuncture vs pelvic floor muscle training (PFMT) plus solifenacin for women with mixed urinary incontinence (MUI). PATIENTS AND METHODS: This randomized controlled noninferiority trial was conducted at 10 hospitals in China between March 1, 2014, and October 10, 2016. Participants were randomized 1:1 to receive electroacupuncture (36 sessions) over 12 weeks with 24 weeks of follow-up or PFMT-solifenacin (5 mg/d) over 36 weeks. The primary outcome was percentage change from baseline to week 12 in mean 72-hour incontinence episode frequency (IEF) measured by the 72-hour bladder diary. It was analyzed in the per-protocol set with a prespecified noninferiority margin of 15%. RESULTS: Of 500 women with MUI who were randomized, 467 (239 in the electroacupuncture group and 228 in the PFMT-solifenacin group) completed treatment per protocol and were included in the primary outcome analysis. At weeks 1 through 12, the percentage of reduction from baseline in mean 72-hour IEF was 37.83% in the electroacupuncture group and 36.49% in the PFMT-solifenacin group (between-group difference, -1.34% [95% CI, -9.78% to 7.10%]; P<.001 for noninferiority), which demonstrates noninferiority; the treatment effect persisted throughout follow-up. Statistically significant improvements were found for secondary outcomes in both groups, with no meaningful difference between treatments. CONCLUSION: In women with moderate to severe MUI, electroacupuncture was not inferior to PFMT-solifenacin in decreasing the 72-hour IEF and shows promise as an effective alternative for the treatment of MUI. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT02047032.