27-Hydroxycholesterol inhibits trophoblast fusion during placenta development by activating PI3K/AKT/mTOR signaling pathway.

Trophoblast cell syncytialization is essential for placental and fetal development. Abnormal trophoblast cell fusion leads to pregnancy pathologies, such as preeclampsia (PE), intrauterine growth restriction (IUGR), and miscarriage. 27-hydroxycholesterol (27-OHC) is the most abundant oxysterol in human peripheral blood synthesized by sterol 27-hydroxylase (CYP27A1) and is considered a critical mediator between hypercholesterolemia and a variety of related disorders. Gestational hypercholesterolemia was associated with spontaneous preterm delivery and low birth weight (LBW) in term infants, yet the mechanism is unclear. In this study, two trophoblast cell models and CD-1 mice were used to evaluate the effects of 27-OHC on trophoblast fusion during placenta development. Two different kinds of trophoblast cells received a dosage of 2.5, 5, or 10 uM 27-OHC. Three groups of pregnant mice were randomly assigned: control, full treatment (E0.5-E17.5), or late treatment (E13.5-E17.5). All mice received daily intraperitoneal injections of saline (control group) and 27-OHC (treatment group; 5.5 mg/kg). In vitro experiments, we found that 27-OHC inhibited trophoblast cell fusion in primary human trophoblasts (PHT) and forskolin (FSK)-induced BeWo cells. 27-OHC up-regulated the expression of the PI3K/AKT/mTOR signaling pathway-related proteins. Moreover, the PI3K inhibitor LY294002 rescued the inhibitory effect of 27-OHC. Inhibition of trophoblast cell fusion by 27-OHC was also observed in CD-1 mice. Furthermore, fetal weight and placental efficiency decreased and fetal blood vessel development was inhibited in pregnant mice treated with 27-OHC. This study was the first to prove that 27-OHC inhibits trophoblast cell fusion by Activating PI3K/AKT/mTOR signaling pathway. This study reveals a novel mechanism by which dyslipidemia during pregnancy results in adverse pregnancy outcomes.

Nuclear Binding Protein 2/Nesfatin-1 Affects Trophoblast Cell Fusion during Placental Development via the EGFR-PLCG1-CAMK4 Pathway.

Previous studies have shown that nuclear binding protein 2 (NUCB2) is expressed in the human placenta and increases with an increase in the syncytialization of trophoblast cells. This study aimed to investigate the role of NUCB2 in the differentiation and fusion of trophectoderm cells. In this study, the expression levels of NUCB2 and E-cadherin in the placentas of rats at different gestation stages were investigated. The results showed that there was an opposite trend between the expression of placental NUCB2 and E-cadherin in rat placentas in different trimesters. When primary human trophoblast (PHT) and BeWo cells were treated with high concentrations of Nesfatin-1, the trophoblast cell syncytialization was significantly inhibited. The effects of NUCB2 knockdown in BeWo cells and Forskolin-induced syncytialization were investigated. These cells showed a significantly decreased cell fusion rate. The mechanism underlying NUCB2-regulated trophoblast cell syncytialization was explored using RNA-Seq and the results indicated that the epidermal growth factor receptor (EGFR)-phospholipase C gamma 1 (PLCG1)-calmodulin-dependent protein kinase IV (CAMK4) pathway might be involved. The results suggested that the placental expression of NUCB2 plays an important role in the fusion of trophoblasts during differentiation via the EGFR-PLCG1-CAMK4 pathway.

Association of maternal HDL2-c concentration in the first trimester and the risk of large for gestational age birth.

BACKGROUND: Maternal lipid levels during pregnancy are critical for fetal development. Recent studies revealed that high-density lipoprotein cholesterol (HDL-c) levels during pregnancy were negatively correlated with birthweight. High-density lipoprotein 2 cholesterol (HDL2-c) is one of the major subclasses of HDL-c, and its relationship with birthweight is unclear. Association of HDL2-c concentration in the first trimester and risk of large for gestational age (LGA) was explored. METHODS: This study recruited pregnant women who registered in Fuxing Hospital from October 2018 to January 2020, had regular obstetric examinations during pregnancy, and delivered between June 2019 and September 2020. Finally, 549 participants were recruited for the study. Maternal demographic characteristics and venous blood were collected at the 6th-14th gestational week, and serum total cholesterol (TC), triglyceride (TG), HDL-c, HDL2-c, high-density lipoprotein 3 cholesterol (HDL3-c), and low-density lipoprotein cholesterol (LDL-c) concentrations were detected. Neonatal characteristics were collected at delivery. A logistic regression model was used to explore the relationship between the first trimester HDL2-c concentration and LGA incidence. A nomogram was developed, and the performance was evaluated with a concordance index. RESULTS: Seventy-five mothers delivered LGA infants, and the LGA incidence was 13.66%. LGA mothers had significantly lower serum HDL-c and HDL2-c concentrations than appropriate for gestational age (AGA) mothers. A logistic regression model showed that HDL2-c concentration was negatively correlated with LGA risk (odds ratio (OR) = 0.237, 95% confidence intervals (CI): 0.099-0.567, P = 0.001) when adjusted for age, prepregnancy body mass index (BMI), and parity. A nomogram was generated using all these risk factors. The area under the curve (AUC) was 0.663 (95% CI: 0.593-0.732). CONCLUSIONS: Maternal HDL2-c concentration in the first trimester was negatively correlated with the risk of LGA.

Changes in serum TG levels during pregnancy and their association with postpartum hypertriglyceridemia: a population-based prospective cohort study.

BACKGROUND: Blood lipid increases during gestation are considered a physiological adaption, and decrease after delivery. However, some adverse pregnancy outcomes are thought to be related to gestational lipid levels. Therefore, it is necessary to have a reference range for lipid changes during gestation. The present study aims to describe triglyceride (TG) changes during pregnancy and 42 days postpartum and to find cut-off points for TG levels during the first, second, and third trimesters. METHODS: A total of 908 pregnant women were followed from recruitment to 42 days postpartum, and their serum lipids were collected at gestational weeks 6-8, 16, 24, and 36 and 42 days postpartum. The major outcome was postpartum hypertriglyceridemia. The association between gestational and postpartum TG levels was analysed by stepwise multiple linear regression. A two-stage approach including a linear mixed-effect model and linear or logistic regression was conducted to explore the contribution of the changes in TG over time in pregnancy to postpartum hypertriglyceridemia. Logistic regression was constructed to examine the association between gestational TG levels and postpartum hypertriglyceridemia. Cut-off points were calculated by receiver operating characteristic (ROC) curves. RESULTS: There was a tendency for serum TG to increase with gestational age and decrease at 42 days postpartum. Prepregnancy overweight, obesity, and GDM intensified this elevation. Higher TG levels at gestational weeks 6-8, 16, 24, and 36 were positively associated with a higher risk of postpartum hypertriglyceridemia [OR 4.962, 95 % CI (3.007-8.189); OR 2.076, 95 % CI (1.303-3.309); OR 1.563, 95 % CI (1.092-2.236); and OR 1.534, 95 % CI (1.208-1.946), respectively]. The trend of the change in TG over time was positively associated with the TG level and risk of postpartum hypertriglyceridemia [OR 11.660, 95 % CI (6.018-22.591)]. Based on ROC curves, the cut-off points of serum TG levels were 1.93, 2.35, and 3.08 mmol/L at gestational weeks 16, 24, and 36, respectively. Stratified analysis of prepregnancy body mass index (pre-BMI) and GDM showed that higher gestational TG was a risk factor for postpartum hypertriglyceridemia in women with normal pre-BMI and without GDM. CONCLUSIONS: Gestational TG and its elevation were risk and predictive factors of postpartum hypertriglyceridemia, especially in pregnant women with normal pre-BMI or without GDM.

Associations between maternal serum HDL-c concentrations during pregnancy and neonatal birth weight: a population-based cohort study.

BACKGROUND: To evaluate the associations between maternal serum concentrations of high-density lipoprotein cholesterol (HDL-c) throughout pregnancy and neonatal birth weight (BW) and small for gestational age (SGA) births. METHODS: A prospective cohort of 2241 pregnant women was followed from recruitment to delivery in three hospitals in Beijing, China between January 2014 and December 2017. Maternal fasting serum lipids concentrations were measured at gestational week 6-12, 16, 24 and 36. Major outcome was neonatal BW. The associations between maternal HDL-c and BW were estimated by linear regression and linear mixed-effects models. Odds ratios (ORs) and 95% confidence intervals of SGA births in relation to HDL-c were evaluated via logistic regression analysis. RESULTS: There was a tendency that mothers with higher HDL-c concentrations throughout gestation gave birth to infants with lower BW. A negative association was found between maternal HDL-c concentrations and BW at 24th and 36th gestational weeks (B = - 34.044, P = 0.034; B = - 53.528, P = 0.000). The HDL-c trend of change was inversely associated with BW (B = - 442.736, P = 0.000). Mothers with SGA neonates had higher serum HDL-c concentration at the 36th gestational week (P < 0.01). The incidences of SGA in the three groups (HDL-c: 1.84-2.23 mmol/L, 2.24-2.59 mmol/L and ≥ 2.60 mmol/L) were higher than the group with the lowest concentration of HDL-c (< 1.83 mmol/L) (P < 0.01, P < 0.01, P < 0.001) at 36th week. Higher maternal HDL-c concentrations at 36th week (HDL-c: 1.84-2.23 mmol/L, 2.24-2.59 mmol/L and ≥ 2.60 mmol/L) were positively associated with the incidence of SGA (OR = 1.900, P = 0.008; OR = 1.893, P = 0.008; OR = 1.975, P = 0.004). The HDL-c trend of change was positively associated with SGA births (OR = 9.772, P = 0.000). CONCLUSIONS: Maternal serum HDL-c concentrations were inversely associated with BW at 24th and 36th gestational weeks. The high concentrations of HDL-c at the 36th gestational week increased the risk of SGA. The maternal HDL-c trend of change across pregnancy was associated with smaller neonatal size.

Elevated serum cholesterol levels during pregnancy as predictors for postpartum hypercholesterolemia: A prospective cohort study.

OBJECTIVE: To identify the associations between gestational cholesterol levels and the risk of postpartum hypercholesterolemia, and to establish trimester-specific reference values. METHODS: Serum lipids at gestational weeks 6-8, 16, 24, and 36, and 42 days postpartum were derived from 905 pregnant women of a prospective cohort. The major outcome was postpartum hypercholesterolemia. Logistic regression and restricted cubic splines were conducted to estimate the associations between cholesterol levels at specific gestational ages and postpartum hypercholesterolemia. Associations of the trend of changes in cholesterol levels during pregnancy with postpartum hypercholesterolemia were evaluated by linear mixed-effect model and linear or logistic regression. Reference values were computed by the receiver operating characteristic curves. RESULTS: Serum concentrations of total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), and the ratios of TC/HDL-C and LDL-C/HDL-C all increased during pregnancy and decreased at 42 days postpartum. The elevated levels of TC and LDL-C, decreased levels of HDL-C in pregnancy, and their rapid change rates were positively associated with higher risks of postpartum hypercholesterolemia. The established reference values from the first to the third trimester were below 5.47, 6.35, and 7.22 mmol/L for TC; below 2.83, 3.82, and 4.21 mmol/L for LDL-C; and more than 1.50, 1.55, and 1.50 mmol/L for HDL-C, respectively. CONCLUSION: Maternal cholesterol levels and their trend of change during pregnancy were predictors of postpartum hypercholesterolemia. Trimester-specific reference values were established in a Chinese population.

Reliability and Validity of a Theory-Based Determinants of Eating and Physical Activity Behaviors Questionnaire for Chinese Elementary School Children.

OBJECTIVE: The primary objective of this study was to develop and validate a Social Cognitive Theory-based instrument to identify psychosocial factors that influence diet and physical activity among Chinese children aged 10-12 years. DESIGN: This is a cross-sectional study, with data collected from questionnaires. SETTING: Two elementary schools in Beijing, China. PARTICIPANTS: Fourth to sixth-grade students (N = 1,486) aged 10-12 years were recruited. VARIABLES MEASURED: Gender, height, weight, nation, and grade were collected. Energy-balanced eating behaviors and their related sociopsychological factors were surveyed. ANALYSIS: Confirmatory factor analysis, Pearson correlations, Cronbach α index, and mediation analysis were used. RESULTS: (1) Confirmatory factor analysis revealed a 6-factor solution (51 items) and all factor loadings > 0.32, indicating that the model fitness was acceptable. (2) All correlation coefficients are statistically significant. All of the Cronbach α indexes were > 0.65, indicating acceptable reliability. (3) The mediating effect of goal intention and outcome expectations between self-efficacy and habit strength was statistically significant (P < 0.01), verifying the theory structure. CONCLUSIONS AND IMPLICATIONS: This questionnaire exhibits good internal consistency, reliability, and structural validity. It can be effectively employed to investigate energy-balanced eating behaviors related to the Social Cognitive Theory in Chinese children.

Dosage-effect of selenium supplementation on blood glucose and oxidative stress in type 2 diabetes mellitus and normal mice.

BACKGROUND: The effectiveness of selenium (Se) supplementation on glycemic control is disparate. OBJECTIVE: This study aims to evaluate the effects of different dosages of Se diets on the blood glucose in type 2 diabetes mellitus (T2DM, db/db) and normal (db/m) mice. METHODS: The db/db and db/m mice were fed with different dosages of Se supplemented diets (0, 0.1, 0.3, 0.9, 2.7 mg/kg) for 12 weeks, respectively. Se concentrations of tissues, physical and biochemical characteristics, oxidative stress indexes and gene expression related to glucose, lipid metabolism and Se transporters of liver were detected. RESULTS: The Se concentrations in tissues were related to the dosages of Se supplementation in db/db (blood: slope=11.69, r = 0.924; skeletal muscle: slope=0.36, r = 0.505; liver: slope=22.12, r = 0.828; kidney: slope=11.81, r = 0.736) and db/m mice (blood: slope=19.89, r = 0.876; skeletal muscle: slope=2.80, r = 0.883; liver: slope=44.75, r = 0.717; kidney: slope=60.15, r = 0.960). Compared with Se2.7 group, the fasting blood glucose (FBG) levels of Se0.1 and Se0.3 group were decreased at week3 in db/db mice. Compared with control (Se0) group, the FBG levels of Se2.7 group were increased from week6 to week12 in db/m mice. The oral glucose tolerance test (OGTT) showed that the area under the curve (AUC) of Se0.3 group was lower than that of Se0.9 and Se2.7 group in db/m mice. Furthermore, compared with control group, the malondialdehyde (MDA) level in skeletal muscle of Se0.1 group was decreased, while that of Se2.7 group was increased in db/db mice; the glutathione peroxidase (GPx) activity in skeletal muscle of Se0.3, Se0.9 and Se2.7 group was increased both in db/db and db/m mice. For db/db mice, glucose-6-phosphatase catalytic (G6pc) expression of other groups were lower and fatty acid synthase (Fasn) expression of Se0.9 group were lower compared with Se0.3 group. For db/m mice, compared with Se0.3 group, (peroxisome proliferative activated receptor gamma coactivator 1 alpha) Pgc-1α expression of control and Se0.9 group were higher; (phosphoenolpyruvate carboxykinase 1) Pck1 expression of Se0.1, Se0.9, and Se2.7 group were higher. CONCLUSION: Low dosages (0.1 and 0.3 mg/kg) of Se supplementation exerted beneficial effects on FBG levels and glucose tolerance through regulating hepatic glycolysis and gluconeogenesis and inhibit the oxidative stress while high dosages of Se (0.9 and 2.7 mg/kg) supplementation enhanced FBG levels, impaired glucose tolerance and aggravate oxidative stress.

Oat ß-glucan supplementation pre- and during pregnancy alleviates fetal intestinal immunity development damaged by gestational diabetes in rats.

Oat ß-glucan (OG) has been shown to improve intestinal microecology in gestational diabetes mellitus (GDM), but the effect on fetal intestine health is unknown. Herein, we aimed to investigate the effects of OG supplementation during gestation in GDM dams on fetal intestinal immune development. OG was supplemented one week before mating until the end of the experiment. GDM rats were made with a high-fat diet (HFD) with a minimal streptozotocin (STZ) dose. The fetal intestines were sampled at gestation day (GD) 19.5, and the intestinal morphology, chemical barrier molecules, intraepithelial immune cell makers, and levels of inflammatory cytokines were investigated. The results showed that OG supplementation alleviated the decrease of the depth of fetal intestinal villi and crypts, the number of goblet cells (GCs), protein expression of mucin-1 (Muc1) and Muc2, the mRNA levels of Gpr41, Gpr43, and T cell markers, and increased the number of paneth cells (PCs), the mRNA levels of defensin-6 (defa6), and macrophage (Mø) marker and the expression of cytokines induced by GDM. In addition, OG supplementation alleviated the function of immune cell self-proliferation, chemotaxis and assembly capabilities, protein, fat, folic acid, and zinc absorption damaged by GDM. As indicated by these findings, OG supplementation before and during pregnancy improved the fetal intestinal chemical barriers, immune cells, cytokines, and the metabolism of nutrients to protect the fetal intestinal immunity.

Lipid Metabolic Genes and Maternal Supraphysiological Hypercholesterolemia: An Analysis of Maternal-fetal Interaction.

CONTEXT: The joint associations of maternal and fetal single nucleotide polymorphisms (SNPs) of lipid metabolic genes with the risk of maternal supraphysiological hypercholesterolemia (MSPH) are unclear. OBJECTIVE: This study aims to investigate the associations of maternal/fetal SNPs of APOE, LPL, LDLR, PCSK9, and SCARB1 with the risk of MSPH and explore whether the maternal-fetal pairing pattern of the risk alleles can affect MSPH risk. METHODS: A nested case-control study was conducted that included 182 pregnant women with MSPH and 182 with maternal physiological hypercholesterolemia. Maternal venous and umbilical venous blood were collected to detect the SNPs of genes. The primary outcome was MSPH. Logistic regression model was used to determine the associations of SNPs with risk of MSPH. RESULTS: The C-allele in maternal APOE rs429358 T > C (adjusted odds ratio [OR] = 1.72, P = 0.033), G-allele in fetal APOE rs440446 C > G (adjusted OR = 1.62, P = 0.012) and T-allele in fetal LPL rs263 C > T (adjusted OR = 1.53, P = 0.011) increased the risk of MSPH. The A-allele in maternal LDLR rs7258950 G > A decreased the risk of MSPH (adjusted OR = 0.67, P = 0.028). For maternal-fetal pairing analysis, the variant concordance of PCSK9 rs2149041, rs7523141, rs7523242, rs7525649, and LDLR rs7258950 were associated with the decreased risk of MSPH under the dominant model. The variant concordance of other SNPs of PCSK9, APOE, LDLR, LPL, and SCARB1 were associated with the increased risk of MSPH. CONCLUSION: This study supports the hypothesis that maternal and fetal genetic polymorphisms of lipid metabolic genes are associated with the risk of MSPH. The maternal-fetal variant concordance is also associated with this risk.

Nobiletin Inhibits Hypoxia-Induced Placental Damage via Modulating P53 Signaling Pathway.

In this study, we aimed to evaluate the effect of Nobiletin (NOB) on the placenta of Sprague-Dawley (SD) rats that had undergone reduced uterine perfusion pressure (RUPP) surgery and to evaluate the safety of NOB intervention during pregnancy. The results showed that NOB alleviated placental hypoxia, attenuated placental cell apoptosis, and inhibited placental damage in RUPP rats. No side effect of NOB intervention during pregnancy was observed. BeWo cell lines with P53 knockdown were then constructed using lentiviral transfection, and the P53 signaling pathway was found to be essential for NOB to reduce hypoxia-induced apoptosis of the BeWo cell lines. In summary, NOB attenuated hypoxia-induced placental damage by regulating the P53 signaling pathway, and those findings may contribute some insights into the role of NOB in placental development and the prevention of placental-related diseases.

Vitamin D3 alleviates cognitive impairment through regulating inflammatory stress in db/db mice.

Patients with type 2 diabetes mellitus (T2DM) have a higher risk to develop cognitive impairment. Several studies reported the potential roles of vitamin D in prevention of cognitive impairment, but the mechanism remains unclear. The present study aims to investigate the protective effects of vitamin D3 on cognitive impairment in db/db mice and to explore the possible mechanism. Twelve-week-old male db/db mice were randomly administrated with low, medium, and high dose of vitamin D3 (LVD, MVD, and HVD groups, respectively) and equivalent volume vitamin D3 solvent (corn oil, DM group) intragastrically. Eight age-matched db/m mice were given equivalent volume corn oil as normal group. After 16 weeks of vitamin D3 treatment, the concentrations of fasting serum glucose in three vitamin D3 groups (especially the 1,000 IU/kg·bw dose) were significantly decreased compared with DM group. Pathology revealed that the neuron damage was reduced in vitamin D3 groups. MVD intervention significantly shortened the escape latency on day 5 and extended time in the target quadrant. Mice in HVD group had significantly higher exploration time and discrimination index compared with the DM group mice. Moreover, vitamin D3 treatment has increased the phosphorylation of cAMP-response element-binding protein and the expression of brain-derived neurotrophic factor and vitamin D receptor. This treatment, meanwhile, has decreased the expression of tumor necrosis factor-α, the phosphorylation of inhibitor kappa Bα (IκBα), and nuclear factor-κB p65 (NF-κB p65) in the hippocampus of db/db mice. These results suggest that vitamin D3 alleviated cognitive impairment in the hippocampus of db/db mice. Down-regulation of the NF-κB signaling pathway-related proteins IκBα and p65 might be one of the possible mechanisms.

The effects of high-density lipoprotein and oxidized high-density lipoprotein on forskolin-induced syncytialization of BeWo cells.

INTRODUCTION: The negative relationship between maternal high-density lipoprotein-cholesterol (HDL-c) level during pregnancy and infant birth weight has been found. Syncytialization (differentiation and fusion) of trophoblast cells is important to fetal development. HDL has an antioxidant effect, and has been proved to protect trophoblast functions including hormone secretion and invasion. However, HDL is susceptible to oxidation, and high concentrations of HDL impair cell growth and oxidized HDL (oxHDL) inhibits cell proliferation and migration. Moreover, the effects of HDL and oxHDL on trophoblast syncytialization have not been characterized. The aim of this study was to investigate the effects of HDL and oxHDL on trophoblast syncytialization. METHODS: Human choriocarcinoma trophoblasts (BeWo cells) were treated with human HDL or oxHDL and then induced to differentiate by forskolin in syncytialization assays. Expression levels of mRNAs and proteins regulating syncytialization were detected by real-time PCR and western blotting, respectively. RESULTS: Treatments of HDL at high concentrations reduced human chorionic gonadotropin (hCG) secretion, placental alkaline phosphatase activity and fusion rates, and decreased the expressions of GCM1 and ERVW-1 mRNA as well as phospho-MAPK1/3 (p-MAPK1/3) and total MAPK1/3 protein in the forskolin-induced syncytialization of BeWo cells. Furthermore, treatment of oxHDL (20 µg/ml) decreased hCG secretion, but increased the expression of p-MAPK1/3 protein. DISCUSSION: These data suggested that both HDL at high concentrations and oxHDL inhibited BeWo cells syncytialization, and might be harmful to placental and fetal development.