Targeted bisulfite resequencing of differentially methylated cytosines in pre-eclampsia reveals a skewed dynamic balance in the DNA methylation of enhancers.

Pre-eclampsia (PE) is a major hypertensive disorder of pregnancy. Widespread differentially methylated cytosines (DMCs) with modest changes in methylation level are associated with PE, whereas their cause and biological significance remain unknown. We aimed to clarify DNA methylation patterns around DMCs in 103 placentas using MethylCap targeted bisulfite re-sequencing (MethylCap-seq) assays of 690 selected DMCs. We verified the MethylCap-seq method, then validated 677 (98.1%) of DMCs (vDMCs) in an independent cohort. The validated DMCs were strongly enriched in active placenta-specific enhancers and showed highly dynamic methylation levels. We found high epigenetic heterogeneity between vDMCs and adjacent CpG sites (r2 < 0.2) and a significant decrease in PE in the discovery and replication cohorts (P = 2.00 × 10-24 and 6.43 × 10-9, respectively). We replicated the methylation changes in a hypoxia/reoxygenation cell model. We constructed 112 methylation haplotype blocks and found that the frequencies of unmethylated haplotypes (UMHs) were dynamic with gestational age (GA) and were altered in maternal plasma of patients with PE. Our results uncovered additional DNA methylation features in PE placentas and suggested a model of skewed DNA methylation balance of enhancers in PE.

Comparison of immune defense and antioxidant capacity between broodstock and hybrid offspring of juvenile shrimp (Macrobrachium nipponense): Response to acute ammonia stress.

Ammonia is a major environmental pollutant in the aquatic system that poses a great threat to the health of shrimp. Macrobrachium nipponense, as one of the large-yield farmed shrimp, is facing germplasm degradation. Genetic improvement through hybridization is one of the effective methods to solve this problem. However, there are few studies on the effects of ammonia nitrogen on the germplasm resources of M. nipponense. In this study, the broodstock populations (Dianshan, DS) and hybrid offspring (DS ♀ × CD [Changjiang, CJ ♂ × Dongting, DT ♀], SCD) were exposed to 0, 5, or 20 mg/L of ammonia for 96 h. The survival rate of the SCD group was greater than the DS group, although there were no significant differences in weight gain rate and length gain rate (p > 0.05). The number of positive cells and apoptosis rates in the DS group were significantly greater than in the SCD group after ammonia exposure (p < 0.05). As the ammonia concentration increased, the antioxidant enzyme activities in the SCD group were significantly higher than DS group, while the hepatotoxicity enzyme activities in the SCD group were significantly lower than DS group (p < 0.05). The trends in the expression of antioxidant- and immune-related genes were generally consistent with the activities of antioxidant enzymes. Our study found that the hybrid population had stronger stress resistance than their parent populations at the same ammonia concentration. This study confirms our speculation that hybrid population has a greater advantage in antioxidant immunity, which also provides reference for the follow-up study of chronic ammonia toxicity.

Transcriptome sequencing of adenomyosis eutopic endometrium: A new insight into its pathophysiology.

The eutopic endometrium has been suggested to play a crucial role in the pathogenesis of adenomyosis. However, the specific genes in eutopic endometrium responsible for the pathogenesis of adenomyosis still remain to be elucidated. We aim to identify differentially expressed genes (DEGs) and molecular pathways/networks in eutopic endometrium from adenomyosis patients and provide a new insight into disease mechanisms at transcriptome level. RNA sequencing (RNA-Seq) was performed with 12 eutopic endometrium from adenomyosis and control groups. Differentially expressed genes in adenomyosis were validated by quantitative real-time PCR (qPCR) and immunochemistry. Functional annotations of the DEGs were analysed with Ingenuity Pathway Analysis (IPA). Quantitative DNA methylation analysis of CEBPB was performed with MassArray system. A total of 373 differentially expressed genes were identified in the adenomyosis eutopic endometrium compared to matched controls. Bioinformatic analysis predicted that IL-6 signalling and ERK/MAPK signalling were activated in adenomyosis endometrium. We also found that the increased expression and DNA hypomethylation of CEBPB were associated with adenomyosis. Our results revealed key pathways and networks in eutopic endometrium of adenomyosis. The study is the first to propose the association between C/EBPß and adenomyosis and can improve the understanding of the pathogenesis of adenomyosis.

Integrated Transcriptome Sequencing Analysis Reveals Role of miR-138-5p/ TBL1X in Placenta from Gestational Diabetes Mellitus.

BACKGROUND/AIMS: The placenta has been suggested to play a crucial role in the pathology of gestational diabetes mellitus (GDM). Placenta-specific microRNAs (miRNAs) and the corresponding targeting genes involved in the pathology of GDM still remain to be elucidated. We aimed to identify the dysregulated miRNAs and the corresponding mRNA targets through an integrated miRNA and mRNA transcriptomic profiles analysis and investigate the role of differentially expressed miR-138-5p/TBL1X in GDM. METHODS: RNA sequencing (RNA-seq) was performed in 16 placentas from GDM and control group. Differentially expressed mRNAs and miRNAs in GDM were validated by quantitative PCR (qPCR). The wound healing assay and transwell migration assay were used to analyze cell migration ability. The cell proliferation was determined by CCK8 assay. Luciferase assay was used to confirm the direct binding of the targeted TBL1X with miR-138-5p. RESULTS: Totally, 281 mRNAs and 32 miRNAs were found to be differentially expressed in the GDM placentas. The biological relationships of the miRNA/mRNA pairs were related to cellular development and function and organ morphology. Among the aberrantly expressed molecules, we selected miR-138-5p from the bioinformatics analysis and found that miR-138-5p significantly inhibited the migration and proliferation of trophoblasts (HTR-8/SVneo) by targeting the 3'-UTR of TBL1X. Furthermore, the aberrant expression of miR-138-5p and TBL1X was significantly correlated with the weight of the placenta. CONCLUSION: We present the first integrative analysis of miRNA and mRNA expression profiles in GDM placenta and uncover a more detailed role for miR-138-5p, as well as its target TBL1X in the pathology of GDM.

The DNA methylation status of genes encoding Matrix metalloproteinases and tissue inhibitors of Matrix metalloproteinases in endometriosis.

Endometriosis is a benign disease, with malignant properties. A necessary step in the progression of endometriosis is tissue remodeling, which is coordinated by the activities of matrix metalloproteinases (MMPs) and tissue inhibitors of matrix metalloproteinases (TIMPs). This study evaluated the regulation of abnormal MMP and TIMP gene expression during endometriosis. Among the two genes families, promoter regions of MMP2, MMP3, MMP7, TIMP3, and TIMP4 were significantly altered in proliferative-phase endometriotic lesions compared to menstrual cycle-matched eutopic tissue from endometriosis-free women. In addition, a negative correlation was found between the DNA methylation status of the promoter region and transcript abundance of MMP2. Our findings suggest that changes in DNA methylation at the promoter region of MMP2 could underlie the changes in its expression in the ectopic endometria from patients with endometriosis.

An adapted carrier for the cryopreservation of human testicular spermatozoa.

RESEARCH QUESTION: Does the adapted carrier Cryoplus improve the quality of cryopreserved spermatozoa compared with the use of conventional containers, and what is the effect of the adapted carrier on clinical outcomes? DESIGN: Semen samples from 27 cases of oligozoospermia were used to investigate whether the adapted carrier improved cryopreserved sperm quality compared with the use of 0.25-ml straws and 2-ml cryogenic vials. Thirty testicular sperm samples were used to study the quality of testicular spermatozoa cryopreserved in the adapted carrier. The retrospective study included a further 104 men with azoospermia to investigate the clinical outcomes of testicular spermatozoa cryopreserved with the adapted carriers. Men with mostly obstructive azoospermia were included in this study. RESULTS: The adapted carrier improved cryopreserved spermatozoa motility of semen samples compared with 2-ml cryogenic vials but not compared with 0.25-ml straws. No differences were found in cryopreserved sperm DNA fragmentation among the three carriers. Fertilization and good-quality embryo rates were similar in ICSI cycles using fresh or cryopreserved testicular spermatozoa. Additionally, no difference was evident between frozen-thawed embryo transfer cycles using fresh or cryopreserved testicular spermatozoa in clinical pregnancy, implantation, miscarriage, live birth rates or birth weight. CONCLUSIONS: The adapted carrier improved the cryopreserved sperm motility compared with the effects of 2-ml cryogenic vials. The outcomes of intracytoplasmic sperm injection and frozen-thawed embryo transfer outcomes indicate that testicular spermatozoa cryopreserved using the adapted carrier is not inferior to fresh testicular spermatozoa. The use of the adapted carrier for cryopreserving human testicular spermatozoa especially from obstructive azoospermia is simple and effective.

Opioid Exposure is Associated with Aberrant DNA Methylation of OPRM1 Promoter Region in a Chinese Han Population.

The µ-opioid receptor (OPRM1) plays an important role in opiate addiction. The OPRM1 gene promoter showed hypermethylation in lymphocytes of opiate addicts as well as opioid medications users, while the methylation status displayed ethnic diversity. The purpose of the study was to investigate the methylation pattern of OPRM1 promoter in the Han Chinese population. We analyzed 22 CpG sites located in OPRM1 promoter in 186 former opiate addicts (94 males and 92 females) and 184 healthy controls (102 males and 82 females). The + 126 CpG site was significantly hypermethylated in the former heroin addicts compared with controls (13.67% versus 8.39%, [Formula: see text], corrected for 36 tests). Six CpG sites were significantly associated with opioid exposure, including the most significant +126 CpG site (opiate addicts 13.57%, control 8.39%, [Formula: see text], corrected for 36 tests), while the +23 GpG site was the only hypomethylated one in former opiate addicts compared with controls (P = 0.0023 after Bonferroni correction). Our results supported that opioid exposure was associated with methylation status of OPRM1 promoter and showed ethnic dependence.

Loci with genome-wide associations with schizophrenia in the Han Chinese population.

BACKGROUND: A large schizophrenia genome-wide association study (GWAS) and a subsequent extensive replication study of individuals of European ancestry identified eight new loci with genome-wide significance and suggested that the MIR137-mediated pathway plays a role in the predisposition for schizophrenia. AIMS: To validate the above findings in a Han Chinese population. METHOD: We analysed the single nucleotide polymorphisms (SNPs) in the newly identified schizophrenia candidate loci and predicted MIR137 target genes based on our published Han Chinese populations (BIOX) GWAS data. We then analysed 18 SNPs from the candidate regions in an independent cohort that consisted of 3585 patients with schizophrenia and 5496 controls of Han Chinese ancestry. RESULTS: We replicated the associations of five markers (P<0.05), including three that were located in the predicted MIR137 target genes. Two loci (ITIH3/4: rs2239547, P = 1.17 × 10(-10) and CALN1: rs2944829, P = 9.97 × 10(-9)) exhibited genome-wide significance in the Han Chinese population. CONCLUSIONS: The ITIH3/4 locus has been reported to be of genome-wide significance in the European population. The successful replication of this finding in a different ethnic group provides stronger evidence for the association between schizophrenia and ITIH3/4. We detected the first genome-wide significant association of schizophrenia with CALN1, which is a predicted target of MIR137, and thus provide new evidence for the associations between MIR137 targets and schizophrenia.

DNA methylome profiling of maternal peripheral blood and placentas reveal potential fetal DNA markers for non-invasive prenatal testing.

Utilizing epigenetic (DNA methylation) differences to differentiate between maternal peripheral blood (PBL) and fetal (placental) DNA has been a promising strategy for non-invasive prenatal testing (NIPT). However, the differentially methylated regions (DMRs) have yet to be fully ascertained. In the present study, we performed genome-wide comparative methylome analysis between maternal PBL and placental DNA from pregnancies of first trimester by methylated DNA immunoprecipitation-sequencing (MeDIP-Seq) and Infinium HumanMethylation450 BeadChip assays. A total of 36 931 DMRs and 45 804 differentially methylated sites (DMSs) covering the whole genome, exclusive of the Y chromosome, were identified via MeDIP-Seq and Infinium 450k array, respectively, of which 3759 sites in 2188 regions were confirmed by both methods. Not only did we find the previously reported potential fetal DNA markers in our identified DMRs/DMSs but also we verified fully the identified DMRs/DMSs in the validation round by MassARRAY EpiTYPER. The screened potential fetal DNA markers may be used for NIPT on aneuploidies and other chromosomal diseases, such as cri du chat syndrome and velo-cardio-facial syndrome. In addition, these potential markers may have application in the early diagnosis of placental dysfunction, such as pre-eclampsia.

Prenatal nutritional deficiency reprogrammed postnatal gene expression in mammal brains: implications for schizophrenia.

BACKGROUND: Epidemiological studies have identified prenatal exposure to famine as a risk factor for schizophrenia, and animal models of prenatal malnutrition display structural and functional brain abnormalities implicated in schizophrenia. METHODS: The offspring of the RLP50 rat, a recently developed animal model of prenatal famine malnutrition exposure, was used to investigate the changes of gene expression and epigenetic modifications in the brain regions. Microarray gene expression analysis was carried out in the prefrontal cortex and the hippocampus from 8 RLP50 offspring rats and 8 controls. MBD-seq was used to test the changes in DNA methylation in hippocampus depending on prenatal malnutrition exposure. RESULTS: In the prefrontal cortex, offspring of RLP50 exhibit differences in neurotransmitters and olfactory-associated gene expression. In the hippocampus, the differentially-expressed genes are related to synaptic function and transcription regulation. DNA methylome profiling of the hippocampus also shows widespread but systematic epigenetic changes; in most cases (87%) this involves hypermethylation. Remarkably, genes encoded for the plasma membrane are significantly enriched for changes in both gene expression and DNA methylome profiling screens (p = 2.37×10(-9) and 5.36×10(-9), respectively). Interestingly, Mecp2 and Slc2a1, two genes associated with cognitive impairment, show significant down-regulation, and Slc2a1 is hypermethylated in the hippocampus of the RLP50 offspring. CONCLUSIONS: Collectively, our results indicate that prenatal exposure to malnutrition leads to the reprogramming of postnatal brain gene expression and that the epigenetic modifications contribute to the reprogramming. The process may impair learning and memory ability and result in higher susceptibility to schizophrenia.

Low flow-resistance solid phase extraction of fluoroquinolones in water and food samples by high-pressure wet spinning porous polyimide microfibers.

In this work, porous polyimide microfibers (PI-µF) were prepared by high-pressure wet spinning method, and successfully applied as adsorbents for solid phase extraction (SPE) of fluoroquinolones (FQs) in water and food samples. The PI-µFs of ∼10, 25, 50, 100 µm in diameter could be controlled by the inner diameter of quartz capillary nozzles. The flow resistance of SPE cartridges packed with 10 µm PI microfiber (10-PI-µF) and 25-PI-µF was comparable to or even lower than that of commercial SPE cartridges, while the flow resistance of 50-PI-µF and 100-PI-µF SPE cartridges was increased obviously due to tiny broken pieces. The 10-PI-µF and 25-PI-µF have a specific surface area of 102 m2 g-1 and 76 m2 g-1, mesopores of 22-32 nm, and large breakthrough volume of 110 mL/5 mg and 85 mL/5 mg for FQs, while the 50-PI-µF and 100-PI-µF had much lower specific surface area and hardly had retention for FQs. FQs from tap water, egg and milk samples were then extracted by PI-µF SPE, and analyzed by high performance liquid chromatography-fluorescence detector (HPLC-FLD). SPE parameters as type of elution solvent, elution solvent volume, pH value of sample solution, flow rate of sample solution, and breakthrough volume were first optimized in detail. Under the optimal conditions, the PI-µF SPE/HPLC-FLD method showed high recoveries (96.8%-107%), wide linearity (0.05-50 µg L-1, or 0.01-10 µg L-1), high determination coefficients (R2 ≥0.9992), and low limits of detection (LODs, 0.005-0.014 µg L-1). For the real tap water, egg and milk samples, the recoveries and RSDs were 81-119% and 0.8-9.8%, respectively. The results show that porous microfiber up to 25 µm in diameter is a promising solid-phase extraction adsorbent with the lowest flow resistance that can be used for trace organic pollutants in water and food samples.

Promoter hypomethylation of TIMP3 is associated with pre-eclampsia in a Chinese population.

A study by Yuen RK, Penaherrera MS, von Dadelszen P, McFadden DE, Robinson WP. DNA methylation profiling of human placentas reveals promoter hypomethylation of multiple genes in early-onset preeclampsia. Eur J Hum Genet 2010;18:1006-1012 based on a Canadian population found the tissue inhibitor of the metalloproteinase 3 (TIMP3) gene to be hypomethylated in pre-eclampsia (PE) placentas and to be a potential prenatal marker for early onset PE. To further explore the role of TIMP3 in PE and to investigate whether the TIMP3 promoter shows the same methylation pattern in the Han Chinese population, we analyzed a complete methylation assay of TIMP3 including the promoter region studied in the Canadian report and the neighboring CpG island in placentas (cases n = 41, controls n = 22) maternal peripheral blood (cases n = 3; controls n = 6) and umbilical cord blood (cases n = 7; controls n = 8) using MassArray EpiTyper (Sequenom, San Diego, CA, USA). Our results confirmed the finding of aberrant TIMP3 promoter methylation in PE placentas (mean = 0.405) compared with those in controls (mean = 0.534, P = 9.40 × 10(-7)). A tissue-specific methylation pattern between placentas (mean = 0.459) and bloods (mean = 0.961, P = 6.91 × 10(-13)) was also demonstrated in our clinical samples. Furthermore, a nearly 2-fold increase in TIMP3 expression for the hypomethylated promoter was found in PE placentas (P = 0.007), pointing to a negative relationship between TIMP3 methylation and the expression (R = -0.758, P = 0.029). In conclusion, we replicated the findings of Yuen et al. in our Han Chinese-based study, confirming that TIMP3 is likely to be involved in the etiology of PE and that hypomethylated and placenta-specific TIMP3 may be a potential marker for early diagnosis of PE in maternal plasma.

Vacuum-assisted thermal bonding of ß-cyclodextrin and its derivatives as chiral stationary phases for high-performance liquid chromatography.

In this work, the vacuum-assisted thermal bonding method was proposed for the covalent coupling of ß-cyclodextrin (ß-CD) (CD-CSP), hexamethylene diisocyanate cross-linked ß-CD (HDI-CSP) and 3, 5-dimethylphenyl isocyanate modified ß-CD (DMPI-CSP) onto the isocyanate silane modified silica gel. Under vacuum conditions, the side reaction due to the water residue from the organic solvent, air, reaction vessels and silica gel could be avoided, and the optimal temperature and time of vacuum-assisted thermal bonding method were determined as 160°C and 3 h. These three CSPs were characterized by FT-IR, TGA, elemental analysis and the nitrogen adsorption-desorption isotherms. The surface coverage of CD-CSP and HDI-CSP on silica gel was determined as ∼0.2 µmol m-2, respectively. The chromatographic performances of these three CSPs were systematically evaluated by separating 7 flavanones, 9 triazoles and 6 chiral alcohols enantiomers under the reversed-phase condition. It was found that the chiral resolution ability of CD-CSP, HDI-CSP and DMPI-CSP was complementary to each other. Among them, CD-CSP could separate all 7 flavanones enantiomers with the resolution of 1.09-2.48. HDI-CSP had a good separation performance for triazoles enantiomers with one chiral center. DMPI-CSP showed excellent separation performance for chiral alcohol enantiomers, among which the resolution of trans-1, 3-diphenyl-2-propen-1-ol reached 12.01. Generally, the vacuum-assisted thermal bonding had been demonstrated as a direct and efficient method for the preparation of chiral stationary phases of ß-CD and its derivatives.

Promoter hypomethylation of COMT in human placenta is not associated with the development of pre-eclampsia.

Disruption of the Catechol-O-methyltransferase (COMT) gene has been shown to be involved in pre-eclampsia (PE). To investigate whether two promoters of the COMT gene are differentially regulated by methylation in PE patients, we have analyzed the genomic DNA extracted from placenta (cases n = 16; controls n = 21), maternal peripheral blood (cases n = 4; controls n = 6) and umbilical cord blood (cases n = 8; controls n = 8) of women with PE and women with normal pregnancy. Bisulfite sequencing identified the predominantly unmethylated MB-COMT promoter in placenta, maternal peripheral blood and umbilical cord blood samples (PE and control). Subsequent quantitative MassArray data confirmed a significant tissue-specific hypomethylation of the S-COMT promoter in placenta (mean = 28.6%) when compared with its densely methylated patterns in blood samples (mean = 74.5%, P < 0.001), consistent with the sequencing data. However, no PE-specific methylation difference was found between cases and controls either in placenta or in blood samples. Moreover, none of the clinical characteristics had an effect on the methylation status of the S-COMT promoter. This study does not support a causal link between methylation regulation of COMT promoters and PE. However, the observed placenta-specific S-COMT promoter may be a potential marker for early prediction of PE in maternal plasma, although this remains to be further evaluated.

Epigenome-wide association data implicate fetal/maternal adaptations contributing to clinical outcomes in preeclampsia.

Aim: To investigate the changes of placental DNA methylome in preeclampsia (PE). Materials & methods: We performed an epigenome-wide association study in a Chinese cohort and six published datasets consisting of 335 samples in total. Results & conclusion: Numerous consistently hypomethylated probes were associated with early-onset PE in different populations, with 2125 reaching epigenome-wide significance. The validated probes were enriched for cytosine-phosphate-guanine dinucleotide (CpG) sites partially methylated and located in genes related to trophoblast fusion. The methylation levels of the validated probes were associated with clinical severity, while the intermediate samples showed antagonistic fetal/maternal outcomes. The DNA methylation patterns of PE and clinically relevant obstetrical syndromes suggested partially common pathophysiologies and complex maternal/fetal adaptive responses contributing to variable clinical outcomes.

Aberrant expression and DNA methylation of lipid metabolism genes in PCOS: a new insight into its pathogenesis.

Background: Polycystic ovary syndrome (PCOS), whose etiology remains uncertain, is a highly heterogenous and genetically complex endocrine disorder. The aim of this study was to identify differentially expressed genes (DEGs) in granulosa cells (GCs) from PCOS patients and make epigenetic insights into the pathogenesis of PCOS. Results: Included in this study were 110 women with PCOS and 119 women with normal ovulatory cycles undergoing in vitro fertilization acting as the control group. RNA-seq identified 92 DEGs unique to PCOS GCs in comparison with the control group. Bioinformatic analysis indicated that synthesis of lipids and steroids was activated in PCOS GCs. 5-Methylcytosine analysis demonstrated that there was an approximate 25% reduction in global DNA methylation of GCs in PCOS women (4.44 ± 0.65%) compared with the controls (6.07 ± 0.72%; P < 0.05). Using MassArray EpiTYPER quantitative DNA methylation analysis, we also found hypomethylation of several gene promoters related to lipid and steroid synthesis, which might result in the aberrant expression of these genes. Conclusions: Our results suggest that hypomethylated genes related to the synthesis of lipid and steroid may dysregulate expression of these genes and promote synthesis of steroid hormones including androgen, which could partially explain mechanisms of hyperandrogenism in PCOS.

Targeted sequencing identifies a novel SH2D1A pathogenic variant in a Chinese family: Carrier screening and prenatal genetic testing.

X-linked lymphoproliferative disease type 1 (XLP1) is a rare primary immunodeficiency characterized by a clinical triad consisting of severe EBV-induced hemophagocytic lymphohistiocytosis, B-cell lymphoma, and dysgammaglobulinemia. Mutations in SH2D1A gene have been revealed as the cause of XLP1. In this study, a pregnant woman with recurrence history of birthing immunodeficiency was screened for pathogenic variant because the proband sample was unavailable. We aimed to clarify the genetic diagnosis and provide prenatal testing for the family. Next-generation sequencing (NGS)-based multigene panel was used in carrier screening of the pregnant woman. Variants of immunodeficiency related genes were analyzed and prioritized. Candidate variant was verified by using Sanger sequencing. The possible influence of the identified variant was evaluated through RNA assay. Amniocentesis, karyotyping, and Sanger sequencing were performed for prenatal testing. We identified a novel de novo frameshift SH2D1A pathogenic variant (c.251_255delTTTCA) in the pregnant carrier. Peripheral blood RNA assay indicated that the mutant transcript could escape nonsense-mediated mRNA decay (NMD) and might encode a C-terminal truncated protein. Information of the variant led to success prenatal diagnosis of the fetus. In conclusion, our study clarified the genetic diagnosis and altered disease prevention for a pregnant carrier of XLP1.

Prevalence of Prediabetes Risk in Offspring Born to Mothers with Hyperandrogenism.

BACKGROUND: Excessive androgen exposure during pregnancy has been suggested to induce diabetic phenotypes in offspring in animal models. The aim of this study was to investigate whether pregestational maternal hyperandrogenism in human influenced the glucose metabolism in offspring via epigenetic memory from mother's oocyte to child's somatic cells. METHODS: Of 1782 reproductive-aged women detected pregestational serum androgen, 1406 were pregnant between 2005 and 2010. Of 1198 women who delivered, 1116 eligible mothers (147 with hyperandrogenism and 969 normal) were recruited. 1216 children (156 children born to mothers with hyperandrogenism and 1060 born to normal mother) were followed up their glycometabolism in mean age of 5years. Imprinting genes of oocyte from mothers and lymphocytes from children were examined. A pregestational hyperandrogenism rat model was also established. FINDINGS: Children born to women with hyperandrogenism showed increased serum fasting glucose and insulin levels, and were more prone to prediabetes (adjusted RR: 3.98 (95%CI 1.16-13.58)). Oocytes from women with hyperandrogenism showed increased insulin-like growth factor 2 (IGF2) expression. Lymphocytes from their children also showed increased IGF2 expression and decreased IGF2 methylation. Treatment of human oocytes with dihydrotestosterone upregulated IGF2 and downregulated DNMT3a levels. In rat, pregestational hyperandrogenism induced diabetic phenotypes and impaired insulin secretion in offspring. In consistent with the findings in human, hyperandrogenism also increased Igf2 expression and decreased DNMT3a in rat oocytes. Importantly, the same altered methylation signatures of Igf2 were identified in the offspring pancreatic islets. INTERPRETATION: Pregestational hyperandrogenism may predispose offspring to glucose metabolism disorder via epigenetic oocyte inheritance. Clinical trial registry no.: ChiCTR-OCC-14004537; www.chictr.org.

The aberrantly expressed miR-193b-3p contributes to preeclampsia through regulating transforming growth factor-ß signaling.

Preeclampsia (PE) is a leading cause of maternal mortality worldwide. Several studies have detected some differentially expressed microRNAs in the preeclamptic placenta, but few of the identified microRNAs demonstrated consistent findings among different research studies. In this study, high-throughput microRNA sequencing (HTS) of 9 preeclamptic and 9 normal placentas was performed. Seventeen microRNAs were identified to be up-regulated, and 8 down-regulated in preeclamptic placentas. Eight differentially expressed microRNAs except one identified in our study were determined to be consistent with at least one previous study, while sixteen were newly found. We performed qRT-PCR with independent 22 preeclamptic placentas and 20 control placentas to verify the differentially expressed microRNAs, and ten microRNAs were validated. The predicted target genes of the aberrantly expressed miR-193b-3p were enriched in the following gene ontology categories: cell motility and migration, cell proliferation and angiogenesis. We also found that miR-193b-3p significantly decreased the migration and invasion of trophoblast (HTR-8/SVneo) cells and that miR-193b-3p could regulate trophoblasts migration and invasion through binding onto the 3'UTR target site of TGF-ß2. In conclusion, we identified a list of differentially expressed microRNAs in PE placentas by HTS and provided preliminary evidence for the role of miR-193b-3p in the pathogenesis of preeclampsia.

Altered DNA methylation in neonates born large-for-gestational-age is associated with cardiometabolic risk in children.

BACKGROUND: Infants being born Large-for-gestational-age (LGA) are prone to developing cardiometabolic disease. However, the underlying mechanisms remain unclear. RESULTS: Clinical investigation showed that children born LGA had significantly higher serum level of total cholesterol (TC), low-density lipoprotein-cholesterol (LDL-c), and insulin, ratio of TC/high-density lipoprotein-cholesterol (HDL-c) compared to children born appropriate for gestational age (AGA). Birth weight (BW) was positively correlated to TC, LDL-c, and the ratio of TC/HDL in serum. Genome-wide DNA methylation analyzed in umbilical cord blood of controls and macrosomia cases. We identified 3459 methylation variable positions (MVPs) achieving genome-wide significance (adjusted P-value < 0.05) with methylation differences of ≥ 5%. A total of 327 MVPs were filtered by methylation differences of ≥ 7% located within an island, which mapped to 213 genes. Function analysis using Ingenuity Pathway Analysis showed 16 genes enriched in "cardiovascular disease". Four genes included contributed to hyperlipidemia. MATERIALS AND METHODS: Fifty-eight children aged 3-6 years born LGA and 123 subjects born AGA were enrolled. Anthropometric parameters and blood pressure (BP) were measured, and metabolic assessment was performed in all subjects. Genome-wide DNA methylation in umbilical blood was assayed by the 450K BeadChip in six AGA and six macrosomia newborns. CONCLUSIONS: Our data indicate that excess birth weight may increase the risk of lipid dysfunction in children aged 3-6 years. It might through reprogramming a group of genes correlated to cardiovascular disease. The genes identified in this study might be potential biomarker for cardiometabolic disease.