Rui Hua

Environmental phenolic endocrine-disrupting chemicals (EDCs) are ubiquitous in the environment and human matrices, raising growing concerns regarding maternal and child health. Here, we present a sensitive analytical method for the simultaneous quantification of 22 phenolic compounds—including 7 bisphenols, 2 alkylphenols, 11 pharmaceutical and personal care product-related phenols, and 2 endogenous estrogens—in human serum. The method combines enzymatic deconjugation, 96-well plate-based protein precipitation and phospholipid removal using HybridSPE technology with dansyl chloride derivatization and liquid chromatography–tandem mass spectrometry, requiring only 100 μL of serum per sample. Validation using fetal bovine serum as a surrogate matrix yielded limits of detection of 0.023–0.227 ng/mL, recoveries of 86–114%, relative standard deviations below 20%, and matrix effects ranging from −35% to −2%. The method was applied to 30 serum samples from pregnant women in southern China, with nine analytes—including bisphenol A, triclosan, and endogenous estrogens—detected in at least 70% of samples. The median concentrations of the target compounds following enzymatic deconjugation ranged from below limits of detection to 28.4 ng/mL (for estrone). This method enables the simultaneous analysis of environmental phenols and endogenous estrogens in limited-volume human serum, supporting biomonitoring studies.

Domoic acid (DA) is a potent neurotoxin that poses public health concerns, especially for children and adolescents during critical neurodevelopmental periods. In the present study, urinary DA concentrations in 216 children and adolescents at the age of 6 to 18 in southern China were determined using a novel dansyl-chloride (DNS-Cl) derivatization high performance liquid chromatography–tandem mass spectrometry (HPLC–MS/MS) method with ultrahigh sensitivity (LOQ: 0.087 ng/mL). The median urinary DA concentration was 2.17 ng/mL (interquartile range (IQR): 0.87–4.08 ng/mL). When analyzed by age group, the medians were 1.40 ng/mL (6–9 years; IQR: 0.55–3.49 ng/mL), 2.16 ng/mL (10–13 years; IQR: 0.94–4.07 ng/mL), and 2.93 ng/mL (14–18 years; IQR: 1.06–5.06 ng/mL). Our findings revealed that urinary DA concentrations increased with age and varied significantly across different body mass index groups (p < 0.05), while no significant gender differences were observed. The estimated daily intake (1.73–374 ng/kg/day) remained below established safety thresholds. This study represents the first systematic biomonitoring of urinary DA exposure in children and adolescents from southern China’s coastal communities, addressing critical knowledge gaps and establishing baseline data amid rising harmful algal bloom frequency.

Subclinical hypothyroidism (SCH) is associated with multiple adverse outcomes in early pregnancy. This study aims to explore the regulatory mechanisms underlying histone lactylation modification in early pregnancy with SCH. Peripheral blood mononuclear cells were collected from early pregnant women with or without SCH. RNA sequencing (RNA-seq) and chromatin immunoprecipitation sequencing (ChIP-seq) analyses were performed to identify the transcriptional pattern and histone lactylation modification in early pregnancy with SCH. RNA-seq analysis revealed that the differentially expressed genes associated with the extracellular matrix exhibited a significant downregulation in early pregnancy with SCH (EP_SCH) compared to early pregnancy without SCH (EP), while those involved in apoptosis were significantly upregulated. In the ChIP-seq analysis, 1660 hypomodified and 766 hypermodified H3K18la-binding peaks were identified in the EP_SCH group compared to the EP group. The hypomodified genes in early pregnancy with SCH compared to its control were enriched in GO terms of apoptotic process and differentiation of immune cells. The genes with increased H3K18 lactylation in early pregnancy with SCH compared to its control were associated with the nervous system, female pregnancy, and the OXT signaling pathway. When RNA-seq data was integrated with ChIP-seq data, we found that the expression and H3K18la enrichment of KCTD7, SIPA1L2, HDAC9, BCL2L14, TXNRD1, and SGK1 were increased in early pregnancy with SCH compared to its control, which was further confirmed by RT-qPCR and ChIP-PCR analyses. This study identifies the changes in histone lactylation modification in early pregnancy with SCH. These findings provide novel insights into the regulatory mechanisms of SCH during early pregnancy.

Human in vitro fertilized embryos exhibit low developmental capabilities, and the mechanisms that underlie embryonic arrest remain unclear. Here using a single-cell multi-omics sequencing approach, we simultaneously analysed alterations in the transcriptome, chromatin accessibility and the DNA methylome in human embryonic arrest due to unexplained reasons. Arrested embryos displayed transcriptome disorders, including a distorted microtubule cytoskeleton, increased genomic instability and impaired glycolysis, which were coordinated with multiple epigenetic reprogramming defects. We identified Aurora A kinase (AURKA) repression as a cause of embryonic arrest. Mechanistically, arrested embryos induced through AURKA inhibition resembled the reprogramming abnormalities of natural embryonic arrest in terms of the transcriptome, the DNA methylome, chromatin accessibility and H3K4me3 modifications. Mitosis-independent sequential activation of the zygotic genome in arrested embryos showed that YY1 contributed to human major zygotic genome activation. Collectively, our study decodes the reprogramming abnormalities and mechanisms of human embryonic arrest and the key regulators of zygotic genome activation.

Background: Polycystic ovary syndrome (PCOS) is a multisystem-related disease whose pathophysiology is still unclear. Several regulators of N6-methyladenosine (m6A) modification were confirmed to play a regulatory role in PCOS. Nonetheless, the roles of m6A regulators in PCOS are not fully demonstrated. Materials and methods: Four mRNA expression profiling microarrays were obtained from the Gene Expression Omnibus (GEO) database. Differentially expressed m6A regulators between PCOS and normal patients were identified by R software. A random forest modal and nomogram were developed to assess the relationship between m6A regulators and the occurrence risk of PCOS. A consensus clustering method was utilized to distinctly divide PCOS patients into two m6A subtypes (m6A cluster A/B). The patterns of differential expression and immune infiltration were explored between the two m6A clusters. Results: In this study, 22 significant m6A regulators were identified between healthy controls and PCOS patients. The random forest model determined three optimal m6A regulators which are related to the occurrence risk of PCOS, including YTHDF1, RBM15 and METTL14. A nomogram was established based on these genes, and its predictive reliability was validated by decision curve analysis. The consensus clustering algorithm distinctly divided PCOS cases into two m6A subtypes. The ssGSEA algorithm found that the immune infiltration was markedly enriched in m6A cluster B than in cluster A. The m6A-pattern related differentially expressed genes (DEGs) of the two m6A subtypes were demonstrated by differential expression analysis. We found that they were enriched in immune-related genes and various infection pathways. Based on the m6A-pattern related DEGs, the PCOS patients were classified into two m6A-pattern related genomic subtypes (gene clusters A and B). Conclusions: The present study provided evidence concerning the different modification patterns of m6A regulators in PCOS compared with normal patients. This study will help clarify the overall impact of m6A modification patterns and related immune infiltration on PCOS.

Background. Nonobstructive azoospermia (NOA) is a complex disease characterized by the spermatogenic dysfunction of testicular tissues. The roles played by long noncoding RNAs (lncRNAs) in NOA pathogenesis have not been extensively studied. Methods. Microarray assays were performed on samples of testicular biopsy tissue obtained from patients with NOA for the purpose of identifying differentially expressed lncRNAs and messenger RNA (mRNA) transcripts, and the results were verified by quantitative real‐time polymerase chain reaction. Mouse‐derived GC‐1 spermatogonia (spg) cells undergoing treatment with Adriamycin (ADR) were used to investigate the biological functions of the selected lncRNAs in vitro. The target microRNAs (miRNAs) of lncRNAs and the target mRNAs of miRNAs were predicted by a bioinformatics analysis. Functional studies performed using the CCK‐8 assay, EdU incorporation assay, apoptosis detection, and senescence‐associated β‐galactosidase (SA‐β‐Gal) staining were conducted using GC‐1 spg cells. Results. Totals of 2,652 lncRNAs and 2,625 mRNAs were found to be differentially expressed in the testicular tissue of NOA patients when compared with patients in a control group. Dynamin 3 opposite strand (DNM3OS) was a provider of pe‐miR‐214‐5p that positively regulates miR‐214‐5p expression in GC‐1 spg cells. The E2 factor (E2F) family of transcription factor 2 (E2F2) was initially predicted and subsequently verified to be a downstream gene of miR‐214‐5p. E2F2 expression was upregulated after DNM3OS knockdown in ADR‐treated GC‐1 spg cells. Moreover, knockdown of either DNM3OS or miR‐214‐5p significantly alleviated ADR‐induced decreases in cellular activity and proliferation, as well as increases in apoptosis and senescence of mouse spermatogonial GC‐1 spg cells. Conclusions. DNM3OS was found to regulate the apoptosis and senescence of spermatogonia by providing miR‐214‐5p and decreasing E2F2 expression, suggesting it as a novel target for gene therapy of male infertility.

Background: Anembryonic pregnancy (AP) is the most severe dysmorphogenesis of human embryo development and a frequent presentation of early pregnancy loss (EPL). Studies have analyzed the association between assisted reproductive technologies (ART) and EPL. However, the specific relationship between ART and AP has not been fully elucidated. Several studies suggested that non-genomic anomalies might be related to AP and ART might increase the risk of epigenetic changes, thus possibly detecting some associations between ART and AP. Our study aims to find out any possible risk factors of AP in ART treatments, and translate the results into clinical practice. Methods: A retrospective cohort study was conducted in Nanfang Hospital. Data from 1,765 singleton pregnancies following fresh or frozen-thawed embryo transfer from January 2014 to December 2017 were collated with the inclusion of EPLs and normal live births (NLB). Participants were divided into three groups: NLB (full-term birth with normal body weight infants), EPL (spontaneous pregnancy loss prior to 13 weeks gestation) with embryos (EE), and APs (embryonic pole was invisible in two consecutive ultrasound examinations). The basic characteristics of the patients and the association between ART-related variables and AP were analyzed using one-way analysis of variance (ANOVA) and multivariable logistic regression model, respectively. Products of conception (POC) from AP and EE patients received karyotype analysis using multiplex ligation-dependent probe amplification (MLPA). Results: Blastocyst culture of non-top-quality cleavage stage embryos almost doubled the percentage of AP in EPL (45.9% vs. 24.4%, P=0.037), and the normal euploid rate was significantly higher in the AP group (50.5% vs. 32.3%, P=0.003). Using multivariable logistic regression model, we found that blastocyst transfer and advanced maternal age might be risk factors for AP (OR >1, PConclusions: The occurrence of AP may be due to epigenetic abnormalities associated with advanced maternal age and extended in vitro embryo culture, while CoQ10 supplementation may be a potential method in preventing AP. Future multi-center prospective cohort studies should be conducted to verify these results.

At present, infertile patients with maturation arrest (MA) are difficult to obtain mature sperm. Spermatogenesis and its molecular mechanism are still not clear. Patients with MA and normal spermatogenesis (NS) were collected. iTRAQ-based proteomic approach was performed to reveal the different proteins between them. To validate the confidence of proteome data, the individual samples were analyzed by Western blotting (WB), quantitative polymerase chain reaction (qPCR), and immunofluorescence. The miR-449a and CEP55 were determined by Luciferase assay. Mouse GC-1 cells were transfected with CEP55 siRNAs, miR-449a mimic, or inhibitor, and cell proliferation was determined. Compared with NS, 27 proteins were differentially expressed in MA, and CEP55 protein was the most significant difference. WB and qPCR showed that CEP55 levels were significantly elevated in NS than MA. In transfected cells, overexpression of miR-449a and knockdown of CEP55 both downregulated CEP55 expression and decreased cell proliferation. miR-449a suppresses mouse spermatogonia proliferation via inhibition of CEP55.

To investigate the associations of occupational, educational and residential factors with the outcomes of in vitro fertilization -embryo transfer (IVF-ET). A retrospective analysis was conducted among 3998 patients undergoing the first treatment with IVF-ET and 3064 perinatal infants in our center between January, 2010 and June, 2014. The relation of the patients' occupation, educational background, and residence registry with IVF-ET outcomes was analyzed. The risk of abortion in the first trimester was significantly higher in female patients engaged in agriculture (OR=4.319, P=0.047). Occupation and educational background of the patients and their spouse were related with the incidence of ectopic pregnancy following IVF-ET. The total incidence rate of perinatal birth defects was 2.1% in this cohort, and was the highest in Shenzhen (5.5%) but this difference was not statistically significant (P>0.05). The patients' occupation and educational background are related with the outcomes of IVF-ET, suggesting the importance of health education and prenatal examination.