Comprehensive analysis of genomic complexity in the 5' end coding region of the DMD gene in patients of exons 1-2 duplications based on long-read sequencing.

BACKGROUND: Dystrophinopathies are the most common X-linked inherited muscle diseases, and the disease-causing gene is DMD. Exonic duplications are a common type of pathogenic variants in the DMD gene, however, 5' end exonic duplications containing exon 1 are less common. When assessing the pathogenicity of exonic duplications in the DMD gene, consideration must be given to their impact on the reading frame. Traditional molecular methods, such as multiplex ligation-dependent probe amplification (MLPA) and next-generation sequencing (NGS), are commonly used in clinics. However, they cannot discriminate the precise physical locations of breakpoints and structural features of genomic rearrangement. Long-read sequencing (LRS) can effectively overcome this limitation. RESULTS: We used LRS technology to perform whole genome sequencing on three families and analyze the structural variations of the DMD gene, which involves the duplications of exon 1 and/or exon 2. Two distinct variant types encompassing exon 1 in the DMD Dp427m isoform and/or Dp427c isoform are identified, which have been infrequently reported previously. In pedigree 1, the male individuals harboring duplication variant of consecutive exons 1-2 in the DMD canonical transcript (Dp427m) and exon 1 in the Dp427c transcript are normal, indicating the variant is likely benign. In pedigree 3, the patient carries complex SVs involving exon 1 of the DMD Dp427c transcript showing an obvious phenotype. The locations of the breakpoints and the characteristics of structural variants (SVs) are identified by LRS, enabling the classification of the variants' pathogenicity. CONCLUSIONS: Our research sheds light on the complexity of DMD variants encompassing Dp427c/Dp427m promoter regions and emphasizes the importance of cautious interpretation when assessing the pathogenicity of DMD 5' end exonic duplications, particularly in carrier screening scenarios without an affected proband.

Developing a Predictive Model for Minimal or Mild Endometriosis as a Clinical Screening Tool in Infertile Women: Uterosacral Tenderness as a Key Predictor.

STUDY OBJECTIVE: To develop a noninvasive predictive model based on patients with infertility for identifying minimal or mild endometriosis. DESIGN: A retrospective cohort study. SETTING: This study was conducted at a tertiary referral center. PATIENTS: A total of consecutive 1365 patients with infertility who underwent laparoscopy between January 2013 and August 2020 were divided into a training set (n = 910) for developing the predictive model and a validation set (n = 455) to confirm the model's prediction efficiency. The patients were randomly assigned in a 2:1 ratio. INTERVENTIONS: Sensitivities, specificities, area under the curve, the Hosmer-Lemeshow goodness of fit test, Net Reclassification Improvement index, and Integrated Discrimination Improvement index were evaluated in the training set to select the optimum model. In the validation set, the model's discriminations, calibrations, and clinical use were tested for validation. MEASUREMENTS AND MAIN RESULTS: In the training set, there were 587 patients with minimal or mild endometriosis and 323 patients without endometriosis. The combination of clinical parameters in the model was evaluated for both statistical and clinical significance. The best-performing model ultimately included body mass index, dysmenorrhea, dyspareunia, uterosacral tenderness, and serum cancer antigen 125 (CA-125). The nomogram based on this model demonstrated sensitivities of 87.7% and 93.3%, specificities of 68.6% and 66.4%, and area under the curve of 0.84 (95% confidence interval 0.81-0.87) and 0.85 (95% confidence interval 0.80-0.89) for the training and validation sets, respectively. Calibration curves and decision curve analyses also indicated that the model had good calibration and clinical value. Uterosacral tenderness emerged as the most valuable predictor. CONCLUSION: This study successfully developed a predictive model with high accuracy in identifying infertile women with minimal or mild endometriosis based on clinical characteristics, signs, and cost-effective blood tests. This model would assist clinicians in screening infertile women for minimal or mild endometriosis, thereby facilitating early diagnosis and treatment.

Rapid and non-invasive diagnostic techniques for embryonic developmental potential: a metabolomic analysis based on Raman spectroscopy to identify the pregnancy outcomes of IVF-ET.

The non-invasive and rapid assessment of the developmental potential of embryos is of great clinical importance in assisted reproductive technology (ART). In this retrospective study, we analyzed the metabolomics of 107 samples provided by volunteers and utilized Raman spectroscopy to detect the substance composition in the discarded culture medium of 53 embryos resulting in successful pregnancies and 54 embryos that did not result in pregnancy after implantation. The culture medium from D3 cleavage-stage embryos was collected after transplantation and a total of 535 (107 × 5) original Raman spectra were obtained. By combining several machine learning methods, we predicted the developmental potential of embryos, and the principal component analysis-convolutional neural network (PCA-CNN) model achieved an accuracy rate of 71.5%. Furthermore, the chemometric algorithm was used to analyze seven amino acid metabolites in the culture medium, and the data showed significant differences in tyrosine, tryptophan, and serine between the pregnancy and non-pregnancy groups. The results suggest that Raman spectroscopy, as a non-invasive and rapid molecular fingerprint detection technology, shows potential for clinical application in assisted reproduction.

Efficacy of dehydroepiandrosterone priming in women with poor ovarian response undergoing IVF/ICSI: a meta-analysis.

Background: Dehydroepiandrosterone (DHEA) may improve the outcomes of patients with poor ovarian response (POR) or diminished ovarian reserve (DOR) undergoing IVF/ICSI. However, the evidence remains inconsistent. This study aimed to investigate the efficacy of DHEA supplementation in patients with POR/DOR undergoing IVF/ICSI. Methods: PubMed, Web of Science, Cochrane Library, China National Knowledge Infrastructure (CNKI) were searched up to October 2022. Results: A total of 32 studies were retrieved, including 14 RCTs, 11 self-controlled studies and 7 case-controlled studies. In the subgroup analysis of only RCTs, DHEA treatment significantly increased the number of antral follicle count (AFC) (weighted mean difference : WMD 1.18, 95% confidence interval(CI): 0.17 to 2.19, P=0.022), while reduced the level of bFSH (WMD -1.99, 95% CI: -2.52 to -1.46, P<0.001), the need of gonadotropin (Gn) doses (WMD -382.29, 95% CI: -644.82 to -119.76, P=0.004), the days of stimulation (WMD -0.90, 95% CI: -1.34 to -0.47, P <0.001) and miscarriage rate (relative risk : RR 0.46, 95% CI: 0.29 to 0.73, P=0.001). The higher clinical pregnancy and live birth rates were found in the analysis of non-RCTs. However, there were no significant differences in the number of retrieved oocytes, the number of transferred embryos, and the clinical pregnancy and live birth rates in the subgroup analysis of only RCTs. Moreover, meta-regression analyses showed that women with lower basal FSH had more increase in serum FSH levels (b=-0.94, 95% CI: -1.62 to -0.25, P=0.014), and women with higher baseline AMH levels had more increase in serum AMH levels (b=-0.60, 95% CI: -1.15 to -0.06, P=0.035) after DHEA supplementation. In addition, the number of retrieved oocytes was higher in the studies on relatively younger women (b=-0.21, 95% CI: -0.39 to -0.03, P=0.023) and small sample sizes (b=-0.003, 95% CI: -0.006 to -0.0003, P=0.032). Conclusions: DHEA treatment didn't significantly improve the live birth rate of women with DOR or POR undergoing IVF/ICSI in the subgroup analysis of only RCTs. The higher clinical pregnancy and live birth rates in those non-RCTs should be interpreted with caution because of potential bias. Further studies using more explicit criteria to subjects are needed. Systematic review registration: https://www.crd.york.ac.uk/prospero/, identifier CRD 42022384393.

Large-scale analysis of de novo mutations identifies risk genes for female infertility characterized by oocyte and early embryo defects.

BACKGROUND: Oocyte maturation arrest and early embryonic arrest are important reproductive phenotypes resulting in female infertility and cause the recurrent failure of assisted reproductive technology (ART). However, the genetic etiologies of these female infertility-related phenotypes are poorly understood. Previous studies have mainly focused on inherited mutations based on large pedigrees or consanguineous patients. However, the role of de novo mutations (DNMs) in these phenotypes remains to be elucidated. RESULTS: To decipher the role of DNMs in ART failure and female infertility with oocyte and embryo defects, we explore the landscape of DNMs in 473 infertile parent-child trios and identify a set of 481 confident DNMs distributed in 474 genes. Gene ontology analysis reveals that the identified genes with DNMs are enriched in signaling pathways associated with female reproductive processes such as meiosis, embryonic development, and reproductive structure development. We perform functional assays on the effects of DNMs in a representative gene Tubulin Alpha 4a (TUBA4A), which shows the most significant enrichment of DNMs in the infertile parent-child trios. DNMs in TUBA4A disrupt the normal assembly of the microtubule network in HeLa cells, and microinjection of DNM TUBA4A cRNAs causes abnormalities in mouse oocyte maturation or embryo development, suggesting the pathogenic role of these DNMs in TUBA4A. CONCLUSIONS: Our findings suggest novel genetic insights that DNMs contribute to female infertility with oocyte and embryo defects. This study also provides potential genetic markers and facilitates the genetic diagnosis of recurrent ART failure and female infertility.

Karyopherin α deficiency contributes to human preimplantation embryo arrest.

Preimplantation embryo arrest (PREMBA) is a common cause of female infertility and recurrent failure of assisted reproductive technology. However, the genetic basis of PREMBA is largely unrevealed. Here, using whole-exome sequencing data from 606 women experiencing PREMBA compared with 2,813 controls, we performed a population and gene-based burden test and identified a candidate gene, karyopherin subunit α7 (KPNA7). In vitro studies showed that identified sequence variants reduced KPNA7 protein levels, impaired KPNA7 capacity for binding to its substrate ribosomal L1 domain-containing protein 1 (RSL1D1), and affected KPNA7 nuclear transport activity. Comparison between humans and mice suggested that mouse KPNA2, rather than mouse KPNA7, acts as an essential karyopherin in embryonic development. Kpna2-/- female mice showed embryo arrest due to zygotic genome activation defects, recapitulating the phenotype of human PREMBA. In addition, female mice with an oocyte-specific knockout of Rsl1d1 recapitulated the phenotype of Kpna2-/- mice, demonstrating the vital role of substrate RSL1D1. Finally, complementary RNA (cRNA) microinjection of human KPNA7, but not mouse Kpna7, was able to rescue the embryo arrest phenotype in Kpna2-/- mice, suggesting mouse KPNA2 might be a homologue of human KPNA7. Our findings uncovered a mechanistic understanding for the pathogenesis of PREMBA, which acts by impairing nuclear protein transport, and provide a diagnostic marker for PREMBA patients.

Mutations in PLCZ1 induce male infertility associated with polyspermy and fertilization failure.

PURPOSE: To investigate the genetic causes of polyspermy and total fertilization failure (TFF) in two independent male patients suffering from male infertility. METHODS: Immunofluorescence (IF) staining was used to detect the localization of the PLCζ protein in sperm and the maternal pronucleus in the zygote. Genomic DNA samples were extracted from the peripheral blood of patients and their families. The ExAC database was used to identify the frequency of corresponding mutations. The PLCZ1 mutations were validated by Sanger sequencing. The pathogenicity of the identified mutations and their possible effects on the protein were assessed using in silico tools and molecular modeling. RESULTS: We identified a reported homozygous mutation c.588C > A (p.Cys196Ter) and a compound heterozygous mutation c.2 T > C(p.Met1Thr)/c.590G > A (p.Arg197His) with one novel mutation in PLCZ1. The IF results showed that these multipronuclear zygotes formed as a result of polyspermy. In silico analysis predicted that the mutations result in disease-causing proteins. IF staining revealed that PLCζ is abnormally localized in the sperm samples from the two affected patients. Assisted oocyte activation (AOA) successfully rescued polyspermy and TFF and achieved pregnancy in two patients with the PLCZ1 mutation. CONCLUSION: We identified a homozygous mutation in PLCZ1 (c.588C > A [p.Cys196Ter]) in a male patient with polyspermy after in vitro fertilization (IVF) as well as a compound heterozygous mutation c.2 T > C(p.Met1Thr)/c.590G > A (p.Arg197His) with one novel mutation in a male patient with fertilization failure after intracytoplasmic sperm injection (ICSI), and we provide evidence that the homozygous mutation can cause polyspermy and the compound heterozygous mutation can cause fertilization failure.

[Effect of complete Y chromosome AZFc microdeletion on embryo euploidy].

OBJECTIVE: Preimplantation genetic testing (PGT) was performed to analyze the embryo euploidy in patients with complete Y chromosome AZFc microdeletion. METHODS: The clinical data of complete AZFc microdeletion underwent PGT from January 2013 to December 2021 in Reproductive Medicine Center of the First Affiliated Hospital of Nanjing Medical University were retrospectively analyzed. The patients with monogenic disease who underwent PGT during the same period were set as the control group. The basic characteristics, fertilization rate, Day 3 high quality embryo rate, blastocyst formation rate, embryo euploid rate, 45, X embryo ratio was compared between the two groups. RESULTS: A total of 220 patients were included, including 91 patients with complete AZFc microdeletion and 129 patients with monogenic disease. There was no significant difference in age between the two groups. In semen parameters, the sperm concentration, total sperm count and progressive motility in AZFc microdeletion group were lower than those in monogenic disease group, and the differences were statistically significant (P=0.001). The fertilization rate of AZFc microdeletion group was lower than that in monogenic disease group (P=0.012), and there was no significant difference in the number of MII oocytes, Day 3 high-quality embryo rate and blastocyst formation rate. A total of 933 blastocysts were successfully tested, including 496 blastocysts in AZFc deletion group and 437 blastocysts in monogenic disease group. The euploid, aneuploid and mosaic rates of the AZFc microdeletion group were 57.26%, 24.60% and 18.14%, respectively, while those of the monogenic disease group were 66.13%, 23.80% and 10.07%, with statistically significant differences between the two groups (P=0.001). Further analysis of the two groups of aneuploid embryos showed that aberrations occurred most commonly in chromosome16 (3.87%), X (3.46%), 13 (2.44%), 22 (2.24%) and 19 (2.03%) in AZFc microdeletion group, respectively, while the monogenic disease group was 22 (4.35%), 16 (2.97%), 7 (2.74%), 15(1.60%) and 2(1.60%), respectively. The proportion of sex chromosome abnormality in AZFc microdeletion group was higher than that in monogenic disease group (P=0.039), and there was no significant difference in the proportion of 45,X between the two groups. CONCLUSION: Compared with monogenic disease group, The embryo euploid rate in AZFc microdeletion patients decreased and the proportion of 45, X embryos did not increase significantly. It is recommended to select euploid female embryos by PGT, which not only avoids vertical transmission of AZFc microdeletion, but also reduces the risk of miscarriage due to aneuploid embryos.

Leukocyte telomere length in children born following blastocyst-stage embryo transfer.

Perinatal and childhood adverse outcomes associated with assisted reproductive technology (ART) has been reported, but it remains unknown whether the initial leukocyte telomere length (LTL), which is an indicator of age-related phenotypes in later life, is affected. Here, we estimated the LTLs of 1,137 individuals from 365 families, including 202 children conceived by ART and 205 children conceived spontaneously from two centers of the China National Birth Cohort, using whole-genome sequencing (WGS) data. One-year-old children conceived by ART had shorter LTLs than those conceived spontaneously (beta, -0.36; P = 1.29 × 10-3) after adjusting for plurality, sex and other potential confounding factors. In particular, blastocyst-stage embryo transfer was associated with shorter LTL (beta, -0.54, P = 2.69 × 10-3) in children conceived by ART. The association was validated in 586 children conceived by ART from five centers using different LTL quantification methods (that is, WGS or qPCR). Blastocyst-stage embryo transfer resulted in shorter telomere lengths in mice at postnatal day 1 (P = 2.10 × 10-4) and mice at 6 months (P = 0.042). In vitro culturing of mice embryos did not result in shorter telomere lengths in the late cleavage stage, but it did suppress telomerase activity in the early blastocyst stage. Our findings demonstrate the need to evaluate the long-term consequences of ART, particularly for aging-related phenotypes, in children conceived by ART.

Congenital absence of the vas deferens with hypospadias or without hypospadias: Phenotypic findings and genetic considerations.

Congenital absence of the vas deferens (CAVD) is a major cause of obstructive azoospermia. Mutations of CFTR and ADGRG2 cause the majority of CAVD. Despite this, 10%-20% of CAVD patients remain without a clear genetic diagnosis. Herein, the CFTR and ADGRG2 genes were first sequenced using Sanger sequencing in 50 CAVD patients. Whole-exome sequencing (WES) was used to further identify potential novel genetic causes in CAVD with hypospadias. In total, 29 of 50 CAVD patients carried at least one CFTR mutation, but no ADGRG2 mutation was found. 5T was found to be the most frequent variant in our CAVD populations. Seven CAVD patients with hypospadias were further analyzed using WES. No homozygous or compound heterozygous mutations related to disorders of sex development (DSDs) or male infertility were identified by WES. CAVD with hypospadias presented lower testicular volume (9.71 ± 2.14 ml vs. 14.45 ± 2.93 ml, p < 0.001) and higher FSH level (FSH: 7.28 ± 3.91 IU/L vs. 4.24 ± 1.96 IU/L, p = 0.027) than CAVD without hypospadias. It is worth noting that neither CFTR or ADGRG2 mutation nor homozygous or compound heterozygous gene mutations were identified in seven CAVD cases with hypospadias. However, nine heterozygous or hemizygous mutations were selected as potential pathogenic genes in CAVD with hypospadias. In conclusion, CFTR variants, especially 5T, play a major role in the Chinese CAVD population. CAVD with hypospadias shows relatively lower testicular spermatogenesis, suggesting a different genetic basis or pathogenic factor from cystic fibrosis/CAVD or unilateral renal agenesis/CAVD.

Peroxiredoxin 4 secreted by cumulus cells ameliorates the maturation of oocytes in vitro.

BACKGROUND: Peroxiredoxin 4 (Prdx4) in the endoplasmic reticulum (ER) is the only secretory member of the antioxidant Prdx family. Our previous studies demonstrated that Prdx4 in cumulus cells (CCs) ameliorated the maturation of oocytes in vitro and enhanced oocyte developmental competence by preventing CCs apoptosis caused by oxidative stress (OS) through gap junctions. In this study, we aimed to determine whether Prdx4 released by CCs can repair meiotic defects in mouse oocytes by co-culturing immature (germinal vesicle) oocytes with CCs from mature oocytes in the absence of gap junctions. RESULTS: The OS-induced meiotic defects in mouse oocytes were impeded by co-culture with CCs, as evidenced by the increased first polar body (PB1) extrusion rate and decreased ROS level. CCs increased Prdx4 expression and lowered IRE1α, Bip expression in H2O2-treated oocytes. After knockdown of Prdx4 expression in CCs, the rate of PB1 extrusion in the oocytes was significantly reduced to the level detected in H2O2 group, and ER stress was not alleviated. CO-IP and immunofluorescence co-localization experiments demonstrated that Prdx4 interacted with PDIA6 in the oocytes and the Pearson's R value was 0.69 calculated using ImageJ. CONCLUSIONS: Cumulus cells can promote the maturation of oocytes in vitro by secreting Prdx4 in a paracrine manner and serve as a promising therapeutic antioxidant for improving the quality of oocytes, especially aging oocytes, in clinical in vitro maturation (IVM).

[Analysis of three Chinese pedigrees affected with recurrent hydatidiform mole due to variants of NLRP7 gene].

OBJECTIVE: To explore the genetic etiology of recurrent hydatidiform mole (RHM) and provide accurate guidance for reproduction. METHODS: Peripheral venous blood samples of the probands with RHM and members from 5 unrelated pedigrees were collected. Genomic DNA was extracted by using routine method, and whole exome sequencing was carried out to detect variants of RHM-associated genes including NLRP7 and KHDC3L. Sanger sequencing and real-time quantitative PCR (RT-qPCR) were used to validate the candidate variants and delineate their parental origin. RESULTS: Homozygous or compound heterozygous variants of the NLRP7 gene were identified in four patients from three pedigrees, which included a homozygous deletion of exon 1 to 4 of NLRP7 in patient P1 and her elder sister, compound heterozygous variants of NLRP7 c.939delG (p.Q314Sfs*6) pat and c.1533delG (p.N512Tfs*4) mat in patient P2, and compound heterozygous variants of NLRP7 c.2389_2390delTC (p.A798Qfs*6) pat and c.2165A>G (p.D722G) mat in patient P4. All variants were interpreted as pathogenic or likely pathogenic according to the American College of Medical and Genomics (ACMG) guidelines. Among these, NLRP7 exons 1 to 4 deletion, c.939delG (p.Q314Sfs*6), c.1533delG (p.N512Tfs*4) and c.2389_2390delTC (p.A798Qfs*6) were unreported previously. CONCLUSION: Variants of the NLRP7 gene probably underlay autosomal recessive RHM in the three pedigrees, and definitive molecular diagnosis is beneficial for accurate genetic counseling. Above finding has also enriched the spectrum of the NLRP7 variants underlying RHM.

Single-Cell Quantitative Proteomic Analysis of Human Oocyte Maturation Revealed High Heterogeneity in In Vitro-Matured Oocytes.

Oocyte maturation is pertinent to the success of in vitro maturation (IVM), which is used to overcome female infertility, and produced over 5000 live births worldwide. However, the quality of human IVM oocytes has not been investigated at single-cell proteome level. Here, we quantified 2094 proteins in human oocytes during in vitro and in vivo maturation (IVO) by single-cell proteomic analysis and identified 176 differential proteins between IVO and germinal vesicle oocytes and 45 between IVM and IVO oocytes including maternal effect proteins, with potential contribution to the clinically observed decreased fertilization, implantation, and birth rates using human IVM oocytes. IVM and IVO oocytes showed separate clusters in principal component analysis, with higher inter-cell variability among IVM oocytes, and have little correlation between mRNA and protein changes during maturation. The patients with the most aberrantly expressed proteins in IVM oocytes had the lowest level of estradiol per mature follicle on trigger day. Our data provide a rich resource to evaluate effect of IVM on oocyte quality and study mechanism of oocyte maturation.

A retrospective study on IVF/ICSI outcomes in patients with persisted positive of anticardiolipin antibody: Effects of low-dose aspirin plus low molecular weight heparin adjuvant treatment.

Antiphospholipid (aPL) antibodies are more frequently detected among infertile women, but the association between aPL and in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) outcomes and whether need to get routine treatment are still controversial. The present study aims to find out whether infertile population with persistent aPL positive need treatment and which therapy is more effective. This retrospective study included 181 persistent aPL positive women, including 149 cases receiving anticoagulant treatment, either low-dose aspirin, low molecular weight heparin (LMWH) or aspirin plus LMWH adjuvant treatment (treated group), and 32 cases not receiving any treatment (untreated group). The treated group were further divided by combination therapy group (using both aspirin and LMWH，52 cases) and monotherapy group (only using aspirin，76 cases). The live birth rate and other clinical outcomes, including pregnancy rate, implantation rate, ongoing pregnancy rate and miscarriage rate were compared. The results show anticoagulant therapy can significantly improve live birth rate (59.06 % VS 34.48 %, P = 0.019), implantation rate (59.64 % VS 46.15 %, P＜0.001), ongoing pregnancy rate (59.73 % VS 34.38 %, P = 0.016), as well as reduce miscarriage rate (8.25 % VS 31.25 %, P＜0.001). Combination treatment of aspirin and LMWH exerts a higher live birth rate than monotherapy (75.00 % VS 53.95 %, P = 0.026). Infertile women with aPL positive might be classified as high-risk and low-risk aPL profiles. Those high-risk aPL positive infertile populations should be identified during IVF/ICSI and given corresponding thromboprophylaxis, and aspirin plus LMWH adjuvant treatment might be recommended.

Pregnancy and Perinatal Outcomes of Patients With Prior Cesarean Section After a Single Embryo Transfer in IVF/ICSI: A Retrospective Cohort Study.

Objective: To study the influence of the previous cesarean section on the pregnancy outcomes and perinatal outcomes in single embryo transfer (SET) cycles in an in vitro fertilization/intracytoplasmic sperm injection-embryo transfer (IVF/ICSI-ET) setting compared to those with previous vaginal delivery (VD). In addition, the association between fertility outcomes and different cesarean scar defect (CSD) sizes was studied. Method: This was a retrospective cohort study conducted in the Reproductive Center of the First Affiliated Hospital of Nanjing Medical University. A total of 4,879 patients with previous delivery history undergoing SET were included between January 2015 and April 2019. Patients were divided into the VD group and cesarean delivery (CD) group according to different modes of previous delivery. The primary outcome was live birth rate. The pregnancy outcomes of CD were analyzed as a subgroup and the relationship between pregnancy outcomes as well as the different sizes of CSD were explored by logistic regression analysis. Results: There were no significant differences in live birth rate, clinical pregnancy rate, and miscarriage rate between the CD group and VD group. The incidence rates of pregnancy complications such as pregnancy hypertension, gestational diabetes mellitus, placenta abnormalities, premature rupture of membrane, and postpartum hemorrhage were similar in the two groups. Live birth rate was significantly lower in the CSD group (23.77% vs 37.01%, aOR: 0.609, 95% CI: 0.476-0.778) comparing to patients without CSD. There were also significant differences in clinical pregnancy rate (37.52% vs 47.64%, aOR: 0.779, 95%CI: 0.623-0.973) and miscarriage rate (34.55% vs 20.59%, aOR: 1.407, 95%CI:1.03-1.923). Large size CSD significantly decreased live birth rate (13.33% vs 26.29%, aOR: 0.422, 95%CI: 0.197-0.902) and clinical pregnancy rate (25.33% vs 40.09%, aOR: 0.503, 95%CI: 0.272-0.930) compared with small size CSD. Conclusion: For women with previous cesarean sections, the pregnancy outcomes were similar to those with previous VD without increased perinatal complications following SET. The presence of CSD was associated with a marked reduction in live birth rate, especially in patients with large size CSD.

Associations between antenatal corticosteroid exposure and neurodevelopment in infants.

BACKGROUND: It has been well recognized that antenatal administration of dexamethasone to pregnant women at risk of preterm delivery may markedly accelerate fetal maturation and reduce the risk of adverse perinatal outcomes in their preterm infants, particularly for births before 34 weeks of gestation. Since 2015, antenatal corticosteroid administration has been extended beyond 34 weeks of gestation by clinical guidelines, as it might have beneficial effects on fetal maturation and perinatal outcomes. However, concerns regarding the potential influence of antenatal corticosteroid treatment on offspring neurodevelopment have been raised. OBJECTIVE: This study aimed to investigate whether maternal antenatal corticosteroid administration was associated with neurodevelopment in infants at 1 year of age. STUDY DESIGN: In this prospective and longitudinal birth cohort study, women were followed up throughout gestation, and their infants underwent a Bayley Scales of Infant and Toddler Development, Third Edition, screening test at 1 year of age between December 2018 and September 2020. Finally, 1609 pregnant women and 1759 infants were included in the current study. Using a generalized linear mixed model, we examined the association between antenatal corticosteroid exposure and infant neurodevelopment in cognitive, receptive communication, expressive communication, fine motor, and gross motor functions. RESULTS: Of the 1759 infants eligible for this study, 1453 (82.6%) were singletons. A total of 710 infants were exposed to antenatal corticosteroids, among whom 415 were dexamethasone exposed and 483 were prednisone exposed. Dexamethasone was prescribed most often in late pregnancy, whereas prednisone was often used before 8 weeks of gestation among women who conceived through assisted reproductive technology. Compared with those who had no exposure, antenatal corticosteroid exposure was associated with an increased risk of infants being noncompetent in the cognitive development domain after adjusting for conventional risk factors (adjusted risk ratio, 1.53; 95% confidence interval, 1.08-2.18; P=.017). For medication-specific exposure, those exposed vs not exposed to antenatal dexamethasone were 1.62-fold (95% confidence interval, 1.10-2.38; P=.014) more likely to be noncompetent in the cognitive development domain at 1 year. The association did not vary markedly between preterm and term infants, singletons and twins, or assisted reproductive technology-conceived and spontaneously conceived infants (all P>.05 for heterogeneity). In contrast, a null association was observed for the risk of being noncompetent in any domain of neurodevelopment with antenatal prednisone exposure at early pregnancy. CONCLUSION: Here, antenatal corticosteroid, particularly dexamethasone exposure, was markedly associated with an increased risk of infants being noncompetent in the cognitive development domain at 1 year of age. These findings may provide new information when weighing the benefits and potential risks of maternal antenatal corticosteroid administration.

Nucleoporin37 may play a role in early embryo development in human and mice.

Maternal-effect genes (MEGs) play an important role in maintaining the survival and development of mammalian embryos at the cleavage stage after fertilization. Despite long-term efforts, the MEGs that regulate preimplantation embryo development remain largely unknown. Here, using whole-exome sequencing and homozygosity mapping, we identified a potential candidate gene associated with early embryo development: nucleoporin37 (NUP37), a nucleoporin gene that encodes a member of the nuclear pore complexes and regulates nuclear pore permeability and nucleocytoplasmic transport. Moreover, we determined the temporal and spatial expression patterns of Nup37 in mouse oocytes and early embryos, and explored the role of NUP37 in oocyte maturation and preimplantation embryo development. Immunoprecipitation assays confirmed that yes-associated protein-1 (YAP1) binds to TEA domain transcription factor 4 (TEAD4) and NUP37. Furthermore, Nup37 gene knockdown reduced the nuclear import of YAP1 and down-regulated the expression of YAP1-TEAD pathway downstream genes Rrm2 and Rpl13 in early embryos. Our study provides evidence that maternal NUP37 contributes to the nuclear import of YAP1 and then activates the YAP1-TEAD pathway, a signalling pathway essential for zygotic genome activation. Nup37 may be a key gene involved in preimplantation embryo development in mammals.

Evaluation of Bone Mineral Density in Children Conceived via Assisted Reproductive Technology.

Objectives: To investigate bone mineral density (BMD) differences between assisted reproductive technology (ART)-conceived children and naturally conceived (NC) children. Study Design: This retrospective cohort study included ART-conceived children and controls aged 1 to 12 years assessed with a follow-up protocol. Maternal and paternal background, birth condition, and growth and development indicators were analyzed. Results: The ART and NC groups exhibited differences in maternal and paternal childbearing age; maternal weight; maternal body mass index (BMI); maternal alcohol consumption; paternal smoking; delivery method; and serum zinc, iron, and lead levels. Multifactor analysis adjusted for relevant factors showed that paternal childbearing age and group significantly affected the BMD Z score. In the subgroup analysis, in vitro fertilization (IVF) (p=0.026) or intracytoplasmic sperm injection (ICSI) (p=0.008) had a positive impact on the BMD Z score. Male infertility only (p=0.010) or male infertility combined with polycystic ovary syndrome (PCOS) (p=0.026) may affect the BMD Z score. In the embryo transfer cycle subgroup analysis, compared with natural conception, both stimulation cycle fresh embryo transfer (p=0.019) and natural cycle frozen embryo transfer (p=0.006) had a positive effect on the BMD Z score. Conclusions: The BMD levels of the ART and control groups were generally in the normal range. Paternal childbearing age and the use of ART independently affected the BMD Z score of the offspring.

Novel biallelic mutations in PADI6 in patients with early embryonic arrest.

Peptidyl arginine deiminase, type VI (PADI6) is a member of the subcortical maternal complex (SCMC), which plays vital roles in mammalian embryogenesis. Most mutations in SCMC members have been reported to cause human embryonic arrest, and a total of 15 mutations in PADI6 have been shown to be responsible for early embryonic arrest according to previous studies. However, the genetic factors behind this phenotype remain to be understood in further detail. Here, we identified 13 novel mutations and 4 previously reported mutations of PADI6 in 14 patients who were diagnosed with abnormal embryonic development caused by early arrest, embryonic fragmentation, and recurrent implantation failure. Most of the mutations were predicted by in silico analysis to be deleterious or damaging to the function of PADI6. In addition, the total and East Asian population frequencies of the mutations were low or absent in the gnomAD database. Our study expands the mutational spectrum in PADI6 and will provide precise targets for genetic counseling in the future.

Androgen Excess Increases Food Intake in a Rat Polycystic Ovary Syndrome Model by Downregulating Hypothalamus Insulin and Leptin Signaling Pathways Preceding Weight Gain.

BACKGROUND: Polycystic ovary syndrome (PCOS) is a common reproductive and metabolic disorder characterized by high androgen levels. The aim of this study was to evaluate the effects of hyperandrogenism on the hypothalamus and subsequently on the food intake and obesity in females. METHODS: A dihydroxy testosterone (DHT)-induced rat model was established to recapitulate the hyperandrogenism features of PCOS patients. Body weight and food intake of the rats were recorded. The food intake of DHT-induced rats was restricted by pair feeding to exclude possible effects of weight gain on the hypothalamus. The expression levels of relevant proteins and mRNAs in the hypothalamus and primary hypothalamic neurons exposed to DHT were analyzed by Western blotting and RT-PCR, respectively. The leptin levels in the serum and cerebrospinal fluid (CSF) were measured, and leptin was injected via the intracerebroventricular (ICV) route to test the leptin sensitivity of the hypothalamus. RESULTS: The excessive prepuberty androgen levels in the DHT-induced rats markedly elevated food intake prior to weight gain. Consistent with this, the expression of neuropeptide Y and agouti-related peptide mRNAs was upregulated, which occurred prior to obesity and even with restricted food intake. In addition, the hypothalamic sensitivity to insulin and leptin was also impaired in the DHT-induced rats before obesity and with restricted food intake. DHT significantly reduced the leptin levels in the CSF, and ICV injection of leptin inhibited the DHT-induced increase in food intake. CONCLUSIONS: Androgen excess increased food intake in rats and promoted obesity by downregulating insulin and leptin signaling in the hypothalamus, most likely by suppressing leptin levels in the CSF.