Comparison of Chromosomal Microarray Analysis and Noninvasive Prenatal Testing in Pregnant Women with Fetal Ultrasonic Soft Markers.

Objective: This study aimed to assess the utility of chromosomal microarray analysis (CMA) and noninvasive prenatal testing (NIPT) in detecting clinically significant chromosomal abnormalities among fetuses presenting ultrasonic soft markers (USMs). Methods: A retrospective observational study, spanning from January 1, 2019, to September 30, 2022, enrolled 539 singleton pregnant women with fetal USMs at our center. Of these, 418 cases (77.6%) underwent NIPT, while 121 cases (22.4%) opted for invasive prenatal diagnosis post-appropriate genetic counseling. Cases with high-risk NIPT results proceeded to invasive prenatal diagnosis, where conventional karyotyping and CMA were concurrently performed. Further stratification was done based on the number of USMs, classifying cases into single-USM and multiple-USM groups. Results: Of the 24 cases (4.5%) exhibiting abnormal findings, 17 presented numerical chromosomal abnormalities, 2 featured clinically significant copy number variations (CNVs), 3 showed variants of unknown significance (VOUS), 1 displayed LOH, and 1 exhibited chromosome nine inversion. Notably, 18 cases (75%) theoretically detectable by karyotyping (eg, sizes above 10Mb) and 16 cases (66.7%) detectable by NIPT for five common aneuploidies were identified. Six submicroscopic findings (25%) were exclusively detectable by CMA. The predominant clinically relevant aberrations were observed in the thickened nuchal-translucency (TNT) group (9/35, 25.7%), followed by the multiple soft markers group (3/32, 9.3%). In the NIPT group, the false positive rate was 1.22%, and the false negative rate was 0%. Conclusion: The prevalence of chromosome aneuploidy exceeded that of submicroscopic chromosomal imbalance in pregnant women with fetal USMs. NIPT demonstrated efficacy, particularly for soft markers like echogenic intracardiac focus. However, for those with TNT and multiple soft markers, invasive prenatal diagnosis, including CMA testing, is recommended as the primary investigative approach.

Clinical Biochemical Indicators and Intestinal Microbiota Testing Reveal the Influence of Reproductive Age Extending from the Mother to the Offspring.

Age is an important factor that determines the physiological functions of the human body, but the changes in maternal physiology, biochemistry, and intestinal flora related to reproductive age and their impact on offspring are not clear. Here, we tested and analyzed the clinical physiological and biochemical indicators and/or intestinal flora, matching the data of 252 parturients and their newborns. We found that 4 clinical indicators, including the white blood cell count and the absolute value of monocytes, were significantly related to the reproductive age (P < 0.05). The composition of the intestinal flora also varied with age, and the intestinal flora of advanced-age women (≥35 years old) was different from that of middle-aged women (>25 and <35 years old). We also found that changes in maternal clinical physiological and biochemical indicators related to reproductive age could reflect changes in the abundance of bacteria, such as Peptococcus and Vibrio, and changes in the intestinal flora spread to offspring. These results provide new evidence to explain the increased adverse pregnancy outcomes of mothers of inappropriate age, describe the increased health risks of newborns, help us examine the importance of age-appropriate birth from a broader perspective, and contribute to the discovery of mother-child bonds for a better understanding of healthy reproduction. IMPORTANCE In this study, we demonstrated that physiological indicators and the gut microbiome fluctuate drastically among parturients of different reproductive ages and that there is a significant correlation between the two changes. Mothers of different ages had different gut microbes, and the gut microbiota varied as the childbearing age became too high. Changes in the gut microbiome with maternal reproductive age affected the offspring, and the influence of reproductive age on the intestinal flora had a synergistic effect between mother and child that was revealed for the first time. The maternal childbearing age might affect the colonization of the offspring's initial flora. The results provide new evidence to explain the increased adverse pregnancy outcomes of mothers of inappropriate age, describe the increased health risks of newborns, and contribute to the discovery of mother-child bonds for a better understanding of healthy reproduction.

Maternal and neonatal viromes indicate the risk of offspring's gastrointestinal tract exposure to pathogenic viruses of vaginal origin during delivery.

A cumulative effect of enterovirus and gluten intake on the risk of celiac disease autoimmunity in infants highlights the significance of viral exposure in early life on the health of children. However, pathogenic viruses may be transmitted to the offspring in an earlier period, raising the possibility that women whose vaginas are inhabited by such viruses may have had their babies infected as early as the time of delivery. A high-resolution intergenerational virome atlas was obtained by metagenomic sequencing and virome analysis on 486 samples from six body sites of 99 mother-neonate pairs. We found that neonates had less diverse oral and enteric viruses than mothers. Vaginally delivered newborns seconds after birth had a more similar oral virome and more viruses of vaginal origin than cesarean-section (C-section) newborns (56.9% vs. 5.8%). Such viruses include both Lactobacillus phage and potentially pathogenic viruses, such as herpesvirus, vaccinia virus, and hepacivirus, illustrating a relatively high variety of the pioneer viral taxa at the time of delivery and a delivery-dependent mother-to-neonate transmission along the vaginal-oral-intestinal route. Neonates are exposed to vaginal viruses as they pass through the reproductive tract, and viruses of vaginal origin may threaten their health. These findings challenge the conventional notion that vaginal delivery is definitely better than cesarean delivery from the perspective of microbial transmission. Screening for vaginal virome before delivery is a worthwhile step to advocate in normal labor to eliminate the risk of intergenerational transmission of pathogenic viruses to offspring.

Deterministic transition of enterotypes shapes the infant gut microbiome at an early age.

BACKGROUND: The succession of the gut microbiota during the first few years plays a vital role in human development. We elucidate the characteristics and alternations of the infant gut microbiota to better understand the correlation between infant health and microbiota maturation. RESULTS: We collect 13,776 fecal samples or datasets from 1956 infants between 1 and 3 years of age, based on multi-population cohorts covering 17 countries. The characteristics of the gut microbiota are analyzed based on enterotype and an ecological model. Clinical information (n = 2287) is integrated to understand outcomes of different developmental patterns. Infants whose gut microbiota are dominated by Firmicutes and Bifidobacterium exhibit typical characteristics of early developmental stages, such as unstable community structure and low microbiome maturation, while those driven by Bacteroides and Prevotella are characterized by higher diversity and stronger connections in the gut microbial community. We further reveal a geography-related pattern in global populations. Through ecological modeling and functional analysis, we demonstrate that the transition of the gut microbiota from infants towards adults follows a deterministic pattern; as infants grow up, the dominance of Firmicutes and Bifidobacterium is replaced by that of Bacteroides and Prevotella, along with shifts in specific metabolic pathways. CONCLUSIONS: By leveraging the extremely large datasets and enterotype-based microbiome analysis, we decipher the colonization and transition of the gut microbiota in infants from a new perspective. We further introduce an ecological model to estimate the tendency of enterotype transitions, and demonstrated that the transition of infant gut microbiota was deterministic and predictable.

Maternal, umbilical arterial metabolic levels and placental Nrf2/CBR1 expression in pregnancies with and without 25-hydroxyvitamin D deficiency.

BACKGROUND: The aim of this case-control study was to document maternal, umbilical arterial metabolic levels and correlations in pregnancies with and without 25-hydroxyvitamin D [25(OH)D] deficiency, while, also investigating the expression of nuclear factor erythroid 2 related factor 2 (Nrf2) and carbonyl reductase 1 (CBR1) in the placenta. METHODS: One hundred participants, 50 deficient for 25(OH)D and 50 normal, were recruited from among hospitalized single-term pregnant women who had elected for cesarean section. Umbilical arterial and placental samples were collected during cesarean section. Metabolic levels were assessed for the 25(OH)D deficiency and control groups' maternal, umbilical arterial samples. Nrf2 and CBR1 expression levels were investigated in the placentas of 12 pregnant women with 25(OH)D deficiency and 12 controls. RESULTS: Compared with the control participants, the 25(OH)D deficient women had significantly higher triglyceride (TG) levels (3.80 ± 2.11 vs. 2.93 ± 1.16 mmol/L, 3.64 ± 1.84 vs. 2.81 ± 1.16 mmol/L, p < .01, .001); lower high density lipoprotein cholesterol (HDL-C) levels (1.54 ± 0.32 vs. 1.82 ± 0.63 mmol/L, 1.41 ± 0.72 vs. 2.44 ± 1.68 mmol/L, p < .001, .01) in both material blood and the umbilical artery. In addition, Nrf2 and CBR1 expression levels were lower in the maternal 25(OH)D deficient placenta. CONCLUSION: 25(OH)D deficient pregnant women have higher TG levels and lower HDL-C levels in both material blood and the umbilical artery. TG level is negatively correlated with 25(OH)D in both the maternal serum and infant umbilical artery. 25(OH)D deficiency also lowers placental expression of Nrf2 and CBR1.Supplemental data for this article is available online at here.

Disorder of Sexual Development Males With XYY in Blood Have Exactly X/XY/XYY Mosaicism in Gonad Tissues.

Y chromosome represents masculinization. The extra Y chromosome of XYY patients usually leads to over-masculinization phenotypes. The occurrence of several DSD cases with XYY in blood is controversial. Is XYY associated with disorder of sex development (DSD)? What is the mechanism behind DSD in males with XYY in blood? To this end, this study retrospectively analyzed blood-karyotype data of 4,437 DSD male children and karyotypes data of 6,259 newborn males as the control. Exome sequencing (ES) was performed to test whether the patients with DSD and with XYY in blood had other variants on known DSD-genes. Testicular biopsy was performed. Fluorescence in situ hybridization (FISH) was used to test whether a sex chromosome mosaicism was present in the oral epithelial cells or gonad tissue of patients with DSD and with XYY in blood. Among 4,437 DSD males who received cytogenetic evaluation, 14 patients with 47,XYY were identified. By contrast, five individuals among the 6,259 controls had 47,XYY. XYY in blood is more frequent among males with DSD than in other males (p = 0.004). The XYY karyotypes were confirmed again by GTG-banding in blood samples and by FISH performed on oral epithelial cells. ES on seven XYY DSD patients was successfully performed, but results did not identify any pathogenic variant on 55 known DSD genes. Gonad biopsy (n = 3) revealed testicular dysplasia and true hermaphroditism. FISH of gonad tissues (n = 3) showed that all of the samples had mosaic for X/XY/XYY. This study is the first to investigate the relationship between XYY in blood and DSD. The knowledge that XYY is in the blood and in oral cells have X/XY/XYY mosaicism in gonadal tissue is new for both researchers and clinicians who seek to understand the genetic basis of DSD males.

The oral microbiome of pregnant women facilitates gestational diabetes discrimination.

The oral microbiota plays an important role in the development of various diseases, whereas its association with gestational diabetes mellitus (GDM) remains largely unclear. The aim of this study is to identify biomarkers from the oral microbiota of GDM patients by analyzing the microbiome of the saliva and dental plaque samples of 111 pregnant women. We find that the microbiota of both types of oral samples in GDM patients exhibits differences and significantly varies from that of patients with periodontitis or dental caries. Using bacterial biomarkers from the oral microbiota, GDM classification models based on support vector machine and random forest algorithms are constructed. The area under curve (AUC) value of the classification model constructed by combination of Lautropia and Neisseria in dental plaque and Streptococcus in saliva reaches 0.83, and the value achieves a maximum value of 0.89 by adding clinical features. These findings suggest that certain bacteria in either saliva or dental plaque can effectively distinguish women with GDM from healthy pregnant women, which provides evidence of oral microbiome as an informative source for developing noninvasive biomarkers of GDM.

FOXD3-AS1 Knockdown Suppresses Hypoxia-Induced Cardiomyocyte Injury by Increasing Cell Survival and Inhibiting Apoptosis via Upregulating Cardioprotective Molecule miR-150-5p In Vitro.

Congenital heart disease (CHD) is the most common type of human innate malformation in fetuses. LncRNAs have been pointed to play critical regulatory roles in various types of cardiac development and diseases including CHD. Our study aimed to explore the effects of lncRNA forkhead box D3 antisense RNA 1 (FOXD3-AS1) on hypoxia-induced injury in AC16 cardiomyocytes and the related molecular mechanism. In vitro cell model of CHD was established by stimulating AC16 cells with hypoxia (1% O2). Expression of FOXD3-AS1 and miR-150-5p was detected by qRT-PCR. Hypoxia-induced injury was evaluated by detecting cell survival, lactate dehydrogenase (LDH) release, apoptosis, and caspase-3/7 activity using MTT, LDH assay, flow cytometry analysis, and caspase-3/7 activity assay, respectively. The regulatory relationship between FOXD3-AS1 and miR-150-5p was explored by luciferase reporter assay, RNA immunoprecipitation (RIP), and qRT-PCR. Results showed that hypoxia exposure caused an upregulation of FOXD3-AS1 and a downregulation of miR-150-5p in AC16 cells. Knockdown of FOXD3-AS1 attenuated reduction of cell survival and increase of LDH release, apoptosis, caspase-3/7 activity, and Bcl-2 associated X (Bax) expression induced by hypoxia in AC16 cells. Notably, we demonstrated that FOXD3-AS1 directly interacted with miR-150-5p to inhibit its expression. miR-150-5p knockdown reinforced the reduction of survival and induction of apoptosis by hypoxia and attenuated the effects of FOXD3-AS1 silencing on the same parameters in AC16 cells. In conclusion, FOXD3-AS1 knockdown protected AC16 cardiomyocytes from hypoxia-induced injury by increasing cell survival and inhibiting apoptosis through upregulating miR-150-5p.

Identification of Novel Compound Heterozygous Mutations in the GAN Gene of a Chinese Patient Diagnosed With Giant Axonal Neuropathy.

Giant axonal neuropathy (GAN) is a very rare autosomal recessive disorder characterized by abnormally large and dysfunctional neuronal axons. Mutations in the GAN gene have been identified as the cause of this disorder. In this report, we performed a detailed phenotypic assessment of a Chinese patient with GAN. An array-based exon capture test and targeted next-generation sequencing were used to detect the suspected mutation sites. Compound heterozygous mutations of p.S79L (c.236C > T) in the BTB domain and p.T489S (c.1466C > G) in the kelch domain were identified in the proband's genome. S79L was a known mutation, and T489S was reported for the first time. The p.S79L and p.T489S were confirmed in the proband's mother and father, respectively. Both mutations were located in highly conserved regions and affected the predicted protein crystal structures. The proband's sural biopsy revealed the classical GAN phenotype of swollen axons filled with closely packed neurofilaments. The combined application of the next-generation sequencing platform and bioinformatics analyses was an effective method for diagnosing GAN. The novel compound mutations of S79L and T489S in the GAN gene were likely the cause of the patient's GAN symptoms. Our findings enrich the spectrum of mutations associated with this rare type of axonopathy.

Maternal 25-Hydroxyvitamin D Deficiency Promoted Metabolic Syndrome and Downregulated Nrf2/CBR1 Pathway in Offspring.

Metabolic syndrome is a disorder of energy use and storage, which is characterized by central obesity, dyslipidemia, and raised blood pressure and blood sugar levels. Maternal 25-hydroxyvitamin D deficiency is known to cause metabolic changes, chronic disease, and increased adiposity in adulthood. However, the underlying mechanism of induced metabolic syndrome (MetS) in the offspring in vitamin D deficient pregnant mothers remains unclear. We identified that maternal 25-hydroxyvitamin D deficiency enhances oxidative stress, which leads to the development of MetS in the mother and her offspring. Further, immunohistochemical, Western blotting, and qRT-PCR analyses revealed that maternal 25-hydroxyvitamin D deficiency inhibited the activation of the Nrf2/carbonyl reductase 1 (CBR1) pathway in maternal placenta, liver, and pancreas, as well as the offspring's liver and pancreas. Further analyses uncovered that application of 25-hydroxyvitamin D activated the Nrf2/CBR1 pathway, relieving the oxidative stress in BRL cells, suggesting that 25-hydroxyvitamin D regulates oxidative stress in offspring and induces the activation of the Nrf2/CBR1 pathway. Taken together, our study finds that maternal 25-hydroxyvitamin D deficiency is likely to result in offspring's MetS probably via abnormal nutrition transformation across placenta. Depression of the Nrf2/CBR1 pathway in both mothers and their offspring is one of the causes of oxidative stress leading to MetS. This study suggests that 25-hydroxyvitamin D treatment may relieve the offspring's MetS.

The Effect of Mitomycin C on Reducing Endometrial Fibrosis for Intrauterine Adhesion.

BACKGROUND Intrauterine adhesion (IUA) is a common reproductive system disease in women, characterized by endometrial stromal cell proliferation, increasing fibroblasts and increasing extracellular matrix secretion. The purpose of this study was to investigate the effect of mitomycin C on reducing endometrial fibrosis for IUA. MATERIAL AND METHODS Firstly, a rat IUA model was constructed by intrauterine mechanical injury. The endometrial stromal cells and fibroblasts were isolated and treated with mitomycin C. After that, Cell Counting Kit-8 (CCK-8) assay was used to investigate the endometrial stromal cell viability. Furthermore, cell cycle and apoptosis assays of endometrial stromal cells and fibroblasts were performed, respectively. Finally, the cell viability of human endometrial cells or human uterus adhesion fibroblasts treated with mitomycin C was determined using CCK-8 assay with or without estradiol. RESULTS Endometrial stromal cells were isolated from a rat IUA model. Cell cycle assay results showed that mitomycin C inhibited cell viability and promoted G1 cell cycle arrest and apoptosis in rat IUA endometrial stromal cells. Fibroblasts were also isolated from the rat IUA model. We found that mitomycin C inhibited the synthesis and secretion of collagen type I by western blotting analysis. Furthermore, mitomycin C promoted G1 cell cycle arrest and apoptosis in IUA rat uterine fibroblasts. We found that estradiol decreased the inhibitory effects of cell viability of human endometrial cells and human uterus adhesion fibroblasts by mitomycin C. CONCLUSIONS Our findings revealed that mitomycin C could reduce endometrial fibrosis for intrauterine adhesion.

Evaluation of ovarian function using three dimensional ultrasound in perimenopausal women.

To evaluate the feasibility and clinical value of three-dimensional ultrasound in evaluating ovarian function in perimenopausal women. In this prospective cohort study, 102 patients with clinically suspected perimenopause and 90 patients with menopause were enrolled. These patients were classified into three groups according to the level of follicle stimulating hormone (FSH) and estradiol (E2): menopause group, perimenopause group, and normal group. Perimenopause group: There were significant differences in volume, vascularization index (VI), flow index (FI), and vascularization-flow index (VFI) in the ovaries after treatment. Cycle 1 > cycle 0 (p < .05) and cycle 3 cycle 0 (p < .05), and in FSH: cycle 3 < cycle 0 (p < .05). Three-dimensional ultrasound in ovarian quantitative measurement can objectively reflect the change in the ovarian function, predicting the effect of drug treatment, and provided an objective information for early intervention to menopausal.

Comparison of Efficiencies of Non-invasive Prenatal Testing, Karyotyping, and Chromosomal Micro-Array for Diagnosing Fetal Chromosomal Anomalies in the Second and Third Trimesters.

In this study, we aimed to compare the efficiency of non-invasive prenatal testing (NIPT), karyotyping, and chromosomal micro-array (CMA) for the diagnosis of fetal chromosomal anomalies in the second and third trimesters. Pregnant women, who underwent amniocenteses for prenatal genetic diagnoses during their middle and late trimesters, were recruited at the Prenatal Diagnosis Center of Taizhou City. Maternal blood was separated for NIPT, and amniotic fluid cells were cultured for karyotyping and CMA. The diagnostic efficiency of NIPT for detecting fetal imbalanced anomalies was compared with karyotyping and CMA. A total of 69 fetal chromosomal imbalances were confirmed by CMA, 37 were diagnosed by NIPT and 35 were found by karyotyping. The sensitivities of NIPT and karyotyping for diagnosing aneuploidy were 96.3% and 100% respectively. Only one mosaic sexual chromosome monosomy was misdiagnosed by NIPT, whereas the sensitivity of NIPT and karyotyping was 70% and 30%, respectively, for detecting pathogenic deletions and duplications sized from 5-20 Mb. Taken together, our results suggest that the efficiency of NIPT was similar to the formula karyotyping for detecting chromosome imbalance in the second and third trimesters.

Fibroblast growth factor 21 protects the heart from angiotensin II-induced cardiac hypertrophy and dysfunction via SIRT1.

AIMS: This study investigated the mechanism through which fibroblast growth factor 21 (FGF21) protects against angiotensin II (Ang II)-induced cardiac hypertrophy and dysfunction. METHODS: Male silent information regulator 1 (SIRT1) flox/flox and cardiomyocyte-specific inducible SIRT1 knockout mice (SIRT1-iKO) were generated and treated with Ang II (1.1 mg/kg/day for 4 weeks) at the age of 8-12-week-old. FGF21 treatment [2.5 mg/kg/day for 4 weeks by intraperitoneal (i.p.) injection] was initiated at the same time as the Ang II infusion. For in vitro studies, neonatal rat cardiomyocytes (NRCMs), H9c2 rat cardiomyocytes and isolated adult mouse cardiomyocytes were treated with Ang II (1 µM) and FGF21 (20 nM) for 24 h with or without SIRT1 silencing. RESULTS: FGF21 treatment significantly attenuated Ang II-induced cardiac hypertrophy and dysfunction. SIRT1 knockout abolished the ability of FGF21 to prevent Ang II-induced cardiac hypertrophy, fibrosis, and apoptosis, without affecting the beneficial effects of FGF21 in Ang II-induced hypertension, and did not influence the hypertension itself. FGF21 markedly increased the deacetylase activity of SIRT1 and promoted the interaction of SIRT1 with liver kinase B1 (LKB1) and forkhead box protein O1 (FoxO1), resulting in decreased acetylation of these SIRT1 target proteins. Consequently, FGF21 promoted the activation of the LKB1 target adenosine monophosphate-activated protein kinase (AMPK) and altered the transcriptional activity of FoxO1 on its downstream target genes catalase (Cat), MnSOD (Sod2), and Bim, resulting in reduced reactive oxygen species (ROS) accumulation and cardiomyocyte apoptosis. CONCLUSIONS: FGF21 improves cardiac function and alleviates Ang II-induced cardiac hypertrophy in a SIRT1-dependent manner.

miR-9 knockdown inhibits hypoxia-induced cardiomyocyte apoptosis by targeting Yap1.

AIMS: Aberrantly expressed miRNAs are demonstrated to be involved in the development of congenital heart disease (CHD). miR-9 was proposed to be upregulated in cardiac tissues from CHD cases. However, the role of miR-9 in hypoxia-induced cardiomyocytes and the potential mechanism are far from being addressed. MAIN METHODS: qRT-PCR and western blot analysis were performed to detect miR-9 and Yes-associated protein 1 (Yap1) expressions in hypoxic H9c2 cells. CCK-8, flow cytometry analysis, caspase-3/7 activity assay were applied to evaluate cell proliferation, apoptosis, and caspase-3/7 activity, respectively. The interaction between miR-9 and Yap1 was explored by luciferase reporter assay, qRT-PCR and western blot. KEY FINDINGS: miR-9 was upregulated and Yap1 was downregulated in H9c2 cells in response to hypoxia in a time-dependent manner. Knockdown of miR-9 promoted cell proliferation, and inhibited apoptosis and caspase-3/7 activity in hypoxic H9c2 cells, while miR-9 overexpression exerted the opposite effects on hypoxic H9c2 cells. In addition, Yap1 was a direct target of miR-9 in H9c2 cells. Yap1 knockdown suppressed cell proliferation and promoted apoptosis in hypoxia-exposed H9c2 cells. Yap1 knockdown attenuated the effect of anti-miR-9 on cell proliferation and apoptosis in hypoxia-exposed H9c2 cells. SIGNIFICANCE: miR-9 knockdown inhibited hypoxia-induced cardiomyocyte apoptosis by targeting Yap1. Our study provided a novel insight into the mechanism of the adaptation of cardiomyocytes to chronic hypoxia.

Short-Term and Mid-Term Results of Minimally Invasive Occlusion of Ventricular Septal Defects via a Subaxillary Approach in a Single Center.

This retrospective, single-center study evaluated short-term and mid-term results of minimally invasive surgery to occlude ventricular septal defects (VSDs) using a subaxillary approach. The procedure was performed on 429 children (224 boys, 205 girls; age 2.4 ± 2.5 years; mean weight 12.7 ± 10.1 kg) between January 2014 and December 2016 at the Children's Heart Center of Henan Province People's Hospital. An approximately 2-cm subaxillary incision was made between the third and fifth ribs, and the appropriate right atrium or ventricle was punctured under the guidance of transencephalographic echocardiography (TEE). The VSD was then occluded under TEE guidance. The mean size of the VSDs was 4.2 ± 1.0 mm, and the occluder measured 5.3 ± 1.3 mm. Asymmetrical occluders were used in 44 patients and symmetrical occluders in 385 patients. The operative time was 60.7 ± 21.3 min, and time in the intensive care unit was 20.9 ± 6.5 h. Blood loss was 12.4 ± 14.4 ml. There were no deaths among these patients. Occluder displacement occurred in two cases. There were no complications (e.g., third-degree atrioventricular block, new aortic regurgitation, reoperation for massive bleeding, serious infection). All patients were followed for 6-48 months, during which time there were ten cases of a postoperative residual shunt, which self-closed in eight during follow-up. The other two cases are still being followed. No complications occurred during follow-up (e.g., reoperation, aortic regurgitation, atrioventricular block, occluder abscission). Occluding VSDs using the subaxillary approach is safe and effective. Short-term and mid-term results are satisfactory. Further follow-up is required regarding long-term results.

[Association of coagulation factor V gene polymorphism with unexplained recurrent spontaneous abortion among ethnic Hans from Wenzhou area].

OBJECTIVE: To assess the association of coagulation factor V gene polymorphisms with unexplained recurrent spontaneous abortion (URSA) among ethnic Han Chinese from Wenzhou area. METHODS: Ninety-six patients with URSA and 103 females with a history of normal pregnancy were recruited. Genotypes of coagulation factor V gene were determined through target sequence capture and high-throughput sequencing. The results were confirmed with a MassARRAY system. Allelic and genotypic frequencies between the two groups were compared. RESULTS: Nineteen single nucleotide polymorphism (SNPs), except coagulation factor V Leiden, were identified in the two groups. The frequencies of rs9287090 allele A, rs1046712 allele T and rs1800594 allele G of the URSA group were lower than those of the control group (6.77% vs. 16.50%, 3.12% vs. 13.11%, 10.94% vs. 18.45%, respectively). After Bonferroni and false discovery rate correction, rs9287090 and rs1046712 were significantly associated with URSA (corrected P<0.05). Although genotypic distribution of rs9287090 and rs1046712 also differed between the two groups, the corrected P value showed no significance (corrected P>0.05). A complete linkage disequilibrium (r2=1, D'=1) of rs6022 and rs6029 was observed for the haplotype block rs6022-rs6029-rs6028. The frequencies of rs6022 allele A and rs6029 allele T were higher in the URSA group with corrected insignificance (75.00% vs. 65.53%, corrected P>0.05). Furthermore, significantly more A-T-T haplotype was found in the URSA group (75.00% vs. 65.50%, OR=1.578, 95%CI:1.021-2.438, χ2=4.248, P<0.05). CONCLUSION: The decreased rate of rs9287090 allele A, rs1046712 allele T, and rs1800594 allele G may contribute to the susceptibility to URSA among ethnic Han Chinese from Wenzhou area. The rs6022 allele A and rs6029 allele T may also predispose to URSA.

Dysbiosis of maternal and neonatal microbiota associated with gestational diabetes mellitus.

OBJECTIVE: The initial colonisation of the human microbiota and the impact of maternal health on neonatal microbiota at birth remain largely unknown. The aim of our study is to investigate the possible dysbiosis of maternal and neonatal microbiota associated with gestational diabetes mellitus (GDM) and to estimate the potential risks of the microbial shift to neonates. DESIGN: Pregnant women and neonates suffering from GDM were enrolled and 581 maternal (oral, intestinal and vaginal) and 248 neonatal (oral, pharyngeal, meconium and amniotic fluid) samples were collected. To avoid vaginal bacteria contaminations, the included neonates were predominantly delivered by C-section, with their samples collected within seconds of delivery. RESULTS: Numerous and diverse bacterial taxa were identified from the neonatal samples, and the samples from different neonatal body sites were grouped into distinct clusters. The microbiota of pregnant women and neonates was remarkably altered in GDM, with a strong correlation between certain discriminatory bacteria and the oral glucose tolerance test. Microbes varying by the same trend across the maternal and neonatal microbiota were observed, revealing the intergenerational concordance of microbial variation associated with GDM. Furthermore, lower evenness but more depletion of KEGG orthologues and higher abundance of some viruses (eg, herpesvirus and mastadenovirus) were observed in the meconium microbiota of neonates associated with GDM. CONCLUSION: GDM can alter the microbiota of both pregnant women and neonates at birth, which sheds light on another form of inheritance and highlights the importance of understanding the formation of early-life microbiome.

A novel factor X mutation Cys81 by Arg and a reported factor VII polymorphism Arg353 replaced by Gln co-occured in a patient.

: Coagulation factor X and factor VII (FVII) are both very important components in blood coagulation. To study the molecular pathogenic mechanism of the inherited factor X and FVII deficiency, the factor X activity (FX:C) and FVII activity were tested with one-stage clotting methods. The factor X antigen and factor FVII antigen were tested with ELISA. All the exons, intron-exon boundaries and 5',3'-flanking regions of F10 and F7 genes were amplified by PCR with direct sequencing. The ClustalX software was used to analyze the conservative property of the mutation sites. The PolyPhen-2 and Sorting Intolerant From Tolerant (SIFT) online bioinformatics softwares were taken to predict whether the point mutation could affect protein function. The software Swiss-pdb Viewer was brought to analyze the impact of mutations on the structure and function of the protein. The thrombin generation tests were applied to evaluate whether there were obstacles in producing thrombin about the mutant protein. The FX:C and FVII activity of the proband were reduced to 35 and 42%, and the factor X antigen and factor FVII antigen were decreased to 43 and 55%, simultaneously. Correspondingly, a FX:Cys81Arg (Cys81 by Arg) mutation and a FVII:Arg353 replaced by Gln polymorphism were detected in the proband. The Cys81 of factor X was conserved among homologous species, but the Arg353 of FVII was not. All softwares analysis results indicated protein features and structures might be affected by the mutation and the polymorphism. And the thrombin generation tests showed that the mutant protein had obstacles in thrombin generation. We identified a FX:Cys81Arg mutation and a FVII:Arg353 replaced by Gln polymorphism in the proband. And they accounted for the decrease of the activity and antigen of factor X and FVII. Of note, the Cys81Arg of factor X was first reported in the world.

CRISPR/Cas9-loxP-Mediated Gene Editing as a Novel Site-Specific Genetic Manipulation Tool.

Cre-loxP, as one of the site-specific genetic manipulation tools, offers a method to study the spatial and temporal regulation of gene expression/inactivation in order to decipher gene function. CRISPR/Cas9-mediated targeted genome engineering technologies are sparking a new revolution in biological research. Whether the traditional site-specific genetic manipulation tool and CRISPR/Cas9 could be combined to create a novel genetic tool for highly specific gene editing is not clear. Here, we successfully generated a CRISPR/Cas9-loxP system to perform gene editing in human cells, providing the proof of principle that these two technologies can be used together for the first time. We also showed that distinct non-homologous end-joining (NHEJ) patterns from CRISPR/Cas9-mediated gene editing of the targeting sequence locates at the level of plasmids (episomal) and chromosomes. Specially, the CRISPR/Cas9-mediated NHEJ pattern in the nuclear genome favors deletions (64%-68% at the human AAVS1 locus versus 4%-28% plasmid DNA). CRISPR/Cas9-loxP, a novel site-specific genetic manipulation tool, offers a platform for the dissection of gene function and molecular insights into DNA-repair pathways.