Sex Differences in Attitudes Toward Marriage and Childbearing Based on the Assumption of Being BRCA1/2 Mutation Carriers Among Young People.

PURPOSE: This study investigated changes in attitudes toward marriage and childbearing assuming a BRCA1/2 mutation carrier status among healthy, unmarried individuals in Korea. METHODS: A nationally representative sample of healthy, unmarried individuals aged 20-39 years was surveyed. A questionnaire on marriage and childbearing intentions was administered to the participants before and after providing them with information on BRCA1/2 mutation carriers' breast and ovarian cancer risks and their autosomal dominant inheritance pattern. The participants were asked about their attitudes toward childbearing through preimplantation genetic diagnosis (PGD). RESULTS: Of the participants who initially wanted to marry, the assumption that they or their partners had BRCA1/2 mutation caused 25.3% to no longer want to get married and 36.2% to change their attitude from wanting to bear children to no longer wanting them. Females were more likely than males to change their attitudes toward marriage and childbearing. The participants who had negative attitudes toward genetic testing were more likely to change their attitudes regarding marriage and childbearing than those who were favorable toward both disclosure and testing. More than 50% of the participants who did not want children were willing to bear children through PGD when it was assumed that they were BRCA mutation carriers. CONCLUSION: On the assumption of being carriers, general, young, and healthy females were more likely than males to negatively change their attitudes toward marriage and childbearing. Public education on the implications of living with mutation carriers and reproductive options may be required.

Attitudes toward Risk-Reducing Mastectomy and Risk-Reducing Salpingo-oophorectomy among Young, Unmarried, Healthy Women in Korea.

PURPOSE: This study investigated the attitudes toward risk-reducing mastectomy (RRM) and risk-reducing salpingo-oophorectomy (RRSO) as cancer prevention options for BRCA1/2 carriers in healthy, young, unmarried Korean women. MATERIALS AND METHODS: A nationally representative sample of 600 women, aged 20-39 years, completed a questionnaire on sociodemographic variables, preference for genetic testing, and intention to undergo risk-reducing surgeries after receiving information on the cancer risk of BRCA1/2 mutations and benefits of risk-reducing surgeries. RESULTS: A total of 54.7% and 57.7% had the intention to undergo RRM and RRSO, respectively, on the assumption that they were BRCA1/2 carriers. Older age and no intention to undergo genetic testing were associated with a reduced likelihood of undergoing RRM (odds ratio [OR], 0.30; 95% confidence interval [CI], 0.14 to 0.61 for age 35-39 years and OR, 0.35; 95% CI, 0.20 to 0.62 for no intention for genetic testing) and RRSO (OR, 0.39; 95% CI, 0.19 to 0.79 for age 35-39 years and OR, 0.30; 95% CI, 0.17 to 0.53 for no intention for genetic testing). Women who chose to be single were likely to undergo risk-reducing surgeries (OR, 1.67; 95% CI, 1.07 to 2.60 for RRM and OR, 1.56; 95% CI, 1.00 to 2.44 for RRSO). CONCLUSION: More than 50% of healthy, unmarried, young Korean women were inclined to undergo prophylactic surgeries if they were BRCA1/2 mutation carriers. Further studies on decision-making process for cancer prevention in individuals at high risk for cancer need to be conducted.

Epigenome-wide base-resolution profiling of DNA methylation in chorionic villi of fetuses with Down syndrome by methyl-capture sequencing.

BACKGROUND: Epigenetic mechanisms provide an interface between environmental factors and the genome and are influential in various diseases. These mechanisms, including DNA methylation, influence the regulation of development, differentiation, and establishment of cellular identity. Here, we performed high-throughput methylome profiling to determine whether differential patterns of DNA methylation correlate with Down syndrome (DS). MATERIALS AND METHODS: We extracted DNA from the chorionic villi cells of five normal and five DS fetuses at the early developmental stage (12-13 weeks of gestation). Methyl-capture sequencing (MC-Seq) was used to investigate the methylation levels of CpG sites distributed across the whole genome to identify differentially methylated CpG sites (DMCs) and regions (DMRs) in DS. New functional annotations of DMR genes using bioinformatics tools were predicted. RESULTS: DNA hypermethylation was observed in DS fetal chorionic villi cells. Significant differences were evident for 4,439 DMCs, including hypermethylation (n = 4,261) and hypomethylation (n = 178). Among them, 140 hypermethylated DMRs and only 1 hypomethylated DMR were located on 121 genes and 1 gene, respectively. One hundred twenty-two genes, including 141 DMRs, were associated with heart morphogenesis and development of the ear, thyroid gland, and nervous systems. The genes were significantly associated with DS and various diseases, including hepatopulmonary syndrome, conductive hearing loss, holoprosencephaly, heart diseases, glaucoma, and musculoskeletal abnormalities. CONCLUSIONS: This is the first study to compare the whole-epigenome DNA methylation pattern of the chorionic villi cells from normal and DS fetuses at the early developmental-stage using MC-seq. Overall, our results indicate that the chorionic villi cells of DS fetuses are hypermethylated in all autosomes and suggested that altered DNA methylation may be a recurrent and functionally relevant downstream response to DS in human cells. This study provides basic information for future research focused on the pathophysiology of the DS and its potential effects, as well as the role DNA methylation plays in the early developmental stage of DS fetuses.

Effective Fetal Epigenetic Biomarkers for Noninvasive Fetal Trisomy 21 Detections.

INTRODUCTION: Recently, we identified three novel fetal-specific epigenetic DNA regions (FSERs) on chromosome 21 for detection of noninvasive fetal trisomy 21 (T21). In this study, the diagnostic accuracies of the three FSERs were assessed on a larger panel of the first-trimester pregnant women. MATERIAL AND METHODS: This study was conducted with maternal plasma collected from 167 pregnant women carrying 155 chromosomally normal and 12 T21 fetuses (10-13 gestational weeks). Accuracies of FSERs for noninvasive prenatal test of fetal T21 were estimated by the area under the receiver operator characteristic curve (AUC). RESULTS: The levels of all FSERs increased in pregnant women with T21 fetuses when compared with controls (p < 0.001 for all). The levels of the three FSERs did not differ according to maternal age, body mass index, and fetal sex at maternal blood sampling (p > 0.05 for all). In noninvasive fetal T21 detection, the AUC of FSER1, FSER2, and FSER3 were 0.859 (95% CI: 0.746-0.972), 0.919 (95% CI: 0.856-0.982), and 0.868 (95% CI: 0.746-0.990), respectively. DISCUSSION: The findings of this study suggest that all FSERs may be useful for noninvasive fetal T21 detection, regardless of maternal age, body mass index, and fetal sex.

Evaluation of extraction methods for methylated cell-free fetal DNA from maternal plasma.

PURPOSE: Recently, fetal placenta-specific epigenetic regions (FSERs) have been identified for quantification of cell-free fetal DNA (cff-DNA) for non-invasive prenatal testing (NIPT). The aim of this study was to evaluate the efficiencies of a column-based kit and magnetic bead-based kit for quantification of methylated FSERs from maternal plasma. METHODS: Maternal plasma was extracted from normal pregnant women within the gestational age of 10~13 weeks (n = 24). Total cell-free DNA (cf-DNA) was extracted using a column-based kit and magnetic bead-based kit from the plasma of the same pregnant woman, respectively. Methylated FSERs were enriched from the extracted total cf-DNA using a methyl-CpG-binding domain-based protein method. The four FSERs were simultaneously quantified by multiplex real-time polymerase chain reaction. RESULTS: Methylated FSERs were detected in all samples extracted from both kits. However, the amplification of FSERs showed significant differences in the extraction efficiency of methylated FSERs between the two extraction methods. The Ct values of methylated FSERs extracted using the column-based kit were significantly lower than those obtained using the magnetic bead-based kit (P < 0.001 for all FSERs). The quantity of methylated FSERs was significantly higher for extracted DNA using the column-based kit than that extracted using the magnetic bead-based kit (P < 0.001 for all FSERs). Time and cost for the process of extraction were similar for the column kit and magnetic bead-based kit. CONCLUSIONS: Our findings demonstrate that the column-based kit was more effective than the magnetic bead-based kit for isolation of methylated FSERs from maternal plasma as assessed by FSER detection.

Y Chromosome Microdeletions in Infertile Men with Non-obstructive Azoospermia and Severe Oligozoospermia.

BACKGROUND: The purpose of the study was to investigate the frequencies and types of Y chromosome microdeletions in infertile men and to analyze the relationship between the levels of reproductive hormones and Y microdeletions. METHODS: A total of 1,226 infertile men were screened for Y chromosome microdeletions using multiplex PCR assay. Karyotype analysis was performed on peripheral blood lymphocytes with standard G-banding. Serum reproductive hormone levels were measured. RESULTS: Out of 1,226 infertile patients, 134 (10.93%) had Y microdeletions. One hundred seven of 765 (13.99%) non-obstructive azoospermic patients and 27 of 133 (20.30%) severe oligozoospermic patients had Y microdeletions. Among the 134 infertile men with Y microdeletions, the most frequent microdeletions were detected in the AZFc region, followed by AZFbc, AZFb, AZFa, AZFabc(Yq), Yp(SRY)+Yq, and partial AZFc regions. Karyotype analysis was available for 130 of the 134 patients with Y microdeletions. Of them, 36 (27.69%) patients had sex chromosomal abnormalities. Levels of FSH and LH in patients with AZFc microdeletion were significantly lower, while those in patients with Yp(SRY)+Yq were significantly higher than in patients without Y microdeletions. Level of testosterone in patients with AZFabc(Yq) or Yp(SRY)+Yq was significantly lower than that in patients without Y microdeletions. However, there was no significant difference in the levels of reproductive hormones between all patients with and without Y microdeletions. CONCLUSION: These results highlight the need for Y chromosome microdeletion screening for correct diagnosis of male infertility. Obtaining reliable genetic information for assisted reproductive techniques can prevent unnecessary treatment and vertical transmission of genetic defects to offspring.

Bioinspired Dual Stimuli-Responsive Membranous System with Multiple On-Off Gates.

Stimuli-responsive polymers have been widely used for controlled release of several biomolecules. In general, a single stimulus among various stimuli, for instance, temperature, pH, or light, has been used for these polymers. Although some stimuli are applied together, one cannot control each stimulus independently at a given stimulus-responsive polymer. However, to mimic biological system like cell membrane, multiple on-off gates utilizing independent control of dual (or multiple) stimuli should be used. Here, we introduce a stimuli-responsive membrane controlled by two orthogonal stimuli. For this purpose, the top and the bottom parts of anodized aluminum oxide membrane walls are independently grafted by thermoresponsive poly(N-isopropylacrylamide) and pH-responsive poly(acrylic acid), respectively, by using surface-initiated atom transfer radical polymerization. The membrane clearly showed two independent on-off gates depending on temperature and pH. Furthermore, through light irradiation of two different wavelengths (near-infrared and ultraviolet), temperature and pH were also controlled independently and promptly. Thus, this membrane shows two independent on-off gating of the transport of a model biomolecule of fluorescein isothiocyanate-labeled bovine serum albumin. This strategy suggests the potential of independently modified membrane in layers as stimuli-responsive on-off gates for the application of artificial cell membrane.

Multi-floor cascading ferroelectric nanostructures: multiple data writing-based multi-level non-volatile memory devices.

Multiple data writing-based multi-level non-volatile memory has gained strong attention for next-generation memory devices to quickly accommodate an extremely large number of data bits because it is capable of storing multiple data bits in a single memory cell at once. However, all previously reported devices have failed to store a large number of data bits due to the macroscale cell size and have not allowed fast access to the stored data due to slow single data writing. Here, we introduce a novel three-dimensional multi-floor cascading polymeric ferroelectric nanostructure, successfully operating as an individual cell. In one cell, each floor has its own piezoresponse and the piezoresponse of one floor can be modulated by the bias voltage applied to the other floor, which means simultaneously written data bits in both floors can be identified. This could achieve multi-level memory through a multiple data writing process.

Changes in the methylation status of DAT, SERT, and MeCP2 gene promoters in the blood cell in families exposed to alcohol during the periconceptional period.

BACKGROUND: Alcohol exposure has been shown to cause devastating effects on neurobehavioral development in numerous animal and human studies. The alteration of DNA methylation levels in gene-specific promoter regions has been investigated in some studies of human alcoholics. This study was aimed to investigate whether social alcohol consumption during periconceptional period is associated with epigenetic alteration and its generational transmission in the blood cells. METHODS: We investigated patterns of alcohol intake in a prospective cohort of 355 pairs of pregnant women and their spouses who reported alcohol intake during the periconceptional period. A subpopulation of 164 families was established for the epigenetic study based on the availability of peripheral blood and cord blood DNA. The relative methylation changes of dopamine transporter (DAT), serotonin transporter (SERT), and methyl CpG binding protein 2 (MeCP2) gene promoters were analyzed using methylation-specific endonuclease digestion followed by quantitative real-time polymerase chain reaction. RESULTS: The relative methylation level of the DAT gene promoter was decreased in the group of mothers reporting above moderate drinking (p = 0.029) and binge drinking (p = 0.037) during pregnancy. The relative methylation level of the DAT promoter was decreased in the group of fathers reporting heavy binge drinking (p = 0.003). The relative methylation levels of the SERT gene promoter were decreased in the group of newborns of light drinking mothers before pregnancy (p = 0.012) and during pregnancy (p = 0.003). The methylation level in the MeCP2 promoter region of babies whose mothers reported above moderate drinking during pregnancy was increased (p = 0.02). In addition, methylation pattern in the DAT promoter region of babies whose fathers reported heavy binge drinking was decreased (p = 0.049). CONCLUSIONS: These findings suggest that periconceptional alcohol intake may cause epigenetic changes in specific locus of parental and newborn genomes as follows: Alcohol consumption decreases the methylation level of the DAT promoter region of the parent themselves, maternal alcohol drinking during the periconceptional period decreases the methylation level of the SERT promoter region of newborns, and maternal alcohol consumption increases the methylation level of the MeCP2 promoter region of newborns.

Non-invasive detection of fetal trisomy 21 using fetal epigenetic biomarkers with a high CpG density.

BACKGROUND: Non-invasive prenatal test of trisomy 21 (T21) is being researched using fetal specific epigenetic biomarkers present in maternal plasma. We applied a methyl-CpG binding domain-based protein (MBD) method based on epigenetic characteristics of fetal specific-methylated regions with a high CpG density in HLCS on chromosome 21 and RASSF1A on chromosome 3 for the non-invasive detection of fetal T21 and estimated the diagnostic accuracy of the method. METHODS: A nested case-control study was conducted with maternal plasma collected from 50 pregnant women carrying 40 normal and 10 T21 fetuses. A MBD method was used for enrichment of methylated DNA regions in maternal plasma. The levels of methylated HLCS (M-HLCS) and methylated RASSF1A (M-RASSF1A) were simultaneously measured by multiplex qPCR. RESULTS: Levels of M-HLCS and M-RASSF1A were obtained in all cases. Levels were not different according to fetal gender (p>0.05 in both). The level of M-HLCS was significantly increased in women with a T21 fetus compared with controls (p<0.001). The level of M-RASSF1A was not different between two groups (p>0.05). In non-invasive fetal T21 detection, the specificity of M-HLCS level and the epigenetic-epigenetic ratio (EER) using M-HLCS and M-RASSF1A levels were 82.5% and 92.5%, respectively, at 90.0% sensitivity. CONCLUSIONS: Our findings suggest that the EER may be useful as a potential biomarker for the non-invasive detection of fetal T21, regardless of fetal gender. The MBD method can be used as an effective tool in the detection of methylated fetal specific markers with a high CpG density in maternal plasma.

Hierarchically self-organized monolithic nanoporous membrane for excellent virus enrichment.

Enrichment of viruses is essential for making high dose viral stocks for vaccines and virus-related research. Since the widely used ultracentrifugation for concentrating viral stock requires ultra-high speed rotation, it easily destroys the activity of some viruses, for instance, hepatitis c virus (HCV), which has a fragile structure and low virus titer. We introduce a novel method to concentrate HCV virus in stock by using a hierarchically self-organized monolithic nanoporous membrane made by stepwise anodization. The pores at the top part of the membrane have very regular sizes that are suitable for the perfect filtration of the virus particles in the stock. On the other hand, the remaining part has large pores that maintain high flux and mechanical strength of the membrane under the high pressure (up to 10 bar). The enrichment efficiency of HCV in crude stocks by using the membrane became over 91%, which is four times higher than that (∼22%) obtained by conventionally used centrifugation. A very high efficiency results from the perfect filtration and no damage to the virion particles during the enrichment process, whereas significant damage to the HCV occurs during centrifugation. The hierarchically self-organized monolithic nanoporous membrane could be effectively employed for concentrating various fragile viruses in stocks, for instance, rabies virus and human immunodeficiency virus in addition to HCV virus.

Preeclampsia is associated with an elevation of plasma sMet concentrations in the second trimester.

OBJECTIVE: The aim of this study was to investigate the association between anti-angiogenic factor soluble c-Met (sMet) concentrations in maternal plasma and the risk of preeclampsia. METHODS: The pregnant women included in this study (1) had subsequent preeclampsia (n = 52) and were compared to normal controls (n = 104) at the time of amniocentesis (15-20 weeks); and (2) had preeclampsia (n = 63) and were compared to normal controls (n = 112) at the time of diagnosis of preeclampsia (29-40 weeks). sMet concentrations were measured by ELISA. Non-parametric statistics were used for analysis. RESULTS: Maternal plasma sMet concentrations were significantly higher in both women with subsequent preeclampsia (median: 1372.7 ng/ml versus 1100.5 ng/ml; p = 0.036) and women with preeclampsia (median: 1651.9 ng/ml versus 1364.7 ng/ml; p < 0.001) than in the controls. After adjusting for potential confounding factors, the risks of developing preeclampsia were as follows: adjusted odds ratio 2.5 (95% confidence interval, 1.2-5.2; p = 0.016) for second trimester sMet concentration with a cut-off value of 1223.5 ng/ml and 4.4 (95% confidence interval, 2.2-9.1; p < 0.001) for third trimester sMet concentration with a cut-off value of 1460.3 ng/ml. CONCLUSION: Elevated maternal plasma sMet concentrations were independently associated with the increased risk of preeclampsia.

Determination of the carrier frequencies of selected GJB2 mutations in the Korean population.

OBJECTIVE: Mutations in the GJB2 gene are a major cause of hereditary hearing loss. However, only a few studies have investigated carrier frequencies of GJB2 mutations in the general population. The aim of this study was to estimate the carrier frequencies of three GJB2 mutations, including 235delC, V37I, and G45E, in the general Korean population. DESIGN: A standard questionnaire of self-reported hearing loss was used to identify and recruit subjects. Screening for three mutations was performed using an allele-specific polymerase chain reaction (PCR), PCR-restriction fragment length polymorphism, and direct DNA sequencing. STUDY SAMPLE: A total of 1256 unrelated healthy individuals were analysed in the present study. RESULTS: Of the 1256 individuals, 24 had GJB2 mutations; 11 were found to be heterozygous for 235delC, 11 were heterozygous and one was homozygous for V37I, and one was heterozygous for G45E. One individual had a compound heterozygous mutation of 235delC/V37I. The allele frequencies of 235delC, V37I, and G45E mutations were 0.44%, 0.52%, and 0.04%, respectively. The carrier frequency of either the 235delC or V37I mutation was estimated to be 0.88% with 95% binomial CI, 0.44-1.56. CONCLUSIONS: These results will facilitate diagnosis of, and genetic counseling for, hearing loss in Koreans.

Maternal serum and amniotic fluid inhibin A levels in women who subsequently develop severe preeclampsia.

The purpose of this study was to evaluate whether maternal serum (MS) and amniotic fluid (AF) inhibin A levels are elevated in patients who subsequently develop severe preecalmpsia, and to investigate the correlation between MS and AF inhibin A levels in the second trimester. The study included 40 patients who subsequently developed severe preecalmpsia and 80 normal pregnant women. Inhibin A levels in MS and AF were measured with enzyme-linked immunosorbent assay (ELISA). The MS and AF inhibin A levels in patients who developed severe preeclampsia were significantly higher than those in the control group (both for p<0.001). There was a positive correlation between MS and AF inhibin A levels in patients who developed severe preeclampsia (r=0.397, p=0.011), but not in the control group (r=0.185, p=0.126). The best cutoff values of MS and AF inhibin A levels for the prediction of severe preeclampsia were 427 pg/mL and 599 pg/mL, respectively; the estimated ORs that were associated with these cut-off values were 9.95 (95% CI 3.8-25.9, p<0.001) and 6.0 (95% CI 2.3-15.8, p<0.001). An elevated level of inhibin A in MS and AF at the time of second trimester amniocentesis may be a risk factor for the subsequent development of severe preeclampsia.

Rapid prenatal diagnosis of Down Syndrome using quantitative fluorescent PCR in uncultured amniocytes.

Rapid prenatal diagnosis of common chromosome aneuploidies have been successful through quantitative fluorescent PCR (QF-PCR) assays and small tandem repeat (STR) markers. The purpose of our study was to investigate the clinical feasibility for rapid prenatal detection of Down syndrome using the quantitative fluorescent PCR in uncultured amniocytes. DNA was extracted from uncultured amniotic fluid of normal karyotype (n=200) and of Down syndrome (n=21). It was amplified using QF-PCR with four STR markers located on chromosome 21. Among normal samples, the ranges of diallelic peaks for at least one STR marker were 1.0-1.3 for D21S11, 1.0-1.4 for D21S1411 and 1.0-1.5 for D21S1270. Down syndrome samples showed trisomic triallelic patterns or trisomic diallelic patterns. The sensitivity, specificity, and efficiency of the assay for detecting Down syndrome were 95.4%, 100%, and 99.5%, respectively. Rapid prenatal diagnosis of Down syndrome using QF-PCR is a reliable technique that aids clinical management of pregnancy.

De novo chromosomal aberrations in the fetus; genetic counseling and clinical outcome.

The aim of this study was to examine the incidence and clinical outcome of de novo chromosomal aberrations retrospectively and provide useful data for genetic counseling in the prenatal cytogenetic diagnosis. We found 17 cases of de novo chromosomal aberrations in 5501 cases of prenatal cytogenetic analysis and reviewed the karyotype, further study, medical records, fetal ultrasound findings and clinical outcomes. Out of the 17 de novo chromosomal aberrations, 5 had balanced reciprocal translocations and 12 had unbalanced translocations characterized as deletion, addition, or marker. In the case of the five balanced reciprocal translocations, 3 cases without abnormal ultrasound findings were carried to term after comprehensive genetic counseling. Neonates were phenotypically normal and clinical examinations were normal. Two cases with abnormal ultrasound findings were terminated therapeutically. Twelve cases of unbalanced translocations were terminated except one case with a mosaic marker chromosome. High resolution fetal ultrasound and detailed cytogenetic and molecular study will be adjunctive tools for predicting the karyotype/phenotype correlations of fetuses with de novo chromosomal aberrations, although they have limitation to find all phenotypic effects.