High-throughput analysis of candidate imprinted genes and allele-specific expression

In diploid human organisms, the ~20,000 genes are usually functional as two active copies or alleles. Exceptionally, some genes have only one active allele while the other is silenced. Two different groups of genes fall into this minor category; the genes that exhibit random monoallelic expression (e.g. odorant receptor genes and genes coding for immunoglobulins), and those genes exhibiting monoallelic expression in a parent-of-origin specific manner, named imprinted genes. At the outset of this study in October 2006, 56 genes in humans were known to be imprinted and 98 in mice, but the total number of imprinted genes in either species was unknown. I have used high-throughput allele-specific PCR assays to screen human term placental tissue samples for new imprinted genes. Hundreds of genes were tested either because they were predicted to be imprinted or because they were candidates for which the imprinted status was simply unknown in human term placenta. My results suggested that we are reaching saturation in the number of human placentally imprinted genes. I show that ZNF331 is imprinted in human placenta and is part of a primate lineagespecific imprinted locus showing differential methylation. My data also highlights that parental allelic specific expression is a continuum, from imprinted monoallelic expression to partial imprinting (i.e., one parental allele is slightly more (or less) expressed than the other). This continuum suggests a requirement to sequence the transcriptome of every human tissue at each different developmental stage exhaustively to assess genes for parent-of-origin specific expression and to clearly define what imprinting means. Most importantly whether ‘partial’ imprinting has functional significance or is just a part of the dynamic flux of gene expression. Specifically, my results call for thorough investigation of the ZNF331 locus in human development, physiology and parent-of-origin specific diseases

High-throughput analysis of candidate imprinted genes and allele-specific gene expression in the human term placenta.

BACKGROUND: Imprinted genes show expression from one parental allele only and are important for development and behaviour. This extreme mode of allelic imbalance has been described for approximately 56 human genes. Imprinting status is often disrupted in cancer and dysmorphic syndromes. More subtle variation of gene expression, that is not parent-of-origin specific, termed 'allele-specific gene expression' (ASE) is more common and may give rise to milder phenotypic differences. Using two allele-specific high-throughput technologies alongside bioinformatics predictions, normal term human placenta was screened to find new imprinted genes and to ascertain the extent of ASE in this tissue. RESULTS: Twenty-three family trios of placental cDNA, placental genomic DNA (gDNA) and gDNA from both parents were tested for 130 candidate genes with the Sequenom MassArray system. Six genes were found differentially expressed but none imprinted. The Illumina ASE BeadArray platform was then used to test 1536 SNPs in 932 genes. The array was enriched for the human orthologues of 124 mouse candidate genes from bioinformatics predictions and 10 human candidate imprinted genes from EST database mining. After quality control pruning, a total of 261 informative SNPs (214 genes) remained for analysis. Imprinting with maternal expression was demonstrated for the lymphocyte imprinted gene ZNF331 in human placenta. Two potential differentially methylated regions (DMRs) were found in the vicinity of ZNF331. None of the bioinformatically predicted candidates tested showed imprinting except for a skewed allelic expression in a parent-specific manner observed for PHACTR2, a neighbour of the imprinted PLAGL1 gene. ASE was detected for two or more individuals in 39 candidate genes (18%). CONCLUSIONS: Both Sequenom and Illumina assays were sensitive enough to study imprinting and strong allelic bias. Previous bioinformatics approaches were not predictive of new imprinted genes in the human term placenta. ZNF331 is imprinted in human term placenta and might be a new ubiquitously imprinted gene, part of a primate-specific locus. Demonstration of partial imprinting of PHACTR2 calls for re-evaluation of the allelic pattern of expression for the PHACTR2-PLAGL1 locus. ASE was common in human term placenta.RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are

Data analysis issues for allele-specific expression using Illumina's GoldenGate assay.

BACKGROUND: High-throughput measurement of allele-specific expression (ASE) is a relatively new and exciting application area for array-based technologies. In this paper, we explore several data sets which make use of Illumina's GoldenGate BeadArray technology to measure ASE. This platform exploits coding SNPs to obtain relative expression measurements for alleles at approximately 1500 positions in the genome. RESULTS: We analyze data from a mixture experiment where genomic DNA samples from pairs of individuals of known genotypes are pooled to create allelic imbalances at varying levels for the majority of SNPs on the array. We observe that GoldenGate has less sensitivity at detecting subtle allelic imbalances (around 1.3 fold) compared to extreme imbalances, and note the benefit of applying local background correction to the data. Analysis of data from a dye-swap control experiment allowed us to quantify dye-bias, which can be reduced considerably by careful normalization. The need to filter the data before carrying out further downstream analysis to remove non-responding probes, which show either weak, or non-specific signal for each allele, was also demonstrated. Throughout this paper, we find that a linear model analysis of the data from each SNP is a flexible modelling strategy that allows for testing of allelic imbalances in each sample when replicate hybridizations are available. CONCLUSIONS: Our analysis shows that local background correction carried out by Illumina's software, together with quantile normalization of the red and green channels within each array, provides optimal performance in terms of false positive rates. In addition, we strongly encourage intensity-based filtering to remove SNPs which only measure non-specific signal. We anticipate that a similar analysis strategy will prove useful when quantifying ASE on Illumina's higher density Infinium BeadChips.RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are

Risk factors associated with adverse fetal outcomes in pregnancies affected by Coronavirus disease 2019 (COVID-19): a secondary analysis of the WAPM study on COVID-19.

Objectives To evaluate the strength of association between maternal and pregnancy characteristics and the risk of adverse perinatal outcomes in pregnancies with laboratory confirmed COVID-19. Methods Secondary analysis of a multinational, cohort study on all consecutive pregnant women with laboratory-confirmed COVID-19 from February 1, 2020 to April 30, 2020 from 73 centers from 22 different countries. A confirmed case of COVID-19 was defined as a positive result on real-time reverse-transcriptase-polymerase-chain-reaction (RT-PCR) assay of nasal and pharyngeal swab specimens. The primary outcome was a composite adverse fetal outcome, defined as the presence of either abortion (pregnancy loss before 22 weeks of gestations), stillbirth (intrauterine fetal death after 22 weeks of gestation), neonatal death (death of a live-born infant within the first 28 days of life), and perinatal death (either stillbirth or neonatal death). Logistic regression analysis was performed to evaluate parameters independently associated with the primary outcome. Logistic regression was reported as odds ratio (OR) with 95% confidence interval (CI). Results Mean gestational age at diagnosis was 30.6+/-9.5 weeks, with 8.0% of women being diagnosed in the first, 22.2% in the second and 69.8% in the third trimester of pregnancy. There were six miscarriage (2.3%), six intrauterine device (IUD) (2.3) and 5 (2.0%) neonatal deaths, with an overall rate of perinatal death of 4.2% (11/265), thus resulting into 17 cases experiencing and 226 not experiencing composite adverse fetal outcome. Neither stillbirths nor neonatal deaths had congenital anomalies found at antenatal or postnatal evaluation. Furthermore, none of the cases experiencing IUD had signs of impending demise at arterial or venous Doppler. Neonatal deaths were all considered as prematurity-related adverse events. Of the 250 live-born neonates, one (0.4%) was found positive at RT-PCR pharyngeal swabs performed after delivery. The mother was tested positive during the third trimester of pregnancy. The newborn was asymptomatic and had negative RT-PCR test after 14 days of life. At logistic regression analysis, gestational age at diagnosis (OR: 0.85, 95% CI 0.8-0.9 per week increase; pPeer reviewe

Maternal and perinatal outcomes of pregnant women with SARS-CoV-2 infection.

OBJECTIVES: To evaluate the maternal and perinatal outcomes of pregnancies affected by SARS-CoV-2 infection. METHODS: This was a multinational retrospective cohort study including women with a singleton pregnancy and laboratory-confirmed SARS-CoV-2 infection, conducted in 72 centers in 22 different countries in Europe, the USA, South America, Asia and Australia, between 1 February 2020 and 30 April 2020. Confirmed SARS-CoV-2 infection was defined as a positive result on real-time reverse-transcription polymerase chain reaction (RT-PCR) assay of nasopharyngeal swab specimens. The primary outcome was a composite measure of maternal mortality and morbidity, including admission to the intensive care unit (ICU), use of mechanical ventilation and death. RESULTS: In total, 388 women with a singleton pregnancy tested positive for SARS-CoV-2 on RT-PCR of a nasopharyngeal swab and were included in the study. Composite adverse maternal outcome was observed in 47/388 (12.1%) women; 43 (11.1%) women were admitted to the ICU, 36 (9.3%) required mechanical ventilation and three (0.8%) died. Of the 388 women included in the study, 122 (31.4%) were still pregnant at the time of data analysis. Among the other 266 women, six (19.4% of the 31 women with first-trimester infection) had miscarriage, three (1.1%) had termination of pregnancy, six (2.3%) had stillbirth and 251 (94.4%) delivered a liveborn infant. The rate of preterm birth before 37 weeks' gestation was 26.3% (70/266). Of the 251 liveborn infants, 69/251 (27.5%) were admitted to the neonatal ICU, and there were five (2.0%) neonatal deaths. The overall rate of perinatal death was 4.1% (11/266). Only one (1/251, 0.4%) infant, born to a mother who tested positive during the third trimester, was found to be positive for SARS-CoV-2 on RT-PCR. CONCLUSIONS: SARS-CoV-2 infection in pregnant women is associated with a 0.8% rate of maternal mortality, but an 11.1% rate of admission to the ICU. The risk of vertical transmission seems to be negligible. © 2020 International Society of Ultrasound in Obstetrics and Gynecology

Le dépistage prénatal: l'exemple de la trisomie 21

Prenatal screening for Down's syndrome initially targeted high-risk pregnant women (> 35 years old). However, the vast majority of babies with Down's syndrome are born to younger women (as the majority of babies are born in this age category). It was first discovered that some serum analytes were altered in pregnancies affected with Down's syndrome (triple test). In the nineties, the association between an increased nuchal translucency measurement and trisomy 21 was noted. The use of this measurement in combination with serum markers has enabled an increased detection rate but still at the cost of a false positive rate of around five percent (combined test). Recently, major advances in sequencing technologies have allowed reasearchers to make use of the cell free fetal DNA in maternal blood. This new test (named non invasive prenatal test) made it into clinical use as early as 2011 in some countries. Its sensitivity is above 99 % for trisomy 21 and the false positive rate is very low. It is risk-free and much more accurate than previous approaches. It is largely favored over an invasive test by high risk women (advanced maternal age or high-risk combined test). Its use is still restricted by a high cost which is for the moment still entirely beared by the patient. The availability of NIPT in our routine practice and the increased complexity of screening options have highlighted the need for a more dedicated counselling consultation before Down's syndrome screening is performed.SCOPUS: ar.jinfo:eu-repo/semantics/publishe

L'accouchement à domicile

In Belgium, very few women give birth outside the delivery room. In the United Kingdom and in the Netherlands, they are more numerous. Several studies evaluated obstetric and neonatal outcomes of home births compared with hospital births. We selected seven recent and large studies (with cohorts of more than 5.000 women) using PubMed, Science Direct and Cochrane Database of Systematic Reviews. Several questions were examined. Is there any difference in maternal and neonatal outcomes depending on the intended place of birth? Does parity affect outcomes ?What are the characteristics of women who choose to deliver at home ?We conclude that giving birth at home improves obstetric outcomes but is riskier for the baby, especially for the first one. The women delivering at home are mainly white Europeans, between 25 and 35 years old, in a relationship, multiparous and wealthier. In order to avoid this increased risk for the baby while preserving the obstetric advantages, alongside birth centers offer an intermediate solution. They combine the reassuring home-like atmosphere with the safety of the hospital. In Belgium, the first alongside birth center " Le Cocon " (a low technicity unit distinct from the delivery room) offers now this type of alternative place of birth for women in Hôpital Erasme in Brussels.SCOPUS: ar.jinfo:eu-repo/semantics/publishe

Using simulation team training with human's factors components in obstetrics to improve patient outcome: A review of the literature

Objective: to assess the evidence from multidisciplinary simulation team training in obstetrics that integrates human's factors components on patient outcome. Introduction: It has been stated that simulation-based education has the potential to improve technical and nontechnical skills. Reports from enquiries into maternal and newborn adverse outcomes, highlight that the majority of incidents are due to a breakdown of communication and a lack of crisis resource management skills (CRM). It is therefore reasonable to think that a better training on teamwork based on simulation will ultimately improve obstetrics care. In order to explore further that idea, we conducted a literature review on patient outcome after a multidisciplinary simulation training in obstetrics. Method: Pubmed, Advances in health sciences education, BMC in medical education, BMC in pregnancy and Childbirth, BMJ open, BMJ Simulation and technology enhanced learning were searched from inception to May 2020 for full-text publications in English on interprofessional, multidisciplinary, obstetrics, simulation training, non-technical skills, CRM. Searches were limited to studies with a report on patient outcome after a multidisciplinary simulation program that included elements of CRM. Result: Out of the ten studies selected in our review, five were single site before and after prospective studies and five were cluster before and after randomized trials. All the single site studies reported a positive outcome in low and high resource countries. Three single site studies reported a reduction between 41 and 50 % of blood transfusion after simulation team training. Two single studies reported a reduction of maternal mortality by 34 % and a decrease in an adverse obstetrics index outcome from 0.052 to 0.048 with a p-value of 0.05. Cluster studies showed either no change or some improvement in patient outcomes such as a 37 % improvement on weighted obstetrics adverse outcome, a 17 % reduction in the incidence of PPH and a 47 % reduction in the incidence of retained placenta. Stillbirths rate was reduced by 34 % while newborn deaths was down by 62 %. There was also a 15 % reduction of maternal mortality in favor of the trained team after adjustment to the secular mortality trend. Neonatal death from 24 weeks during the first 24 h was also reduced by 83 % in the intervention site compare with an increase by 18 % in the control site. Conclusion: There is evidence that simulation team training that includes CRM is associated with better patient outcome. In order to consolidate this finding, appropriate methodology should be used in future studies with the support of health authorities.SCOPUS: re.jDecretOANoAutActifinfo:eu-repo/semantics/publishe