Chromosomal microarray vs. NIPS: analysis of 5541 low-risk pregnancies.

PURPOSE: To evaluate the diagnostic yield of chromosomal microarray (CMA) in pregnancies with normal ultrasound. METHODS: This retrospective cohort analysis included all pregnancies with normal ultrasound undergoing CMA testing between the years 2010 and 2016. We calculated the rate of detection of clinically significant CMA findings in the whole cohort and according to various indications. RESULTS: Of 5541 CMA analyses, clinically significant findings were yielded in 78 cases (1.4%). Of these, 31 (39.7%) variants could have theoretically been detected by karyotyping (e.g., sized above 10 Mb), and 28 (35.9%) by noninvasive prenatal screening aimed at five common aneuploidies. Of the 47 submicroscopic findings detectable by CMA only, the majority (37 cases, 78.7%) represented known recurrent syndromes. Detection of clinically significant CMA findings in women with no indication for invasive testing was 0.76% (21/2752), which was significantly lower compared with 1.8% in advanced maternal age group (41/2336), 2.8% in abnormal biochemical serum screening (6/211), and 4.1% (10/242) in fetuses with sonographic soft markers. CONCLUSION: Clinically significant CMA aberrations are detected in 1 of 71 pregnancies with normal ultrasound, and in 1 of 131 women with no indication for invasive testing. Thus, CMA might be recommended a first-tier test in pregnancies with normal ultrasound.

Microarray findings in pregnancies with oligohydramnios - a retrospective cohort study and literature review.

Objective To explore the risk for abnormal chromosomal microarray analysis (CMA) findings in pregnancies with oligohydramnios. Methods Data from all CMA analyses performed due to oligohydramnios between 2013 and 2017 were retrospectively obtained from the Israeli Ministry of Health database. The rate of clinically significant (pathogenic and likely pathogenic) findings was compared to a local cohort of pregnancies with normal ultrasound, yielding a 1.4% rate of abnormal CMA results. In addition, a search was conducted through the PubMed database addressing the issue. Results Fifty CMA analyses were performed due to oligohydramnios. The 2% risk for clinically significant CMA finding in pregnancies with oligohydramnios did not differ from the control population of 5541 pregnancies with normal ultrasound - relative risk (RR) 1.4 [95% confidence interval (CI) 0.2-10.2]. Literature search yielded 394 titles, of which four relevant articles were selected, all using fetal karyotyping. Conclusion There is yet insufficient evidence to support invasive prenatal testing in pregnancies with isolated oligohydramnios.

When genotype is not predictive of phenotype: implications for genetic counseling based on 21,594 chromosomal microarray analysis examinations.

PurposeTo compare the frequency of copy-number variants (CNVs) of variable penetrance in low-risk and high-risk prenatal samples and postnatal samples.MethodsTwo cohorts were categorized according to chromosomal microarray analysis (CMA) indication: group I, low-risk prenatal-women with uneventful pregnancy (control group); group II, high-risk prenatal-women whose fetuses had congenital malformations; and group III, postnatal-individuals with unexplained developmental delay/intellectual disability, autism spectrum disorders, or multiple congenital anomalies. CNVs were categorized based on clinical penetrance: (i) high (>40%), (ii) moderate (10-40%), and (iii) low (<10%).ResultsFrom 2013 to 2016, 21,594 CMAs were performed. The frequency of high-penetrance CNVs was 0.1% (21/15,215) in group I, 0.9% (26/2,791) in group II, and 2.6% (92/3,588) in group III. Moderate-penetrance CNV frequency was 0.3% (47/15,215), 0.6% (19/2,791), and 1.2% (46/3,588), respectively. These differences were statistically significant. The frequency of low-penetrance CNVs was not significantly different among groups: 0.6% (85/15,215), 0.9% (25/2,791), and 1.0% (35/3,588), respectively.ConclusionHigh-penetrance CNVs might be a major factor in the overall heritability of developmental, intellectual, and structural anomalies. Low-penetrance CNV alone does not seem to contribute to these anomalies. These data may assist pre- and posttest CMA counseling.

Microarray analysis in pregnancies with isolated unilateral kidney agenesis.

BackgroundThe objective of our study was to examine the risk for submicroscopic chromosomal aberrations among fetuses with apparently isolated solitary kidney.MethodsData acquisition was performed retrospectively by searching Israeli Ministry of Health-computerized database. All cases having chromosomal microarray analysis (CMA), referred because of an indication of isolated unilateral kidney agenesis between January 2013 and September 2016, were included. Rate of clinically significant CMA findings in these pregnancies was compared to pregnancies with normal ultrasound, based on a systematic review encompassing 9,792 cases and local data of 5,541 pregnancies undergoing CMA because of maternal request.ResultsOf the 81 pregnancies with isolated solitary kidney, 2 (2.47%) loss-of-copy number variants compatible with well-described deletion syndromes were reported (16p11.2-16p12.2 and 22q11.21 microdeletion syndromes). In addition, one variant of unknown significance was demonstrated. The relative risk for pathogenic CMA findings among pregnancies with isolated unilateral renal agenesis was not significantly different compared with the control population.ConclusionCMA analysis in pregnancies with unilateral renal agenesis might still be useful, to the same degree as it can be in the general population.

An improved design of optical sensor for long-term measurement of arterial blood flow waveform.

We present here the improved design and development of optical sensor for non-invasive measurements of arterial blood flow waveform. The sensor is based on a physical principle of reflective photoplethysmography (PPG). As the light source we used serially connected infrared diodes whereas NPN silicon phototransistors were used as light detectors. The electronic components were molded into square package and poured with silicone. Such preparation produced an elastic superficies that allowed excellent attachment of the sensor on the skin's surface. Moreover, a serial connection of infrared diodes and phototransistors completely eliminated signal artifacts caused by minor muscle contractions. The sensor recording performances were examined at the photoplethysmographic sites on three different arteries; the commune carotid, femoral and radial and, on each site the sensor demonstrated remarkable capability to make a consistent, reproducible measurements. Because of the advantageous physical and electrical properties, the new sensor is suitable for various cardiovascular diagnostics procedures, especially when long-term measurements of arterial blood flow waveform are required, for monitoring of different parameters in cardiovascular units and for research.

Terminal microdeletions of 13q34 chromosome region in patients with intellectual disability: Delineation of an emerging new microdeletion syndrome.

The increasing use of chromosomal microarray studies in patients with intellectual disability has led to the description of new microdeletion and microduplication syndromes. We report terminal microdeletions in 13q34 chromosome region in 5 adult patients of two unrelated families. Patients harboring 13q34 microdeletions display common clinical features, including intellectual disability, obesity, and mild facial dysmorphism. These individuals can become fairly self-sufficient, however they do not live independently, and require community and social support. Further systematic analysis of the genes comprised in the deleted region will allow the identification of genes whose haploinsufficiency is expected to lead to disease manifestations, in particular intellectual disability.

MicroRNAs are essential for development and function of inner ear hair cells in vertebrates.

MicroRNAs (miRNAs) inhibit the translation of target mRNAs and affect, directly or indirectly, the expression of a large portion of the protein-coding genes. This study focuses on miRNAs that are expressed in the mouse cochlea and vestibule, the 2 inner ear compartments. A conditional knock-out mouse for Dicer1 demonstrated that miRNAs are crucial for postnatal survival of functional hair cells of the inner ear. We identified miRNAs that have a role in the vertebrate developing inner ear by combining miRNA transcriptome analysis, spatial and temporal expression patterns, and bioinformatics. Microarrays revealed similar miRNA profiles in newborn-mouse whole cochleae and vestibules, but different temporal and spatial expression patterns of six miRNAs (miR-15a, miR-18a, miR-30b, miR-99a, miR-182, and miR-199a) may reflect their roles. Two of these miRNAs, miR-15a-1 and miR-18a, were also shown to be crucial for zebrafish inner ear development and morphogenesis. To suggest putative target mRNAs whose translation may be inhibited by selected miRNAs, we combined bioinformatics-based predictions and mRNA expression data. Finally, we present indirect evidence that Slc12a2, Cldn12, and Bdnf mRNAs may be targets for miR-15a. Our data support the hypothesis that inner ear tissue differentiation and maintenance are regulated and controlled by conserved sets of cell-specific miRNAs in both mouse and zebrafish.

The yield of chromosomal microarray analysis among pregnancies terminated due to fetal malformations.

BACKGROUND: Chromosomal microarray analysis (CMA) is preferred for genetic work-up when fetal malformations are detected prenatally. OBJECTIVES: To assess the detection rate of CMA after pregnancy termination due to abnormal ultrasound findings. METHODS: CMA was successfully performed in 71 pregnancies using fetal DNA (mainly from skin) or placenta. Data regarding clinical background, pregnancy work-up, and CMA were analyzed. RESULTS: Findings were abnormal in 17 cases (23.9%), of which 13 were detectable by karyotype. The incremental yield of CMA was 4/71 (5.6%); 1/32 (3.1%) for cases with an isolated anomaly and 3/39 (7.7%) for cases with nonisolated anomalies. CONCLUSIONS: CMA yield from terminated pregnancies was 23.9%. Although most chromosomal abnormalities are detectable by karyotype, CMA does not require viable dividing cells; hence, it is more practical for work-up after termination. In most cases, the diagnosis was followed by consultation regarding the risk of recurrence and recommendations for testing in subsequent pregnancies.

Frequent aneuploidy in primary human T cells after CRISPR-Cas9 cleavage.

Multiple clinical trials of allogeneic T cell therapy use site-specific nucleases to disrupt T cell receptor (TCR) and other genes1-6. In this study, using single-cell RNA sequencing, we investigated genome editing outcomes in primary human T cells transfected with CRISPR-Cas9 and guide RNAs targeting genes for TCR chains and programmed cell death protein 1. Four days after transfection, we found a loss of chromosome 14, harboring the TCRα locus, in up to 9% of the cells and a chromosome 14 gain in up to 1.4% of the cells. Chromosome 7, harboring the TCRß locus, was truncated in 9.9% of the cells. Aberrations were validated using fluorescence in situ hybridization and digital droplet PCR. Aneuploidy was associated with reduced proliferation, induced p53 activation and cell death. However, at 11 days after transfection, 0.9% of T cells still had a chromosome 14 loss. Aneuploidy and chromosomal truncations are, thus, frequent outcomes of CRISPR-Cas9 cleavage that should be monitored and minimized in clinical protocols.

Should We Report 15q11.2 BP1-BP2 Deletions and Duplications in the Prenatal Setting?

Copy number variations of the 15q11.2 region at breakpoints 1-2 (BP1-BP2) have been associated with variable phenotypes and low penetrance. Detection of such variations in the prenatal setting can result in significant parental anxiety. The clinical significance of pre- and postnatally detected 15q11.2 BP1-BP2 deletions and duplications was assessed. Of 11,004 chromosomal microarray tests performed in a single referral lab (7596 prenatal, 3408 postnatal), deletions were detected in 66 cases: 39 in prenatal tests (0.51%) and 27 in postnatal tests (0.79%). Duplications were detected in 94 cases: 62 prenatal tests (0.82%) and 32 postnatal tests (0.94%). The prevalence of deletions and duplications among clinically indicated prenatal tests (0.57% and 0.9%, respectively) did not differ significantly in comparison to unindicated tests (0.49% and 0.78%, respectively). The prevalence of deletions and duplications among postnatal tests performed for clinical indications was similar to the prevalence in healthy individuals (0.73% and 1% vs. 0.98% and 0.74%, respectively). The calculated penetrance of deletions and duplications over the background risk was 2.18% and 1.16%, respectively. We conclude that the pathogenicity of 15q11.2 BP1-BP2 deletions and duplications is low. Opting out the report of these copy number variations to both clinicians and couples should be considered.

Chromosomal Microarray Analysis Results From Pregnancies With Various Ultrasonographic Anomalies.

OBJECTIVE: To examine chromosomal microarray analysis results in pregnancies with various ultrasonographic anomalies and to characterize the copy number variants in diverse fetal phenotypes. METHODS: We retrospectively examined chromosomal microarray analyses of amniocenteses performed nationwide as a result of fetal ultrasonographic anomalies (structural defects, fetal growth restriction, and polyhydramnios) between January 2013 and September 2017. The rate of abnormal chromosomal microarray findings was compared between the different phenotypes and with a previously described control population of 15,225 pregnancies with normal ultrasonographic findings. RESULTS: Clinically significant chromosomal microarray aberrations were detected in 272 of 5,750 pregnancies (4.7%): 115 (2%) karyotype-detectable and 157 (2.7%) submicroscopic. Most commonly detected copy number variants were 22q11.21 deletions (0.4%) followed by 22q11.21 gain of copy number (0.2%). Specific copy number variants detected among pregnancies with abnormal ultrasonographic findings were up to 20-fold more prevalent compared with low-risk pregnancies. Some variants were associated with specific phenotypes (eg, 22q11.21 microdeletions with cardiovascular and 17q12 microdeletions with genitourinary defects). CONCLUSION: The rate of abnormal amniotic chromosomal microarray analysis results is twice that of karyotypic abnormalities in pregnancies with various abnormal ultrasonographic findings.

Identification of a novel hypoxia-inducible factor 1-responsive gene, RTP801, involved in apoptosis.

Hypoxia is an important factor that elicits numerous physiological and pathological responses. One of the major gene expression programs triggered by hypoxia is mediated through hypoxia-responsive transcription factor hypoxia-inducible factor 1 (HIF-1). Here, we report the identification and cloning of a novel HIF-1-responsive gene, designated RTP801. Its strong up-regulation by hypoxia was detected both in vitro and in vivo in an animal model of ischemic stroke. When induced from a tetracycline-repressible promoter, RTP801 protected MCF7 and PC12 cells from hypoxia in glucose-free medium and from H(2)O(2)-triggered apoptosis via a dramatic reduction in the generation of reactive oxygen species. However, expression of RTP801 appeared toxic for nondividing neuron-like PC12 cells and increased their sensitivity to ischemic injury and oxidative stress. Liposomal delivery of RTP801 cDNA to mouse lungs also resulted in massive cell death. Thus, the biological effect of RTP801 overexpression depends on the cell context and may be either protecting or detrimental for cells under conditions of oxidative or ischemic stresses. Altogether, the data suggest a complex type of involvement of RTP801 in the pathogenesis of ischemic diseases.

Effect of viscosity on the wave propagation: Experimental determination of compression and expansion pulse wave velocity in fluid-fill elastic tube.

The velocity by which the disturbance travels through the medium is the wave velocity. Pulse wave velocity is one of the main parameters in hemodynamics. The study of wave propagation through the fluid-fill elastic tube is of great importance for the proper biophysical understanding of the nature of blood flow through of cardiovascular system. The effect of viscosity on the pulse wave velocity is generally ignored. In this paper we present the results of experimental measurements of pulse wave velocity (PWV) of compression and expansion waves in elastic tube. The solutions with different density and viscosity were used in the experiment. Biophysical model of the circulatory flow is designed to perform measurements. Experimental results show that the PWV of the expansion waves is higher than the compression waves during the same experimental conditions. It was found that the change in viscosity causes a change of PWV for both waves. We found a relationship between PWV, fluid density and viscosity.

Dissection of a DNA-damage-induced transcriptional network using a combination of microarrays, RNA interference and computational promoter analysis.

BACKGROUND: Gene-expression microarrays and RNA interferences (RNAi) are among the most prominent techniques in functional genomics. The combination of the two holds promise for systematic, large-scale dissection of transcriptional networks. Recent studies, however, raise the concern that nonspecific responses to small interfering RNAs (siRNAs) might obscure the consequences of silencing the gene of interest, throwing into question the ability of this experimental strategy to achieve precise network dissections. RESULTS: We used microarrays and RNAi to dissect a transcriptional network induced by DNA damage in a human cellular system. We recorded expression profiles with and without exposure of the cells to a radiomimetic drug that induces DNA double-strand breaks (DSBs). Profiles were measured in control cells and in cells knocked-down for the Rel-A subunit of NFkappaB and for p53, two pivotal stress-induced transcription factors, and for the protein kinase ATM, the major transducer of the cellular responses to DSBs. We observed that NFkappaB and p53 mediated most of the damage-induced gene activation; that they controlled the activation of largely disjoint sets of genes; and that ATM was required for the activation of both pathways. Applying computational promoter analysis, we demonstrated that the dissection of the network into ATM/NFkappaB and ATM/p53-mediated arms was highly accurate. CONCLUSIONS: Our results demonstrate that the combined experimental strategy of expression arrays and RNAi is indeed a powerful method for the dissection of complex transcriptional networks, and that computational promoter analysis can provide a strong complementary means for assessing the accuracy of this dissection.