Orsolya Biró

Identifying miRNA regulatory mechanisms in preeclampsia by systems biology approaches

ABSTRACT Background: Preeclampsia (PE) is the major cause of maternal and fetal morbidity and mortality, affecting 3–8% of all pregnancies around the globe. miRNAs are small, noncoding RNA molecules, which negatively regulate gene expression. Abnormally expressed miRNAs contribute to pregnancy complications such as PE. The aim of our study was to find possible regulatory mechanisms by system biology approaches, which are connected to the pathogenesis of PE. Methods: We integrated publicly available miRNA and gene expression profiles and created a network from the significant miRNA–mRNA pairs with the help of MAGIA and Cytoscape softwares. Two subnetworks were expanded by adding protein–protein interactions. Differentially expressed miRNAs were identified for the evaluation of their regulatory effect. We analyzed the miRNAs and their targets using different bioinformatics tools and through literature research. Results: Altogether, 52,603 miRNA–mRNA interactions were generated by the MAGIA web tool. The top 250 interactions were visualized and pairs with q < 0.0001 were analyzed, which included 85 nodes and 80 edges signalizing the connections between 52 regulated genes and 33 miRNAs. A total of 11 of the regulated genes are PE related and 9 of them were targeted by multiple miRNAs. A total of 8 miRNAs were associated with PE before, and 13 miRNAs regulated more than 1 mRNA. Hsa-mir-210 was the highest degree node in the network and its role in PE is well established. Conclusions: We identified several miRNA–mRNA regulatory mechanisms which may contribute to the pathogenesis of PE. Further investigations are needed to validate these miRNA–mRNA interactions and to enlighten the possibilities of developing potential therapeutic targets.

Utilization of benchtop next generation sequencing platforms ion torrent PGM and miseq in noninvasive prenatal testing for chromosome 21 trisomy and testing of impact of in silico and physical size selection on its analytical performance

Objectives The aims of this study were to test the utility of benchtop NGS platforms for NIPT for trisomy 21 using previously published z score calculation methods and to optimize the sample preparation and data analysis with use of in silico and physical size selection methods. Methods Samples from 130 pregnant women were analyzed by whole genome sequencing on benchtop NGS systems Ion Torrent PGM and MiSeq. The targeted yield of 3 million raw reads on each platform was used for z score calculation. The impact of in silico and physical size selection on analytical performance of the test was studied. Results Using a z score value of 3 as the cut-off, 98.11% - 100% (104-106/106) specificity and 100% (24/24) sensitivity and 99.06% - 100% (105-106/106) specificity and 100% (24/24) sensitivity were observed for Ion Torrent PGM and MiSeq, respectively. After in silico based size selection both platforms reached 100% specificity and sensitivity. Following the physical size selection z scores of tested trisomic samples increased significantly—p = 0.0141 and p = 0.025 for Ion Torrent PGM and MiSeq, respectively. Conclusions Noninvasive prenatal testing for chromosome 21 trisomy with the utilization of benchtop NGS systems led to results equivalent to previously published studies performed on high-to-ultrahigh throughput NGS systems. The in silico size selection led to higher specificity of the test. Physical size selection performed on isolated DNA led to significant increase in z scores. The observed results could represent a basis for increasing of cost effectiveness of the test and thus help with its penetration worldwide.

Various levels of circulating exosomal total-miRNA and miR-210 hypoxamiR in different forms of pregnancy hypertension

INTRODUCTION Hypertension is a common complication during pregnancy, affecting 10% of pregnant women worldwide. Several microRNA (miRNA) were shown to be involved in hypertensive disorders of pregnancy. In preeclampsia (PE), placental dysfunction causes the enhanced release of extracellular vesicle-derived miRNAs. The hypoxia-sensitive hsa-mir-210 is the most common PE-associated miRNA, but its exosomal profile has not been investigated. OBJECTIVES Our aims were to measure exosomal total-miRNA concentration and to perform expression analysis of circulating exosomal hsa-miR-210 in women affected by chronic hypertension (CHT) gestational hypertension (GHT) or PE. MATERIALS AND METHODS We collected plasma samples from women with CHT, GHT, PE (moderate: mPE and severe: sPE) and from normotensive pregnancies. Exosomal miRNAs were extracted and miRNA concentration was measured. RT-PCR was carried out with hsa-miR-210-3p-specific primers and relative expression was calculated using the comparative Ct method. RESULTS The total-miRNA concentration was different in the disease subgroups, and was significantly higher in mPE and sPE compared to the other groups. We found a significant difference in the relative exosomal hsa-miR-210-3p expression between all hypertensive groups compared to the normotensive samples, but significant upregulation was only observed in case of mPE and sPE patients. Both the level of total-miRNA and hsa-miR-210 expression was higher in case of severe PE. CONCLUSIONS The level of circulating exosomal total-miRNA and hsa-miR-210 was elevated in women with PE, and it was higher in the severe form. We showed that hsa-miR-210 is secreted via exosomes, which may have a role in the pathomechanism of the disease.

Circulating cell-free nucleic acids: characteristics and applications

Liquid biopsy is becoming a very popular sample obtaining procedure, replacing the invasive sampling methods for the diagnostic protocols. The advantages of this method include the possibility to isolate cell-free nucleic acids (cfNAs) for diagnostic or screening purposes. A comprehensive review combining all current and perspective applications of cell-free nucleic acids is missing. Published articles are dealing with one type of cfNAs, or discuss them from the perspective of single disorder. We collected here all known types of cfNAs which are known to be present in biological fluids and could be involved in further studies to find out the exact biological role of them in normal physiological and pathological conditions. Beyond doubt, cfNAs will have a tremendous effect in future screening, diagnosis, prognosis, follow-up and treatment of cardiovascular diseases, cancer, diabetes and other diseases.

Relationship between cardiovascular diseases and circulating cell-free nucleic acids in human plasma

Cardiovascular diseases (CVDs) are the main cause of human morbidity and mortality worldwide. Early diagnosis could improve the efficiency of treatments. New biomarkers are needed for the identification of high-risk populations in order to make accurate diagnosis and therapy monitoring. Circulating cell-free nucleic acids (cf-NAs) offer a promising new noninvasive tool. These have a role in the regulation of normal physiological functions and in the development of pathological alterations. There is extended research on the clinical application and utilization of cell-free genomic DNA, mtDNA, mRNA, miRNA and long noncoding RNA in CVDs. These molecules could serve as components of new generation therapeutics. Our review focuses on the role of cf-NAs in the pathogenesis of CVDs and we are discussing also possible diagnostic applications and therapeutic implications.

Noninvasive prenatal testing for congenital heart disease – cell-free nucleic acid and protein biomarkers in maternal blood

Abstract Context: Congenital heart disease (CHD) is the most common fetal malformation. Prenatal ultrasonography is routinely applied for the screening of CHD but many factors influence its diagnostic accuracy. The introduction of new biomarkers could facilitate the identification of high-risk pregnancies. Objective: In our review, our aim was to collect expression studies of cell-free nucleic acids and proteins in maternal circulation. Syndromic CHDs which can be detected by noninvasive prenatal testing (NIPT) techniques were also discussed. Methods: PubMed and Web of Science databases were screened for studies where the levels of potential CHD biomarkers were measured in maternal blood samples. Available NIPT tests were collected from the providers’ resources. Results: There are nine CHD-associated chromosomal abnormalities, five aneuploidies, and four microdeletions, which are included in NIPT panels. We found eight articles from which five included the analysis of specific cell-free RNA expression and three measurements of protein levels. Conclusions: Most of the common heart-related chromosomal aberrations can be diagnosed by NIPT. Specific cell-free RNAs and circulating proteins seem to be potential biomarkers for fetal CHDs. The application of these new biomarkers could improve the detection rate at early pregnancy, making it possible to provide optimal perinatal and perioperative management.