Unit:
Faculty of MedicineLibrary of the School of Health Sciences
Author:
Lykoudi Alexandra
Dissertation committee:
Παπαντωνίου Νικόλαος, Ομότιμος Καθηγητής, Ιατρική Σχολή, ΕΚΠΑ
Μαύρου Καλπίνη Αριάδνη, Ομότιμη Καθηγήτρια, Ιατρική Σχολή, ΕΚΠΑ
Τζέτη Μαρία, Καθηγήτρια, Ιατρική Σχολή, ΕΚΠΑ
Traeger Συνοδινού Ιωάννα, Καθηγήτρια, Ιατρική Σχολή, ΕΚΠΑ
Δασκαλάκης Γεώργιος, Καθηγητής, Ιατρική Σχολή, ΕΚΠΑ
Ελευθεριάδης Μακάριος, Αναπληρωτής Καθηγητής, Ιατρική Σχολή, ΕΚΠΑ
Αραβαντινός Λέων, Επίκουρος Καθηγητής,Ιατρική Σχολή, ΕΚΠΑ
Original Title:
Μελέτη του προτύπου έκφρασης των miRNA στην προεκλαμψία
Translated title:
Dysregulated miRNA in early and late onset preeclampsia
Summary:
Introduction: Preeclampsia is a hypertensive obstetric complication characterized by generalized maternal systemic inflammation and endothelial dysfunction. The complication is associated with substantial maternal and fetal morbidity and mortality. MicroRNAs (miRNA) belong to a family of small non-coding RNAs that regulate gene expression at the stage of post-transcriptional level by degrading or blocking translation of messenger RNA (mRNA).
Aim: The aim of the study was to characterize miRNAs associated with the pathogenic mechanism of Preeclampsia analyzing their expression profiles both in placental tissue biopsies at delivery and plasma samples from women in the first trimester of pregnancy that subsequently develop late onset Preeclampsia (after 20th week of gestation), compared to uncomplicated pregnancies. Furthermore, we aimed to evaluate the potential use of these miRNAs as non-invasive diagnostic and putative prognostic biomarkers for Preeclampsia.
Materials and Methods: On the first stage of the study, the expression profile of placental miRNAs was analyzed in 16 placental tissue biopsies from women with Preeclampsia (11 with early onset and 5 with late onset Preeclampsia) and 8 placental tissue biopsies from uncomplicated pregnancies, using miRNA microarrays. The samples were analyzed based on the stage of onset and the severity of clinical manifestation of the complication.
During the second stage of the study, maternal peripheral blood samples were obtained from a non-selected population of 2437 Caucasian pregnant women at 11+0 to 13+6 weeks of gestation undergoing first trimester prenatal screening for fetal aneuploidies. Based on these data, 34 samples were retrieved for miRNA profiling analysis, comprising 17 from women who subsequently developed late onset Preeclampsia and required delivery at >34 weeks of gestation and 17 from uncomplicated pregnancies matching for maternal age, gestational age and duration of storage. The expression profile of miRNAs was analyzed in 5 first trimester plasma samples from women who developed late onset Preeclampsia and 5 controls using deep sequencing analysis. The study was focused in late onset Preeclampsia (delivery >34 weeks of gestation) that consist 80% of complication cases.
For statistical analysis, the MATLAB® simulation environment was applied. Differences in clinical characteristics between the two groups were assessed using one-way ANOVA for independent variables or Mann-Whitney U-test for continuous variables. Receiver operating characteristic (ROC) curve was performed to evaluate the diagnostic potential of differentially expressed miRNAs with 95% standard error and 95% confidence intervals. Differentially expressed miRNAs were further analyzed and enriched for known functions and their potential target genes. In gene ontology analysis (GO), the number of genes corresponding to GO entries namely Biological Process, Molecular function and Cellular Component, were determined and enrichment score was reported as the -log (p-value). The results of miRNAs expression profiles both of microarrays and deep sequencing were verified in all samples using Quantitative Real-Time Polymerase Chain Reaction, a method of increased sensitivity and specificity for the detection of these sequences.
Results: Expression profile analysis of miRNAs in placental tissue biopsy from pregnancies complicated by Preeclampsia compared to healthy uncomplicated pregnancies revealed a more than two fold increased in expression levels of 8 miRNAs (miR-500a, miR-383, miR-518a-5p/miR-527, miR-431, miR-423-3p, miR-124*, miR-1183, miR-130b) and a more than two-fold decrease in expression levels of 2 miRNAs (miR-3942, miR-544b) in Preeclampsia cases. ROC analysis revealed statistical significant correlation of miRNAs with Preeclampsia subtypes and severity of clinical symptoms. MiR-423, miR-124* and miR-431 were over-expressed while miR-544b and miR-3942 were down regulated in early onset Preeclampsia cases as compared to late onset Preeclampsia and controls. Importantly, over-expression of miR-518a-5p/miR-527 in cases of early onset Preeclampsia was further associated with intra uterine growth restriction and severe clinical manifestation, emerging its diagnostic potential on prediction of severity of the complication.
Deep sequencing analysis of circulating miRNA in plasma sample of pregnant women with late onset Preeclampsia compared to those with uncomplicated pregnancies revealed 2 miRNAs with significant increased expression profile (miR-525-5p, miR-548e-3p) and 2 miRNAs with significant reduced expression (miR-99b-5p, miR-23b-5p). In silico analysis showed that target genes of these miRNAs are associated with signaling pathways of angiogenesis, immune response, lymphocyte differentiation and insulin resistance. Since these miRNAs implicate in pathogenic mechanisms leading to the induction of late onset Preeclampsia, we found that in combination with clinical symptoms could reveal predictive models for diagnosis with >60% sensitivity and >70% specificity. Interestingly, the predictive model of miR-525-5p - PAPP-A - MAP showed 100% specificity and 73% sensitivity while the predictive model of miR-548e-3p - PAPP-A - UtPI showed 90% sensitivity and 76% specificity.
Conclusions: Our findings provided significant insights about the role of miRNAs in pathogenesis of Preeclampsia and emphasize the possibility of using these molecules as biomarkers for the prevention, prognosis and non-invasive diagnosis of the complication. Since specific miRNAs can differentiate the subtypes of Preeclampsia, these molecules might also guide the development of novel therapeutic interventions. The results of this study are preliminary and thus further studies using larger heterogeneous cohorts are required to validate the data obtained, assess their clinical value and establish the performance of miRNAs as biomarkers of Preeclampsia.
Main subject category:
Health Sciences
Keywords:
microRNAs, Preeclampsia, Εarly onset preeclampsia, Late onset preeclampsia, placenta, NGS, Microarrays
Number of references:
288
