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Study of extracellular placental specific microRNAs in maternal circulation and their utilization in clinical diagnostics of pregnancy-related complications
Kotlabová, Kateřina ; Hromadníková, Ilona (advisor) ; Černá, Marie (referee) ; Korabečná, Marie (referee)
Objectiees: Our etudy inieetigated the poeeible utliiaton of extracellular microRNAAe ae noiel non-iniaeiie biomarkere for diaignoetce of preignancy-related complicatone euch ae igeetatonal hyperteneion (GH), preeclampeia (PE) and fetal igrowth reetricton (FGR). Firet, we identifed appropriate preignancy-aeeociated (placenta epeciifc) microRNAAe in maternal circulaton in preignanciee with normal couree of igeetaton. Then, we quantifed eelected extracellular C19MC microRNAAe in maternal circulaton oiertme in normally proigreeeinig preignanciee. Subeequently, we compared C19MC microRNAA expreeeion proiflee in maternal circulaton between preignanciee with clinically eetabliehed preignancy-related complicatone (PE, FGR, GH) and igeetatonal-aige-matched controle. Finally, monitorinig of eelected placenta epeciifc C19MC microRNAAe in maternal circulaton within the ifret trimeeter of igeetaton wae performed with the aim to identfy extracellular C19MC microRNAAe able to diferentate between normal preignanciee and thoee at riek of eubeequent deielopment of preignancy- related complicatone. Reeultes: The leiele and expreeeion proiflee of extracellular placental epeciifc microRNAAe in circulaton of non-preignant indiiiduale and preignant women were performed ueinig real- tme RT-PCR. The eelecton of...
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The expression profile of cardiovascular disease associated microRNAs in pregnancies with clinical manifestation of gestational hypertension, preeclampsia and intrauterine growth restriction
Bohatá, Jana ; Hromadníková, Ilona (advisor) ; Korabečná, Marie (referee)
MicroRNA (miRNA) are small non-coding 21-23 nucleotides long one strand RNAs. They are among the major posttranscriptional regulators of gene expression that regulate both physiological and pathological processes. Some of microRNAs, amount of their expression respectively, are specific only for certain type of tissue or pathological condition. The hypothesis for my diploma thesis was that gene expression of 28 cardiovascular disease associated microRNAs (miR-1-3p, miR-16-5p, miR-17-5p, miR- 20a-5p, miR-20b-5p, miR-21-5p, miR-23a-3p, miR-24-3p, miR-26a-5p, miR-29a-3p, miR-92a-3p, miR-100-5p, miR-103a-3p, miR-125b-5p, miR-126-3p, miR-130b-3p, miR-133a-3p, miR-143-3p, miR-145-5p, miR-146a-5p, miR-181-5p, miR-195-5p, miR- 199a-5p, miR-210-3p, miR-221-3p, miR-342-3p, miR-499a-5p, miR-574-3p) would differ in umbilical cord blood between groups of women with physiological pregnancies (FG), gestational hypertension (GH), preeclampsia (PE) and fetal growth restriciton (FGR). The studied cohort consisted of 184 pregnant women involving 44 controls, 47 GH pregnancies, 56 PE pregnancies and 37 FGR pregnancies. Relative quantification of microRNAs was performed by quantitative real-time PCR. Results showed a trend to miR-195-5p down-regulation in umbilical cord blood of GH patients. On the other hand, mild PE...
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Relation between cardiovascular microRNAs and pregnancy-related complications
Dalecká, Linda ; Hromadníková, Ilona (advisor) ; Daňková, Pavlína (referee)
During the pregnancy, there are many complications, which can affect a fetal development and eventually, its or mother's life. Pre-eclampsia is one of these complications. It is a hypertensive disorder, which appears during the pregnancy. Another example of these complications is an intrauterine growth restriction. It is a condition, when the fetus is not able to reach its genetical growth potential. Both of those pathological disorders are accompanied by changes in microRNA gene expression in placenta, for example miR-16, miR-21, miR-210. Knowledge about these changes in gene expression could represent unique instruments in the field of noninvasive prenatal diagnosis of these disorders. This could be benefitial due to the fact, that both of these disorders are responsible for increasing maternal and perinatal morbidity and mortality. This work focused on microRNAs, their expression in aformentioned disorders and benefits, which could bring in diagnostics. In this work these two disorders, characteristics of microRNAs and their biogenesi are described. MicroRNAs are interesting for us as potential biomarkers for noninvasive prenatal diagnostics becouse they are present in body fluids, such as plasma, breast milk or amniotic fluid and they are not subject of rapid degradation. Key words:...
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The Role of Heat Shock Proteins in Pathogenesis of Placental Insufficinecy
Slabá, Kristýna ; Hromadníková, Ilona (advisor) ; Černý, Jan (referee)
Heat shock proteins (Hsp) are highly conserved proteins that are part of the universal stress response of the cell. Their main function is to protect cells against structural and functional damage. Organisms exposed to different forms of stress, such as e.g. a lack of nutrients or water, hypoxia, infection or inflammation, demonstrated an increased gene expression of these proteins. Pregnancy complications cause stress conditions for maternal and fetal organism, which may result in an increased gene expression of Hsp. In my thesis, I examined the concentration of extracellular mRNA for five different heat shock proteins (Hsp27, Hsp60, Hsp70, Hsp90, HspBP1) in the plasma of pregnant women and wheather this concentration is affected by possible pregnancy complications (preeclampsia, fetal growth restriction and gestational hypertension). I also investigated a possible correlation between mRNA plasma concentration for Hsp and pulsatility index values (PI) obtained by Doppler ultrasound. This research should help to invent a new predictive method for pregnancy complications, based on a detection of specific biomarkers in the first trimester of pregnancy. The research was conducted on plasma samples obtained from peripheral blood of pregnant women, whose collection was performed during clinical manifestations of...
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Study of placental specific microRNA expression in pacients with spontaneous preterm birth and preterm prelabor rupture of membranes
Vintrová, Iva ; Hromadníková, Ilona (advisor) ; Daňková, Pavlína (referee)
MicroRNAs (miRNAs) are small non-coding RNAs with a length of 18 to 25 nucleotides playing a pivotal role in post-transcriptional regulation of gene expression. There are miRNAs whose expression is limited to a certain tissue type and diseases which are characterized by a unique miRNA expression profile. I assumed spontaneous preterm birth (PTB) and preterm prelabor rupture of membranes (PPROM) would be characterized by a unique miRNA expression profile. I observed the gene expression of 15 placental specific miRNAs (miR-512-5p, miR-515-5p, miR-516b-5p, miR-517-5p, miR-518b, miR-518f-5p, miR-519a-5p, miR-519d-3p, miR-519e- 5p, miR-520a-5p, miR-520h, miR-524-5p, miR-525-5p, miR-526a and miR-526b-5p) in placental tissue of pacients with PTB, PPROM and women with term in labor pregnancies (FG). PTB group consisted of 24 pacients, PPROM group of 75 pacients and FG group of 20 pacients. Quantitative real-time PCR was used to quantify gene expression. In the group of PTB pregnancies I identified 3 significantly upregulated miRNAs (miR-516b-5p, miR-519d-3p and miR-524-5p) and 4 miRNAs (miR-518b, miR-519a-5p, miR-520h and miR-526a) with a trend to upregulation compared to controls (FG). In the group of PPROM pregnancies I identified 3 miRNAs (miR-519d-3p, miR-520h and miR-256b-5p) with a trend to...
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The role of microRNAs in pathogenesis of placental insufficiency - related pregnancies
Fárková, Eva ; Hromadníková, Ilona (advisor) ; Korabečná, Marie (referee)
MicroRNAs (miRNAs) are small noncoding RNAs of length ranging from 18 to 25 nucleotides that regulate gene expression at posttranscriptional level. Expression of some miRNAs is tissue specific. This task assumed that placental insufficiency related pregnancy complications may be characterised by the dysregulation of microRNAs that are crutial for the regulation of cardiovascular system and cardiovascular diseases. The gene expresion of 32 cardiovascular miRNAs was studied in two types of biological material - placental tissue and maternal whole peripheral blood. Patient cohort of clinically manifested preeclampsia (PE), gestational hypertension (GH) and intrauterine growth restriction (IUGR) was compared with samples derived from normal pregnancies (FG). In total, 127 placental tissue samples (20 FG, GH 20, 20 IUGR , 67 PE) and 80 maternal whole peripheral blood samples (20 FG, GH 20, 20 IUGR , 20 PE) were processed. Before the research, two pilot studies focused on the selection of endogenous controls for data normalization of gene expression in both types of biological material were performed. Detection and quantification was carried out by quantitative real-time PCR. In a group of placental tissue several miRNAs showed elevated levels in GH (miR-1, miR-16, miR-17, miR-20a, miR-21, miR-23a,...
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NK cell involvement in the pathogenesis of autoimmune arthritis
Richter, Jan ; Fišerová, Anna (advisor) ; Hromadníková, Ilona (referee) ; Turánek, Jaroslav (referee)
Rheumatoid arthritis (RA) is a worldwide problem representing one of the most prevalent autoimmune diseases in the world. Despite the commonness of the disea- se, its pathogenesis has not been fully described. Immune cells ranging from antigen- presenting cells to T, B and NK cells playing various roles participate in the rheumatic process. In this work we concentrated on NK cells expressing a repertoire of activating and inhibitory receptors which influence their function in health and disease. We focused on the analysis of NK cell function and described its possible modulation by rheumatic autoantigens and multivalent glycodendrimers bearing 4 (GN4C) or 8 (GN8P) N-acetyl glucosamine moieties. The effect on NK cells and the glycosylation pathways was further studied in vitro. Finally, an in vivo study was performed on an animal model of RA - col- lagen-induced arthritis (CIA) to assess the effect of the compounds on clinical develop- ment of the disease and selected immune parameters. Comparison of NK cell cytotoxicity in patients suffering from RA, other inflam- matory diseases and healthy donors showed its impairment particularly in RA patients. Peripheral blood NK cells reacted to GN8P glycoconjugate by inhibition of their effector function in CD161 high-expressing samples. The MGAT5...
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Detection of extracellular microRNAs in maternal circulation - diagnosis and prognosis of pregnancy related complications
Ondráčková, Markéta ; Hromadníková, Ilona (advisor) ; Daňková, Pavlína (referee)
MicroRNAs (miRNAs) are small noncoding RNAs of length 18 to 25 nucleotides that regulate gene expression posttranscriptionally. Expression of some miRNAs is tissue specific. I assumed that pregnancy induced complications associated with placental insufficiency could be characterized by a unique profil of placental-specific miRNAs in maternal circulation. I measured concentration and gene expression of selected miRNAs (miR-516-5p, miR-517*, miR-518b, miR-520a*, miR-520h, miR-525 and miR-526a) in the plasma of patients with preeclampsia (PE), fetal growth restriction (FGR) and gestational hypertension (GH). The control group consisted of patients with a normal course of pregnancy (FG). I processed 168 plasma samples, the representation of individual diagnosis were as follows: PE 63, FGR 27, GH 23, FG 55. Detection and quantification was carried out by quantitative real-time PCR. I identified three miRNAs with elevated levels in a group of preeclamptic patients: miR-517*, miR-520a* and miR-525. The severity of the PE, which was characterized by a form of the disease (mild or severe PE) and term (before or after the 34th week of pregnancy), did not have a statistically significant effect on the levels of miRNAs. More than a quarter of patients had PE superimposed on previous hypertension. Previous history of...
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Heat shock proteins - - their role in diagnosis and prognosis of pregnancy related complications
Dvořáková, Lenka ; Hromadníková, Ilona (advisor) ; Růžičková, Šárka (referee)
Heat shock proteins increase their gene expression after exposure of cells or organisms to some forms of stress, which may be high temperature, infection, inflammation, hypoxia, lack of nutrients and water. Stressful situations for the body are also pregnancy-related complications associated with placental insufficiency - preeclampsia and IUGR, as well as other pregnancy-related complications such as fetal growth restriction and gestational hypertension. Therefore, I examined whether the occurrence of pregnancy-related complications (preeclampsia, fetal growth retardation, gestational hypertension) affect the gene expression of heat shock proteins. Five hsp systems was detected, namely Hsp27, Hsp60, Hsp70, Hsp90 and HspBP1 in placental tissue samples and whole maternal peripheral blood. Samples came from women with physiological pregnancy and from women with certain pregnancy-related complications (PE, FGR, GH). RNA was isolated from the samples. Detection of hsp expression was performed by using real-time RT-PCR and the comparative Ct method. Changes in gene expression between the test samples and reference sample were examined. To assess the difference between physiological pregnancies and pregnancies with selected pregnancy- related complications, an analysis of variance (ANOVA) was used....
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