Biomarker selection from mirnas obtained from mesenchymal stem cells for the early diagnosis of the preeclampsia

Preeclampsia (PE) is a pregnancy-specific hypertensive disease. Abnormal placentation is a distinctive feature of preeclampsia and causes increased placental oxidative stress. Oxidative stress triggers the release of anti-angiogenic factors that cause local damage to the decidual vascular structure. Afterwards, the damage spreads systematically and results in maternal clinical symptoms. Mesenchymal stem cells (MSC) at the maternal-fetal interface are thought to play an important role in the pathogenesis of preeclampsia. Decidua basalis mesenchymal stem cells (dMSC) are found in a vascular microenvironment. In preeclampsia, dMSCs are exposed to abnormally high levels of oxidative stress and inflammatory factors circulating in the maternal blood. dMSC exposed to oxidative stress is thought to play a role in the development of preeclampsia. Micro RNAs (miRNAs) are short (19-25 nucleotides), single-stranded, non-coding RNAs. They regulate gene expression by binding to 3'non-coding regions of target mRNAs. They are known to play an important role in many biological processes such as development, differentiation, apoptosis and oncogenesis. MiRNAs from the class of small non-coding RNAs have been shown to have a significant impact on the differentiation, maturation and function of MSCs. Recent researches on miRNAs have assert to the idea that these RNAs can be used as new molecular markers in PE. MSCs are multipotent precursor cells that can differentiate into many cell types and are abundant at the maternal fetal interface. They have the ability to promote angiogenesis and neurogenesis through the substances they secrete. Studies have shown that the expression of miRNAs in MSCs obtained from placenta of pregnant women with PE differs from that of normal pregnant women. This thesis covers miRNA expression analysis of MSCs originated decidua and peripheral blood obtained from PE patient and healthy pregnant women and is a preliminary study to investigate putative biomarkers. In this thesis, both decidua and peripheral blood samples were studied from total 14 pregnant women, 7 PE patients and 7 healthy subjects. In our research with a total of 28 samples, the number of cells was increased by MSC culture and then MSCs were sorted using by specific cell surface markers to obtain pure MSCs. After isolation of total RNA from MSCs, let-7b-3p, let-7f-1-3p, miR-191-3p, miR-550a-5p, miR-33b-3p and miR-425-3p were used for miRNA analysis and U6 snRNA was used for normalization of the samples. MiRNA analysis was performed by droplet digital polymerase chain reaction (ddPCR) method and the obtained results were evaluated using "GraphPad Prism 8.1.2" program. Significance tests were performed using non-parametric "Mann Whitney Test". Sensitivity and specificity tests were performed using "Receiver Operating Characteristic" (ROC) analysis. As a result of the analysis, it was observed that the levels of hsa-miR-33b-3p significantly (AUC: 0.93, p = 0.04, fold change: 4.5) increased in dMSC of PE patients compared to healthy controls. However, let-7b-3p, let-7f-1-3p, miR-191-3p, miR-550a-5p, and miR-425-3p were not considered as significant because they did not meet the p<0,05 requirement.