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Browsing by Subject "DNA-metylaatio"

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  • Laukkanen, Ida (2024)
    Epigenetics is the study of changes in gene expression without alterations in the DNA sequence. Epigenetic modifications, of which DNA methylation (DNAm) is the most known, are crucial for many biological events, especially for normal development and genomic imprinting. In imprinted genes, only one of the parental alleles is consistently monoallelically expressed. The epigenome can be altered by various environmental factors such as diet and chemicals. Moreover, evidence indicates that assisted reproductive technology (ART) is associated with distinct DNAm patterns. Still, it is unclear whether these alterations are a consequence of ART procedures themselves or the underlying subfertility. This thesis aimed to study whether ART procedures and subfertility are associated with aberrant DNAm at the imprinted DLK1-DIO3 locus in the human placenta. The genes encoded from this paternally imprinted locus are essential in mammals' fetal and placental development. Moreover, recent evidence suggests that downregulation of the DLK1 gene is associated with ART and subfertility. Therefore, two gene regions, the IG-DMR and the DLK1 promoter, were chosen as study objects. The IG-DMR acts as the main regulator of gene expression at the whole DLK1-DIO3 locus, and promoter regions are considered important regulators of the expression of their corresponding genes. Eight ART, four subfertility, and six control samples of human placental DNA were studied. The subfertility group consisted of couples who were committed to initiating fertility treatments but eventually got pregnant spontaneously. The study was performed using traditional bisulfite sequencing, after which the differences in DNAm levels between study groups were statistically analyzed. As expected, significantly decreased DNAm level was observed in the placentas of subfertile couples compared to controls. Surprisingly, no significant differences were addressed between ART and control groups in either region. The partly unexpected results are explained by the fact that aberrant methylation at distinct imprinted DMRs, including the IG-DMR, is caused by the in vitro culture media. Moreover, the results indicate that the regulation of DLK1 expression is more complicated than that solely by the IG-DMR and the DLK1 promoter. Further research should be dedicated to differentiating the impacts of ART and subfertility on the epigenome and phenotype to better understand the health implications of ART.
  • Ahola, Laura (2021)
    Environment is known to be a strong mediator of embryonal development and the future health of an individual. According to earlier studies, early pregnancy is especially vulnerable to environmental influence. Early embryogenesis is a critical period when epigenetic reprogramming occurs and epigenetic modifications are established. Alcohol is an environmental factor and a teratogen that affects normal epigenetic reprogramming and embryonal development. Prenatal alcohol exposure may contribute to the development of abnormal phenotype or diseases such as fetal alcohol spectrum disorders, FASD. This master’s thesis is part of the epiFASD study at the Environmental Epigenetic Laboratory, University of Helsinki. The study focuses on the environmental impact on the epigenetic mechanisms of FASD and finding possible future biomarkers of early disease. The research group has collected biological samples from a cohort of control and alcohol exposed newborns and their parents. The main aim of the study is to reveal the effects of prenatal alcohol exposure to the epigenetic reprogramming of the newborn. If there are epigenetic fingerprints to be seen in the first developing cells of the embryo, these fingerprints may spread to other cells and tissues by cell proliferation. The main aim of this master’s thesis was to optimize a DNA extraction protocol for the collected buccal cell samples. The optimization was expected to enhance the concentration and purity of the DNA samples for future studies. The group had found earlier prenatal alcohol exposure associated changes on the DNA methylation of alcohol-exposed placentas by genome-wide microarrays. The second aim of the thesis was to observe if similar DNA methylation patterns are found in both buccal epithelial cells and placental tissue. The optimization of the DNA extraction protocol enhanced the concentration but not significantly the purity of the buccal cell DNA samples. The earlier microarray studies with placental tissue revealed an interesting candidate gene and the locus-specific EpiTYPER-analysis confirmed the results: the regulatory regions of the studied gene were less methylated in alcohol-exposed placentas compared to controls. EpiTYPER also showed that methylation levels of the placenta and buccal epithelial cells did not correlate with each other although the changes were similar. Further research needs to be done to confirm if the methylation changes could be used as biomarkers in the diagnosis of alcohol-related disorders.