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DNA methylation as a potential biomarker for radiation exposure

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dc.contributor.advisor정해원-
dc.contributor.author이영현-
dc.date.accessioned2017-07-13T17:21:53Z-
dc.date.available2017-07-13T17:21:53Z-
dc.date.issued2014-02-
dc.identifier.other000000016955-
dc.identifier.urihttps://hdl.handle.net/10371/120778-
dc.description학위논문 (박사)-- 서울대학교 보건대학원 : 보건학과, 2014. 2. 정해원.-
dc.description.abstractEpigenetic changes, including DNA methylation, microRNA expression and histone modification, are heritable and stable alterations in regulation of gene transcription. Although the changes are required for normal health and development, they can also contribute to diseases including cancers. The epigenetic changes could be a missing link among radiation exposure, radiation-induced genomic instability, and radiation-induced carcinogenesis. While recent studies have shown that ionizing radiation (IR) can affect epigenetic changes in animal and cancer cell lines, there is a lack of information on epigenetic alterations following irradiation and involved mechanisms in human.
This study aimed to evaluate DNA methylation changes, one of the epigenetic alterations, in normal human blood irradiated with gamma rays in vitro and investigate the effects of low-dose radiation exposure on DNA methylation in peripheral blood DNA from nuclear power plant workers. I also evaluated whether radiation-induced DNA methylation states were associated with genomic instability.
I evaluated global DNA methylation levels using DNA methylation quantification kits and examined the methylation levels of specific locus and repeat element such as long interspersed element-1 (LINE-1) and Satellite 2 (Sat2) by MethyLight assay in human blood. I also evaluated the association between DNA methylation and the frequency of micronuclei or chromosomal aberrations, indicators of genomic instability in radiation exposed workers. The expression of DNA methyltransferases (DNMTs) was measured by using quantitative real-time PCR to investigate the mechanisms of radiation-induced DNA methylation alterations.
In vitro study revealed that g-irradiation led to global DNA hypomethylation in normal human leukocytes and the expression of DNMTs was decreased with g-irradiation. These results indicated that global hypomethylation detected in this study may be the consequence of decreased DNMTs expression by g-irradiation. LINE-1 hypermethylation after 0.5, 1, and 2 Gy of g irradiation was observed in g-ray exposed blood. It was also observed to be significantly methylated in Mut L homologue-1 (MLH1) promoter region in cells irradiated with 2Gy g radiation, and the expression of MLH1 mRNA was also decreased.
This study found that global DNA methylation levels were lower in radiation-exposed workers than in controls. The methylation levels were negatively associated with recent 1.5 year radiation dose, while the levels were increased with total cumulative dose in radiation-exposed workers. Global DNA methylation levels were increased with DNMT1 expression, and the expression levels were correlated with recent 1.5 year radiation dose. These findings suggest that recent 1.5 year radiation exposure caused a decrease of DNMT1 expression, which resulted in global DNA hypomethylation. LINE-1 methylation levels were higher in radiation-exposed workers than in controls, while Sat2 methylation levels had no significant differences between workers and controls. Total cumulative radiation dose was significantly associated with LINE-1 methylation levels after adjusting for age, smoking status and alcohol intake in radiation-exposed workers.
Global DNA methylation levels were also correlated with chromosomal aberrations in workers, while the association was not found in controls. In contrary to this result, the frequency of micronuclei was not correlated with global DNA methylation and repeat element methylation levels. This finding suggests that global DNA hypomethylation is associated with radiation-induced double strand breaks.
These results support the hypothesis that exposure to ionizing radiation may influence DNA methylation in human blood. These findings of this study suggest the possible use of DNA methylation levels as a potential biomarker to monitor IR exposure for radiation-related workers. In addition, the findings of this study support a link between global DNA methylation and genomic instability in radiation-exposed individuals.
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dc.description.tableofcontentsABSTRACT
CONTENTS
LIST OF TABLES
LIST OF FIGURES
CHAPTER I: Introduction
1.Background
1.1. Epigenetic alterations by environmental toxicants
1.2. Ionizing radiation biomarkers
1.3. The studies on radiation-induced DNA methylation and their limitations
1.3.1. Radiation-induced DNA methylation
1.3.2. Radiation-induced DNA methylation and genomic instability
2. Purposes
CHAPTER II: DNA methylation alterations in response to radiation exposure in vitro
1. Introduction
2. Materials and Methods
2.1. Blood culture and Irradiation
2.2. DNA extraction and Bisulfite modification
2.3. RNA extraction and cDNA synthesis
2.4. Assessment of global DNA methylation
2.5. Repeat Element DNA Methylation Assay
2.6. Specific Locus DNA Methylation Assay
2.7. Quantitative real-time PCR for gene expression
2.8. Statistical analysis
3. Results
3.1. Effect of g-radiation on global methylation level in normal human leukocytes
3.2. Effect of g-radiation on repeat element methylation level in normal human leukocytes
3.3. Specific locus DNA methylation
3.4. mRNA expression levels of DNMTs
4. Discussion
4.1. Global methylation changes induced by ionizing radiation
4.2. Repeat element methylation changes induced by ionizing radiation
4.3. Change in MLH1 methylation levels induced by g radiation
CHAPTER III: DNA methylation alterations in the workers occupationally exposed to low dose radiation
1. Introduction
2. Materials and Methods
2.1. Study population and blood collection
2.2. DNA extraction and Bisulfite modification
2.3. RNA extraction and cDNA synthesis
2.4. Assessment of global DNA methylation
2.5. Repeat Element DNA Methylation Assay
2.6. Specific Locus DNA Methylation Assay
2.7. Quantitative RT-PCR to detect DNA methyltransferase mRNAs
2.8. Blood culture for chromosome aberration analysis and cytokinesis-block micronucleus assay
2.9. Chromosome aberration analysis
2.10. Cytokinesis-block micronucleus assay
2.11. Statistical analysis
3. Results
3.1. General characteristics of nuclear power plant workers and controls
3.2. Effect of ionizing radiation on global DNA methylation
3.2.1. Levels of global DNA methylation in workers and controls
3.2.2. Levels of global DNA methylation with radiation exposure in radiation exposed workers
3.2.3. Levels of global DNA methylation with duration of work in radiation exposed workers
3.3. Effect of radiation exposure on repeat element methylation
3.3.1. Levels of repeat element methylation in workers and controls
3.3.2. Levels of repeat element methylation with radiation exposure in radiation exposed workers
3.3.3. Levels of repeat element methylation with duration of work in radiation exposed workers
3.4. Gene-specific promoter methylation changes
3.5. DNMTs expression in response to radiation exposure
3.6. Relationship between genomic instability and DNA methylation patterns
4. Discussion
4.1. Change in global DNA methylation induced by ionizing radiation
4.2. Change in repeat elements methylation induced by ionizing radiation
4.3. The role of DNMTs in radiation-induced methylation changes
4.4. Change in methylation of various genes involved in cellular responses to irradiation
4.5. Association between radiation-induced genomic instability and DNA methylation
4.6. Association between radiation-induced methylation changes and other variables known to affect the methylation
4.7. Summary and Limitation
CHAPTER IV. Overall discussion and conclusion
References
Abstract in Korean (국문초록)
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dc.formatapplication/pdf-
dc.format.extent2394397 bytes-
dc.format.mediumapplication/pdf-
dc.language.isoen-
dc.publisher서울대학교 대학원-
dc.subjectionizing radiation-
dc.subjectDNA methylation-
dc.subjectDNA methyltransferases-
dc.subjectgenomic instability-
dc.subjectchromosome aberrations-
dc.subject.ddc614-
dc.titleDNA methylation as a potential biomarker for radiation exposure-
dc.typeThesis-
dc.description.degreeDoctor-
dc.citation.pagesxiv, 168-
dc.contributor.affiliation보건대학원 보건학과-
dc.date.awarded2014-02-
Appears in Collections:
Graduate School of Public Health (보건대학원)Dept. of Public Health (보건학과)Theses (Ph.D. / Sc.D._보건학과)
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