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DNA methylation alterations in an association between bisphenol A and endometrial condition : 비스페놀 A와 자궁내막질환의 연관성에서 DNA 메틸화 변이의 역할

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dc.contributor.advisor정해원-
dc.contributor.author이선영-
dc.date.accessioned2017-07-13T17:22:30Z-
dc.date.available2017-07-13T17:22:30Z-
dc.date.issued2015-08-
dc.identifier.other000000056823-
dc.identifier.urihttps://hdl.handle.net/10371/120789-
dc.description학위논문 (박사)-- 서울대학교 보건대학원 : 보건학과 분자역학전공, 2015. 8. 정해원.-
dc.description.abstractBisphenol A is one of the endocrine disrupting chemicals and extensively used as protective coatings on food-storage containers, baby bottles, bottle tops, water pipes, and medical equipment. Bisphenol A binds to estrogen receptors, and acts as an estrogen agonist. Bisphenol A also induces epigenetic modifications. Endometrium expressed estrogen receptors, thus it could be a target tissue for bisphenol A. Several studies explored relationship between bisphenol A and gynecologic disorders. However, there are few studies about cytotoxic and epigenetic effects of bisphenol A on endometrium. Therefore, it is necessary to evaluate the relationships between bisphenol A and endometrial disorders.
This study evaluated cytotoxicity and DNA methylation changes in bovine endometrial cells induced by bisphenol A. And urinary bisphenol A concentrations were assessed from women with endometrial disorders, then the relationship was explored between urinary bisphenol A levels and methylation levels of endometrial tissue DNA.
When bovine endometrial cells were treated with 100 μM of bisphenol A for 3 h, the cell viability was significantly decreased. Also, apoptotic cells were significantly induced after 24 h treatment of bisphenol A at 100 μM. S phase arrest was observed after 3 h treatment, and G2/M cell cycle arrest was observed after 24 h treatment. After 1 h treatment, intracellular ROS was significantly increased. DNA damage was induced after 3 h treatment of bisphenol A. After 3 h treatment, global DNA methylation and HOXA 10 methylation levels were decreased. Meanwhile, the methylation level of RASSF1A, one of tumor suppressor genes, was increased after 24 h treatment of 100 μM bisphenol A.
Endometrial tissue DNAs were obtained from 44 women with endometriosis, myoma, or adenomyosis. The relationships between urinary bisphenol A and DNA methylation levels, and the diseases and DNA methylation status were evaluated. There were no significant differences about urinary bisphenol A level among the diseases, also the relationships between urinary bisphenol A levels and DNA methylation status were not observed. However, global DNA methylation level in proliferative phase was significantly higher than that in secretory phase. Sat2 methylation level was significantly higher in adenomyosis than that in myoma. Also, HOXA 10 methylation was significantly associated with endometrial polyps.
In conclusion, bisphenol A can induce cytotoxicity and modify DNA methylation level in bovine endometrial cells. The relations between urinary bisphenol A concentration and endometrial disorders, or urinary bisphenol A concentration and DNA methylation could not find in the epidemiologic study. On the other hand, global DNA hypomethylation was associated with endometrium phase, and endometrial polyps showed HOXA 10 hypermethylation. Therefore, HOXA 10 methylation can be used as a biomarker for endometrial polyp.		-
dc.description.tableofcontentsABSTRACT i
CONTENTS iv
LIST OF TABLES ix
LIST OF FIGURES xii
Chapter 1. Introduction 1
1. Background 2
2. Objective 15
Chapter 2. Cytotoxicity and DNA methylation changes in Bovine Endometrial cells induced by bisphenol A 16
1. Introduction 17
2. Materials and methods 21
2.1. Materials 21
2.2. Cell culture 21
2.3. Evaluation of cell viability 22
2.4. Single-cell gel electrophoresis (comet assay) 22
2.5. Analysis of apoptosis and cell cycle 23
2.6. Measurement of ROS 23
2.7. Extract of DNA and bisulfite modification 24
2.8. Global DNA methylation 24
2.9. Methylation-Specific PCR 24
2.10. Statistical analysis 25
3. Results 27
3.1. Cell viability 27
3.2. Effect of BPA on the viability of BEND cells 29
3.3. Induction of cellular apoptosis by BPA 31
3.4. Cell cycle arrest after treatment with BPA 33
3.5. Increase of intracellular ROS 35
3.6. Extent of DNA damage as indicated by the comet assay 37
3.7. Effect of BPA on global DNA methylation level 39
3.8. Specific locus DNA methylation 41
3.8.1. HOXA 10 (Homeobox 10) 41
3.8.2. RASSF1A (Ras-association domain family 1, isoform A) 43
4. Discussion 45
Chapter 3. DNA methylation changes in human endometrial cells after bisphenol A exposure 51
1. Introduction 52
2. Materials and methods 53
2.1. Materials 53
2.2. Cell culture 53
2.3. Extract of DNA and bisulfite modification 54
2.4. Repetitive element DNA methylation assay 54
2.5. Specific locus DNA methylation assay 55
2.6. Statistical analysis 55
3. Results 56
3.1. The methylation of repetitive elements after BPA exposure 56
3.1.1. The level of LINE-1 methylation 56
3.1.2. The level of Sat2 methylation 58
3.2. Specific locus DNA methylation 59
3.2.1. HOXA 10 methylation level 59
3.2.2. Progesterone receptor (PR) methylation level 60
4. Discussion 61
Chapter 4. Urinary bisphenol A concentrations and DNA methylation alterations in women with endometrial disorders 64
1. Introduction 65
2. Materials and methods 73
2.1. Study design and populations 73
2.2. Urine sample collection and total BPA measurements 74
2.3. Extract of DNA and bisulfite modification 74
2.4. Global DNA methylation 75
2.5. Repetitive element DNA methylation assay 75
2.6. Specific locus DNA methylation assay 76
2.7. Statistical analysis 76
3. Results 78
3.1. General characteristics of study population 78
3.2. Geometric means and quartiles of urinary BPA concentrations 80
3.2.1. Distribution of urinary BPA concentrations 80
3.2.2. Association between BMI and urinary BPA concentration 83
3.3. Effect of BPA on global DNA methylation 87
3.3.1. Levels of global DNA methylation in the disease groups 87
3.3.2. Global DNA methylation level according to age, BMI, parity, smoking status, endometrial phase, and endometrial pathology 89
3.3.3. Correlation between global DNA methylation and urinary BPA concentration 92
3.4. Effect of BPA on repeat element methylation 94
3.4.1. Levels of repeat element methylation in the groups 94
3.4.2. Levels of repeat element methylation according to age, BMI, smoking status, parity, exercise, endometrial phase, and endometrial pathology 97
3.4.3. Levels of repeat element methylation according to BPA level by the disease groups 102
3.5. Effect of BPA on specific DNA methylation 105
3.5.1. Association between BPA levels and HOXA 10 methylation 107
3.5.1.1. Correlation between HOXA 10 methylation and BPA concentration 110
3.5.2. Association between BPA level and PR methylation 112
3.5.2.1. Correlation between PR methylation and BPA concentration 115
3.5.3. Association between BPA levels and PR methylation 117
3.5.3.1. Correlation between RASSF1A methylation and BPA concentration 120
3.6. The level of DNA methylation by endometrium phase 122
3.6.1. Association between global DNA methylation and endometrium phase 122
3.6.2. Association between repeat elements methylation and endometrium phase 123
3.6.3. Association between HOXA 10 methylation and endometrium phase 125
3.6.4. Association between PR methylation and endometrium phase 126
3.6.5. Association between RASSF1A methylation and endometrium phase 127
3.7. The level of DNA methylation by endometrial pathology 128
3.7.1. Association between global DNA methylation and endometrial polyps 128
3.7.2. Association between repeat elements methylation and endometrial polyps 129
3.7.3. Association between HOXA 10 methylation and endometrial polyps 131
3.7.4. Association between PR methylation and endometrial polyps 132
3.7.5. Association between RASSF1A methylation and endometrial polyps 133
4. Discussion 134
Chapter 5. Summary and conclusion 146
References 150
Abstract in Korean 170		-
dc.formatapplication/pdf-
dc.format.extent2769358 bytes-
dc.format.mediumapplication/pdf-
dc.language.isoen-
dc.publisher서울대학교 보건대학원-
dc.subjectbisphenol A-
dc.subjectendometrium-
dc.subjectendometrial cells-
dc.subjectendometriosis-
dc.subjectmyoma-
dc.subjectadenomyosis-
dc.subjectendometrial polyp-
dc.subjectDNA methylation-
dc.subjectHOXA 10-
dc.subject.ddc614-
dc.titleDNA methylation alterations in an association between bisphenol A and endometrial condition-
dc.title.alternative비스페놀 A와 자궁내막질환의 연관성에서 DNA 메틸화 변이의 역할-
dc.typeThesis-
dc.contributor.AlternativeAuthorSunyeong Lee-
dc.description.degreeDoctor-
dc.citation.pages173-
dc.contributor.affiliation보건대학원 보건학과-
dc.date.awarded2015-08-
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