Identification of atheroprone shear stress responsiv enhancer elements in endothelial cells:
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| Format: | Abschlussarbeit Buch |
| Sprache: | English |
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2017
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| Beschreibung: | 126 Blätter Illustrationen 30 cm |
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| 245 | 1 | 0 | |a Identification of atheroprone shear stress responsiv enhancer elements in endothelial cells |c vorgelegt von Olga Bondareva |
| 264 | 1 | |a Münster |c 2017 | |
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Datensatz im Suchindex
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| adam_text | TABLE OF CONTENT
SUMMARY........................................................................
4
ACKNOWLEDGMENTS....................................................................................................................................
6
LIST OF
ABBREVIATIONS.................................................................................................................................9
1.
INTRODUCTION.................................................................................................................................
11
1.1. SHEAR STRESS AND
ENDOTHELIUM...................................................................................................11
1.1.1. ENDOTHELIUM - A DYNAMIC INTERFACE BETWEEN CIRCULATING FLUID AND
ORGANS
................
11
1.1.2. DIFFERENT SHEAR STRESS PROFILES CAUSE MORPHOLOGICAL AND
FUNCTIONAL CHANGES IN
ENDOTHELIAL
CELLS..................................................................................................................................
11
1.2. EPIGENETIC MECHANISMS OF GENE
REGULATION............................................................................
13
1.2.1. DNA
METHYLATION.................................................................................................................14
1.2.2. HISTONE
MODIFICATIONS........................................................................................................
14
1.2.3. ENHANCERS AS CENTRAL REGULATORY ELEMENTS IN GENOME
...................................................
17
1.3. SHEAR STRESS INDUCES EPIGENETIC REPROGRAMMING OF ENDOTHELIAL CELLS
.................................
22
1.3.1. ATHEROSCLEROSIS AS A MAJOR BURDEN IN MODERN
SOCIETY.................................................. 22
1.3.2. EPIGENETIC CHANGES FOUND IN ATHEROSCLEROTIC VESSELS
.....................................................
25
1.4. AIM OF
STUDY.................................................................................................................................
27
2. MATERIALS AND
METHODS.............................................................................................................
28
2.1. MATERIALS AND
EQUIPMENT......................................................................................................
28
2.2. BUFFERS AND
SOLUTIONS.............................................................................................................
31
2.3. CELL
CULTURE...............................................................................................................................34
2.4. SHEAR STRESS
EXPERIMENTS.......................................................................................................
35
2.5. REAL-TIME
PCR.........................................................................................................................
36
2.6. WESTERN
BLOT...........................................................................................................................
40
2.7. BRDU CELL PROLIFERATION
ASSAY..................................................................................................43
2.8. FLUORESCENT
MICROSCOPY.........................................................................................................44
2.9. CHROMATIN IMMUNOPRECIPITATION
(CHIP)..............................................................................
45
2.10. CHIP AND RNA SEQUENCING (CHIP-SEQ AND
RNA-SEQ)............................................................49
2.11. BIOINFORMATICAL
ANALYSIS.........................................................................................................
49
2.12. CLONING AND PLASMID
PRODUCTION...........................................................................................
51
2.5. LUCIFERASE
ASSAY......................................................................................................................
55
2.6. STATISTICAL
ANALYSIS...................................................................................................................56
3.
RESULTS..........................................................................................................................................
57
3.1. RNA-SEQ
DATA...............................................................................................................................57
3.1.1. PRE-SEQUENCING RNA QUALITY CONTROL BY
RT-PCR.............................................................. 57
3.1.2. QUALITY CONTROLS OF THE RAW DATA AFTER
SEQUENCING.........................................................58
3.1.3. DIFFERENTIALLY EXPRESSED GENES IN OSCILLATORY VERSUS STATIC SHEAR
STRESS TREATED HUVEC
61
3.1.4. VALIDATION WITH
RT-PCR.......................................................................................................
64
3.2. CHIP-SEQ FOR
H3K27AC.................................................................................................................66
3.2.1. PRE-SEQUENCING QUALITY CONTROLS OF THE CHIP
...................................................................
67
3.2.2. QUALITY CONTROLS OF THE SEQUENCING
DATA..........................................................................
69
3.2.3. PEAK CALLING AND IDENTIFICATION OF DIFFERENTIAL PEAKS IN
OSCILLATORY VERSUS STATIC
CONDITIONS.............................................................................................................................................72
3.2.4. CHIP-SEQ VALIDATION -
HEATMAPS.......................................................................................73
3.2.5. ANNOTATION OF PEAKS TO THE NEARBY
GENES........................................................................75
3.2.6. DAVID GO
ANNOTATION........................................................................................................
77
3.2.7. TF BINDING MOTIF
ANALYSIS....................................................................................................84
3.3. API TRANSCRIPTION FACTOR ACTIVATION UNDER UNIDIRECTIONAL VERSUS
OSCILLATORY SHEAR STRESS .89
3.3.1. API TRANSCRIPTION FACTOR FAMILY MEMBERS EXPRESSION
...................................................
89
3.3.2. JUN ACTIVATION BY PHOSPHORYLATION AND NUCLEAR TRANSLOCATION
.....................................
90
3.3.3. API ACTIVATION BY LUCIFERASE
ASSAY....................................................................................92
3.4. YAP/TAZ TRANSCRIPTION FACTOR ACTIVATION IN HUVEC BY OSCILLATORY
SHEAR STRESS
....................
94
3.4.1. YAP/TAZ TARGET GENES
EXPRESSION.....................................................................................94
3.4.2. PROLIFERATION RATE OF HUVEC SUBJECTED TO 6 H OF SHEAR STRESS
.......................................
95
3.5. LUCIFERASE ASSAY VALIDATION OF
ENHANCERS................................................................................
98
3.5.1. VALIDATION OF THE OSCILLATORY SHEAR STRESS RESPONSIVE ENHANCERS
..................................
98
3.5.2. MUTATION OF TF BINDING SITES IN THE OSCILLATORY SHEAR STRESS
RESPONSIVE ENHANCERS .100
4.
DISCUSSION...................................................................................................................................103
4.1. TRANSCRIPTOMIC AND EPIGENOMIC ANALYSIS OF ENDOTHELIAL RESPONSE TO
OSCILLATORY SHEAR
STRESS 104
4.2. API TRANSCRIPTION FACTOR ACTIVATION BY SHEAR
STRESS..........................................................108
4.3. YAP/TAZ ACTIVATION BY OSCILLATORY SHEAR STRESS IN
HUVEC..................................................109
REFERENCES.............................................................................................................................................114
LEBENSLAUF..............................................................................................................................................125
PUBLICATIONS, CONFERENCE
ATTENDANCE.................................................................................................126
8
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| any_adam_object | 1 |
| author | Bondareva, Olga 1992- |
| author_GND | (DE-588)1154451623 |
| author_facet | Bondareva, Olga 1992- |
| author_role | aut |
| author_sort | Bondareva, Olga 1992- |
| author_variant | o b ob |
| building | Verbundindex |
| bvnumber | BV045218977 |
| ctrlnum | (OCoLC)1136403825 (DE-599)DNB1154795756 |
| discipline | Medizin |
| format | Thesis Book |
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| spelling | Bondareva, Olga 1992- Verfasser (DE-588)1154451623 aut Identification of atheroprone shear stress responsiv enhancer elements in endothelial cells vorgelegt von Olga Bondareva Münster 2017 126 Blätter Illustrationen 30 cm txt rdacontent n rdamedia nc rdacarrier Dissertation Westfälische Wilhelms-Universität Münster 2018 Beschränkt für den Austausch (DE-588)4113937-9 Hochschulschrift gnd-content B:DE-101 application/pdf http://d-nb.info/1154795756/04 Inhaltsverzeichnis DNB Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=030607616&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Bondareva, Olga 1992- Identification of atheroprone shear stress responsiv enhancer elements in endothelial cells |
| subject_GND | (DE-588)4113937-9 |
| title | Identification of atheroprone shear stress responsiv enhancer elements in endothelial cells |
| title_auth | Identification of atheroprone shear stress responsiv enhancer elements in endothelial cells |
| title_exact_search | Identification of atheroprone shear stress responsiv enhancer elements in endothelial cells |
| title_full | Identification of atheroprone shear stress responsiv enhancer elements in endothelial cells vorgelegt von Olga Bondareva |
| title_fullStr | Identification of atheroprone shear stress responsiv enhancer elements in endothelial cells vorgelegt von Olga Bondareva |
| title_full_unstemmed | Identification of atheroprone shear stress responsiv enhancer elements in endothelial cells vorgelegt von Olga Bondareva |
| title_short | Identification of atheroprone shear stress responsiv enhancer elements in endothelial cells |
| title_sort | identification of atheroprone shear stress responsiv enhancer elements in endothelial cells |
| topic_facet | Hochschulschrift |
| url | http://d-nb.info/1154795756/04 http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=030607616&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |
| work_keys_str_mv | AT bondarevaolga identificationofatheroproneshearstressresponsivenhancerelementsinendothelialcells |
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