Expression, function and regulation of methylthioadenosine phosphorylase in the pathogenesis of chronic liver disease:
Gespeichert in:
| Beteilige Person: | |
|---|---|
| Format: | Hochschulschrift/Dissertation Buch |
| Sprache: | Englisch |
| Veröffentlicht: |
2013
|
| Schlagwörter: | |
| Links: | http://epub.uni-regensburg.de/28956/ https://nbn-resolving.org/urn:nbn:de:bvb:355-epub-289569 http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=026842221&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |
| Umfang: | III, 89 Bl. Ill., graph. Darst. |
Internformat
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| 245 | 1 | 0 | |a Expression, function and regulation of methylthioadenosine phosphorylase in the pathogenesis of chronic liver disease |c von Barbara Czech |
| 264 | 1 | |c 2013 | |
| 300 | |a III, 89 Bl. |b Ill., graph. Darst. | ||
| 336 | |b txt |2 rdacontent | ||
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| 502 | |a Regensburg, Univ., Diss., 2013 | ||
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Datensatz im Suchindex
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| DE-BY-TUM_katkey | 1959730 |
| _version_ | 1831256899374284801 |
| adam_text | TABLE OF CONTENTS
1 ZUSAMMENFASSUNG 1
2 SUMMARY 2
3 INTRODUCTION 3
3.1 LIVER FUNCTION AND PHYSIOLOGY IN HEALTH AND DISEASE 3
3.1.1 LIVER ANATOMY 3
3.1.2 LIVER FUNCTION 4
3.2 CHRONIC LIVER DISEASE 5
3.2.1 FIBROSIS 5
3.2.2 CIRRHOSIS 6
3.3 TRIGGER OF CHRONIC LIVER DISEASE 6
3.3.1 NON ALCOHOLIC FATTY LIVER DISEASE (NAFLD) 7
3.3.2 EFFECTS OF REACTIVE OXYGEN SPECIES (ROS) ON FIBROSIS 8
3.3.2.1 EFFECTS OF ROS FORMATION ON METHYLATION 8
3.4 CHARACTERISATION OF POLYAMINE METABOLISM AND METHIONINE PATHWAYS 9
3.4.1 METHYLTHIOADENOSINE PHOSPHORYLASE (MTAP) 9
3.4.1.1 MTAP STRUCTURE 10
3.4.1.2 MTAP EXPRESSION IN VIVO 10
3.4.1.3 EFFECT OF MTAP MANIPULATION IN VIVO AND IN VITRO 11
3.4.1.4 REGULATION OF MTAP 12
3.4.2 5 -DEOXY-5 -(METHYLTHIO)ADENOSINE (MTA)- STRUCTURE AND ABUNDANCE..
12
3.4.2.1 EFFECTS OF MTA IN VIVO AND IN VITRO 14
3.4.3 ADOMET: MAT 15
3.5 AIM OF THE STUDY 17
4 MATERIALS AND METHODS 18
4.1 CHEMICALS AND REAGENTS 18
4.2 LABORATORY EXPENDABLES 19
4.3 LABORATORY INSTRUMENTS 19
4.4 BUFFERS 21
4.5 PLASMIDS 21
4.6 WORKING WITH BACTERIA 21
4.6.1 BACTERIAL STRAINS 21
4.6.2 LIQUID MEDIA AND AGAR PLATES 21
HTTP://D-NB.INFO/1045157899
TABLE OF CONTENTS II
4.6.3 BACTERIAL CULTURE 22
4.6.4 TRANSFORMATION 22
4.6.5 ISOLATION OF PLASMID DNA (MINI AND MIDI PREPARATION) 22
4.7 CELL CULTURE 23
4.7.1 CELL CULTURE MEDIUM 23
4.7.2 CULTIVATION OF CELL LINES 23
4.7.3 CELL LINE OF ACTIVATED MURINE HEPATIC STELLATE CELLS 24
4.7.4 ISOLATION OF PRIMARY HUMAN HEPATOCYTES 24
4.7.5 ISOLATION OF HEPATIC STELLATE CELLS 24
4.7.6 DETERMINATION OF CELL NUMBER AND VIABILITY 25
4.7.7 FREEZING CELLS FOR STORAGE 25
4.7.8 TRANSIENT SIRNA TRANSFECTION 26
4.7.9 TRANSIENT PLASMID TRANSFECTION 26
4.7.10 LUCIFERASE REPORTER GENE ASSAY 27
4.8 ISOLATION AND ANALYSIS OF RNA 27
4.8.1 RNA ISOLATION AND DETERMINATION OF RNA CONCENTRATION 27
4.8.2 REVERSE TRANSCRIPTION OF RNA TO CDNA 28
4.8.3 QUANTITATIVE REAL TIME POLYMERASE CHAIN REACTION 28
4.9 PROTEIN ANALYSIS 30
4.9.1 PREPARATION OF PROTEIN EXTRACTS 30
4.9.2 DETERMINATION OF PROTEIN CONCENTRATION 30
4.9.3 SDS POLYACRYLAMID GEL ELECTROPHORESIS (SDS-PAGE) 31
4.9.4 WESTERN BLOTTING 32
4.9.5 QUANTIFICATION OF CASPASE-3/7 ACTIVITY 33
4.9.6 5 -DEOXY-5 -(METHYLTHIO)ADENOSINE (MTA) EXTRACTION AND ANALYSIS..
33
4.10 FLOW CYTOMETRY 34
4.10.1 ANNEXIN V / PROPIDIUM IODIDE DOUBLE STAINING 34
4.11 MEASUREMENT OF REACTIVE OXYGEN FORMATION (ROS) 36
4.12 ANIMAL EXPERIMENTS 36
4.12.1 BDL 36
4.12.2 NASH 36
4.13 HISTOLOGY AND IMMUNOHISTOCHEMISTRY 36
4.13.1 HISTOLOGY AND IMMUNOHISTOCHEMISTRY 36
4.13.2 IMMUNOFLUORESCENT CELL STAINING 37
TABLE OF CONTENTS III
4.14 STATISTICAL ANALYSIS 37
5 RESULTS 39
5.1 EXPRESSION AND FUNCTION OF MTAP IN CHRONIC LIVER DISEASE 39
5.1.1 MTAP EXPRESSION IN CHRONIC LIVER DISEASE 39
5.1.2 MTAP EXPRESSION IN NON-ALCOHOLIC STEATOEPATITIS (NASH) 40
5.2 MTAP EXPRESSION IN HEPATIC STELLATE CELLS 43
5.3 FUNCTIONAL ROLE OF MTAP IN ACTIVATED HEPATIC STELLATE CELLS 47
5.3.1 FUNCTIONAL ROLE OF MTAP SUPPRESSION 47
5.4 EFFECT OF MTA ON HEPATIC STELLATE CELLS 52
5.4.1 DURING ACTIVATION 52
5.4.2 IN ACTIVATED HSCS 52
5.4.3 FUNCTIONAL EFFECT OF MTAP/MTA ON SURVIVIN EXPRESSION IN ACTIVATED
HEPATIC STELLATE CELLS 55
5.4.4 ROLE OF METHYLATION AN THE GENERATION OF ROS ON MTAP REGULATION IN
ACTIVATED HSCS 58
6 DISCUSSION 62
7 REFERENCES 65
8 ABBREVIATIONS 78
9 APPENDIX 82
9.1 CURRICULUM VITAE 82
9.2 ADVANCED TRAINING AND COURSES 83
9.3 PUBLICATIONS 83
9.4 PRESENTATIONS 84
9.5 AWARDS/ GRANTS 85
9.6 DANKSAGUNG 86
9.7 SELBSTSTANDIGKEITSERKLARUNG 89
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| indexdate | 2024-12-20T16:46:27Z |
| institution | BVB |
| language | English |
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| physical | III, 89 Bl. Ill., graph. Darst. |
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| publishDate | 2013 |
| publishDateSearch | 2013 |
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| spellingShingle | Czech, Barbara 1983- Expression, function and regulation of methylthioadenosine phosphorylase in the pathogenesis of chronic liver disease Pathogenese (DE-588)4115512-9 gnd Leberkrankheit (DE-588)4034937-8 gnd Phosphorylase (DE-588)4359674-5 gnd Chronische Krankheit (DE-588)4010176-9 gnd |
| subject_GND | (DE-588)4115512-9 (DE-588)4034937-8 (DE-588)4359674-5 (DE-588)4010176-9 (DE-588)4113937-9 |
| title | Expression, function and regulation of methylthioadenosine phosphorylase in the pathogenesis of chronic liver disease |
| title_auth | Expression, function and regulation of methylthioadenosine phosphorylase in the pathogenesis of chronic liver disease |
| title_exact_search | Expression, function and regulation of methylthioadenosine phosphorylase in the pathogenesis of chronic liver disease |
| title_full | Expression, function and regulation of methylthioadenosine phosphorylase in the pathogenesis of chronic liver disease von Barbara Czech |
| title_fullStr | Expression, function and regulation of methylthioadenosine phosphorylase in the pathogenesis of chronic liver disease von Barbara Czech |
| title_full_unstemmed | Expression, function and regulation of methylthioadenosine phosphorylase in the pathogenesis of chronic liver disease von Barbara Czech |
| title_short | Expression, function and regulation of methylthioadenosine phosphorylase in the pathogenesis of chronic liver disease |
| title_sort | expression function and regulation of methylthioadenosine phosphorylase in the pathogenesis of chronic liver disease |
| topic | Pathogenese (DE-588)4115512-9 gnd Leberkrankheit (DE-588)4034937-8 gnd Phosphorylase (DE-588)4359674-5 gnd Chronische Krankheit (DE-588)4010176-9 gnd |
| topic_facet | Pathogenese Leberkrankheit Phosphorylase Chronische Krankheit Hochschulschrift |
| url | http://epub.uni-regensburg.de/28956/ https://nbn-resolving.org/urn:nbn:de:bvb:355-epub-289569 http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=026842221&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |
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