The mitochondrial phenotype of human mesenchymal stem cells undergoing adipogenic differentiation:
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| 1. Verfasser: | |
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| Format: | Abschlussarbeit Buch |
| Sprache: | English |
| Veröffentlicht: |
2013
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| Schlagworte: | |
| Online-Zugang: | Inhaltsverzeichnis |
| Beschreibung: | III, 133 Bl. Ill., graph. Darst. |
Internformat
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| 245 | 1 | 0 | |a The mitochondrial phenotype of human mesenchymal stem cells undergoing adipogenic differentiation |c von Andreas Hofmann |
| 264 | 1 | |c 2013 | |
| 300 | |a III, 133 Bl. |b Ill., graph. Darst. | ||
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| 502 | |a Dresden, Techn. Univ., Diss., 2013 | ||
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Datensatz im Suchindex
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| adam_text | IMAGE 1
TABLE OF CONTENTS
1. SUMMARY 1
2. INTRODUCTION 3
2.1 HMSC: HUMAN MESENCHYMAL STEM CELLS/STROMAL CELLS 3
2.2 BIOLOGY O F MITOCHONDRIA 6
2.2.1 MITOCHONDRIAL FUNCTIONS 6
2.2.2 STRUCTURE O F MITOCHONDRIA AND MT NETWORK MORPHOLOGY/DISTRIBUTION/
DYNAMICS 6
2.2.3 MT GENETICS AND GENE EXPRESSION 7
2.2.4 MT BIOGENESIS 11
2.2.5 THE OXIDATIVE PHOSPHORYLATION SYSTEM 12
2.3 STEM CELL METABOLISM AND THE STEM CELL NICHE CONCEPT 15
2.4 HYPOXIA AND ITS CONSEQUENCES FOR THE CELL PHYSIOLOGY 17
2.5 AIMS O F THE THESIS AND DEFINITION O F THE MT PHENOTYPE 20
3. RESULTS 21
3.1 ANALYSIS O F HMSC DIFFERENTIATION 21
3.1.1 PROOF O F DIFFERENTIATION CAPABILITY (OSTEOGENIC AND ADIPOGENIC)
21
3.1.2 ADIPOGENIC DIFFERENTIATION O F HMSCS 23
3.2 CHANGES O F THE MITOCHONDRIAL PHENOTYPE DURING ADIPOGENIC
DIFFERENTIATION 24
3.2.1 PHYSIOLOGICAL CHANGES 24
3.2.1.1 OXYGEN CONSUMPTION DURING DIFFERENTIATION 24
3.2.1.2 ROS PRODUCTION DURING DIFFERENTIATION 26
3.2.2 GENETICAL CHANGES 27
3.2.2.1 ANALYSIS O F THE MTDNA COPY NUMBER 27
3.2.2.2 ESTABLISHMENT O F A MLTIWELL GENE EXPRESSION SYSTEM 29
3.2.2.3 EXPRESSION ANALYSIS O F MT BIOGENESIS AND OXPHOS GENES 30
- 3.2.3 CHANGES O F OXPHOS ORGANIZATION - 2D-BN/SDS-PAGE ANALYSES 34
3.2.4 CHANGES O F MT NETWORK MORPHOLOGY 44
3.3 IMPACT O F OXYGEN TENSION ON MT BIOGENESIS AND DIFFERENTIATION 46
3.3.1 ANALYSIS O F RELATIVE MT PROTEIN CONTENT 46
3.3.2 ANALYSIS O F MTDNA COPY NUMBER UNDER HYPOXIC CULTURE CONDITIONS 47
3.3.3 INFLUENCE O F OXYGEN TENSION ON OXPHOS ORGANIZATION 48
3.3.4 EFFICIENCY O F ADIPOGENIC DIFFERENTIATION UNDER HYPOXIA 50
HTTP://D-NB.INFO/1037411552
IMAGE 2
3.4 IMPACT O F DIFFERENTIATION AND OXYGEN TENSION ON THE MT PROTEOME O F
HMSCS-
TWO-DIMENSIONAL DIFFERENTIAL GEL ELECTROPHORESIS (2D-DIGE) 51
4. EXPERIMENTAL OUTLOOK 63
4.1 BN-QMS 63
4.2 GENERATION O F P-HMSCS 64
5. DISCUSSION 67
5.1 PHYSIOLOGICAL PARAMETERS LINKED TO MITOCHONDRIA 67
5.2 GENETICAL PARAMETERS 68
5.3 MT NETWORK MORPHOLOGY 72
5.4 OXPHOS SYSTEM ORGANIZATION 73
5.5 IMPACT O F OXYGEN TENSION ON MT BIOGENESIS AND DIFFERENTIATION 77
5.6 THE IMPACT O F DIFFERENTIATION AND OXYGEN TENSION ON THE MT PROTEOME
80
5.7 CONCLUSION 83
6. MATERIALS AND METHODS 85
6.1 MATERIALS 85
6.1.1 HMSCS (ISOLATION AND CHARACTERIZATION) 85
6.1.2 CHEMICALS AND REAGENTS 85
6.1.3 KITS 88
6.1.4 ANTIBODIES 88
6.1.5 BUFFERS 89
6.1.6 MARKERS 89
6.1.7 PRIMERS 90
6.1.8 MEDIA COMPOSITION 90
6.2 METHODS 91
6.2.1 CELL CULTURE O F HMSCS 91
6.2.2 CELL CULTURE O F HELA CELLS 91
6.2.3 ADIPOGENIC DIFFERENTIATION O F HMSCS 91
6.2.4 OSTEOGENIC DIFFERENTIATION O F HMSCS 92
6.2.5 MICROSCOPY 92
6.2.5.1 OIL RED O STAINING 93
6.2.5.2 ALKALINE PHOSPHATASE STAINING 93
6.2.6 OXYGEN CONSUMPTION MEASUREMENT 93
6.2.7 FLOW CYTOMETRY 94
6.2.8 RELATIVE QUANTIFICATION O F MTDNA COPY NUMBER 94
IMAGE 3
6.2.9 ABSOLUTE QUANTIFICATION O F MTDNA COPY 95
6.2.10 GENE EXPRESSION ANALYSIS 95
6.2.11 DETERMINATION O F RNA QUALITY USING BIOANALYZER 98
6.2.12 ISOLATION O F MITOCHONDRIA 99
6.2.13 DETERMINATION O F PROTEIN CONCENTRATION 99
6.2.14 2D BLUE NATIVE/ SDS POLYACRYLAMIDE GEL ELECTROPHORESIS (PAGE) 99
6.2.15 WESTERN BLOT ANALYSIS 100
6.2.16 HYPOXIC CELL CULTURE 101
6.2.17 TWO DIMENSIONAL DIFFERENTIAL GEL ELECTROPHORESIS (2D-DIGE) 101
6.2.18 ANALYSIS O F THE MT PROTEOME USING 2D-DIGE 105
6.2.19 2D-DIGE ANALYSIS 105
6.2.20 COLLOIDAL COOMASSIE STAINING, SPOT CUTTING AND MASS SPECTROMETRY
(MS) 107
6.2.21 PRODUCTION O F VIRAL PARTICLES 108
6.2.22 VIRAL TRANSDUCTION O F HMSCS 108
6.2.23 DETERMINATION O F TRANSDUCTION EFFICIENCIES AND EVALUATION O F
VIRAL VECTORS .... 109
6.2.24 STATISTICAL ANALYSIS 109
7. REFERENCES 110
8. ABBREVIATIONS ; 122
9. FIGURES AND TABLES 125
10. APPENDIX 127
10.1 THE MITOBIOX QPCR ARRAY 127
10.2 WB EXPOSURE TIME KINETICS 128
10.3 2D-DIGE QUANTIFICATION DATA 129
10.3.1 QUANTITATION TABLES (DELTA2D) 129
10.3.2 EXPRESSION PROFILES (DELTA2D) 132
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| author | Hofmann, Andreas 1981- |
| author_GND | (DE-588)1033684953 |
| author_facet | Hofmann, Andreas 1981- |
| author_role | aut |
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| dewey-hundreds | 600 - Technology (Applied sciences) |
| dewey-ones | 610 - Medicine and health |
| dewey-raw | 610 |
| dewey-search | 610 |
| dewey-sort | 3610 |
| dewey-tens | 610 - Medicine and health |
| discipline | Biologie Medizin |
| format | Thesis Book |
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| spelling | Hofmann, Andreas 1981- Verfasser (DE-588)1033684953 aut The mitochondrial phenotype of human mesenchymal stem cells undergoing adipogenic differentiation von Andreas Hofmann 2013 III, 133 Bl. Ill., graph. Darst. txt rdacontent n rdamedia nc rdacarrier Dresden, Techn. Univ., Diss., 2013 (DE-588)4113937-9 Hochschulschrift gnd-content DNB Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=026051265&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Hofmann, Andreas 1981- The mitochondrial phenotype of human mesenchymal stem cells undergoing adipogenic differentiation |
| subject_GND | (DE-588)4113937-9 |
| title | The mitochondrial phenotype of human mesenchymal stem cells undergoing adipogenic differentiation |
| title_auth | The mitochondrial phenotype of human mesenchymal stem cells undergoing adipogenic differentiation |
| title_exact_search | The mitochondrial phenotype of human mesenchymal stem cells undergoing adipogenic differentiation |
| title_full | The mitochondrial phenotype of human mesenchymal stem cells undergoing adipogenic differentiation von Andreas Hofmann |
| title_fullStr | The mitochondrial phenotype of human mesenchymal stem cells undergoing adipogenic differentiation von Andreas Hofmann |
| title_full_unstemmed | The mitochondrial phenotype of human mesenchymal stem cells undergoing adipogenic differentiation von Andreas Hofmann |
| title_short | The mitochondrial phenotype of human mesenchymal stem cells undergoing adipogenic differentiation |
| title_sort | the mitochondrial phenotype of human mesenchymal stem cells undergoing adipogenic differentiation |
| topic_facet | Hochschulschrift |
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