12/15-Lipoxygenase mediates GC-induced anti-inflammatory and LPS-induced pro-inflammatory mechanisms in monocytes:
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| Format: | Thesis Book |
| Language: | English |
| Published: |
2015
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| Subjects: | |
| Online Access: | Inhaltsverzeichnis Inhaltsverzeichnis |
| Physical Description: | 144 S. Ill., graph. Darst. 21 cm |
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| 246 | 1 | 3 | |a Twelve/15-Lipoxygenase mediates GC-induced anti-inflammatory and LPS-induced pro-inflammatory mechanisms in monocytes |
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| adam_text | CONTENTS
CONTENTS
1 INTRODUCTION 1
1.1 THE IMMUNE SYSTEM: PRINCIPLES OF DEFENSE 1
1.1.1 INNATE IMMUNITY 1
1.1.2 ADAPTIVE IMMUNITY 2
1.2 MONOCYTES AND MACROPHAGES 4
1.2.1 ORIGIN, MARKERS AND DIFFERENTIATION OF MONOCYTES/MACROPHAGES 4
1.2.2 FUNCTION OF MONOCYTES AND MACROPHAGES 7
1.3 ROLE OF GC IN MODULATION OF INFLAMMATION/INFLAMMATORY RESPONSE 14
1.3.1 BIOSYNTHESIS AND METABOLISM OF GCS 15
1.3.2 GC ACTIVATION BY THE GC RECEPTOR 15
1.3.3 GR ACTION MODULATE GENE EXPRESSION 16
1.3.4 GCS AFFECT MONOCYTES 18
1.4 12/15-LIPOXYGENASE INFLUENCES MONOCYTE FUNCTION 19
1.4.1 EVOLUTION AND CLASSIFICATION OF LOXS 20
1.4.2 STRUCTURE AND PROPERTIES OF 12/15-LOX 22
1.4.3 12/15-LOX SHOWS DIFFERENT EXPRESSION PATTERN 23
1.4.4 (POST)-TRANSLATIONAL REGULATION OF 12/15-LOX 23
1.4.5 SIGNALING OF 12/15-LOX IN IMMUNE CELLS...:: 24
1.4.6 THE EICOSANOID SWITCH AND THE GENERATION OF ANTI-INFLAMMATORY AND
PRO-RESOLVING
LIPID MEDIATORS.... 25
1.5 THE INFLAMMASOME 28
1.5.1 STRUCTURAL CHARACTERISTICS OF THE INFLAMMASOME 28
1.5.2 ACTIVATION OF THE INFLAMMASOMES 29
1.5.3 CASPASES AND IL-1(3 30
1.6 AIM OF THE STUDY 34
2 MATERIALS AND METHODS 35
2.1 GENERAL LAB MATERIAL 35
2.1.1 CHEMICALS AND REAGENTS 35
2.1.2 DEVICES 37
2.1.3 USED KITS : 38
2.1.4 MOLECULAR WEIGHT STANDARDS 38
2.1.5 BUFFERS 38
HTTP://D-NB.INFO/1074706285
CONTENTS
2.1.6 BACTERIA AND PARASITES 41
2.1.7 MOUSE STRAINS 42
2.1.8 MODIFIED MOUSE STRAINS 42
2.2 METHODS FOR PREPARATION AND CULTIVATION OF MURINE IMMUNE CELLS 43
2.2.1 ISOLATION OF BONE MARROW CELLS 43
2.2.2 DENSITY GRADIENT CENTRIFUGATION 43
2.2.3 CELL COUNTING 43
2.2.4 ISOLATION OF BONE MARROW DERIVED MONOCYTES USING MACS TECHNOLOGY
44
2.2.5 CULTIVATION OF BONE MARROW DERIVED MONOCYTES 45
2.2.6 HARVESTING OF BONE MARROW DERIVED MONOCYTES 45
2.2.7 IN VITRO APPLICATION OF INHIBITORS 45
2.3 MOLECULAR METHODS FOR PREPARATION OF NUCLEIC ACIDS 46
2.3.1 RNA PREPARATION 46
2.3.2 CDNA SYNTHESIS 47
2.3.3 TRANSFECTION OF MONOCYTES USING POLYETHYLENIMINE 49
2.4 BIOCHEMICAL METHODS 50
2.4.1 LYSIS OF CELLS 50
2.4.2 TRICHLOROACETIC ACID PRECIPITATION 50
2.4.3 DETERMINATION OF PROTEIN CONCENTRATION VIA BRADFORD ASSAY 51
2.4.4 SDS-POLYACRYLAMIDE GELELECTROPHORESIS 51
2.4.5 WESTERN BLOTTING 52
2.5 FUNCTIONAL ASSAYS FOR TESTING BONE MARROW DERIVED MONOCYTES 53
2.5.1 ADHESION ASSAY 53
2.5.2 MIGRATION AND CHEMOTAXIS 54
2.5.3 OXIDATIVE BURST 54
2.5.4 PHAGOCYTOSIS OF LATEX BEADS AND CARBOXYLATE-MODIFIED LATEX BEADS
54
2.5.5 OPSONIZATION AND LABELING OF BACTERIA 55
2.5.6 PHAGOCYTOSIS OF BACTERIA 56
2.5.7 KILLING ASSAY 56
2.5.8 NITRIC OXIDE ASSAY 57
2.5.9 CYTOMETRIC BEAD ASSAY 57
2.5.10ENZYME-LMKED IMMUNOSORBENT ASSAY 58
2.5.11SPONTANEOUS AND STAUROSPORINE INDUCED APOPTOSIS 59
2.6 FLUORESCENCE MICROSCOPY AND FLOW CYTOMETRY 60
2.6.1 CYTOSPINS 60
CONTENTS
2.6.2 IMMUNOFLUORESCENCE STAINING 60
2.6.3 FLOW CYTOMETRY 61
2.7 INFLUENZA INFECTION 62
2.7.1 INFLUENZA INFECTION IN VITRO 62
2.7.2 PLAQUE ASSAY 62
2.8 STATISTICAL ANALYSIS 63
3 RESULTS 64
3.1 EXPRESSION OF LOX IN HUMAN AND MURINE MONOCYTES 64
3.1.1 VALIDATING THE RESULTS OF THE AFFYMETRIX MICROARRAY 65
3.1.2 EXPRESSION PROFILE OF 12/15-LOX IN MURINE BM-DERIVED MONOCYTES 65
3.2 FUNCTIONAL ASSAYS DETERMINING DEX-INDUCED EFFECTS MEDIATED BY
12/15-LOX 67
3.2.1 12/15-LOX DID NOT REGULATE PRO-INFLAMMATORY CYTOKINE SECRETION 68
3.2.2 DEX INDUCED CHANGES IN ADHESION AND MIGRATION OF MONOCYTES BUT
THEY WERE NOT
DEPENDENT ON EXPRESSION OF 12/15-LOX 69
3.2.3 THE DEX-INDUCED CHANGES IN PHAGOCYTIC ACTIVITY OF MONOCYTES WERE
INDEPENDENT OF
12/15-LOX 71
3.2.4 PHAGOCYTOSIS OF BACTERIA 73
3.2.5 KILLING OF 5. AUREUS IS INDEPENDENT OF 12/15-LOX 74
3.2.6 DEX-INDUCED REDUCTION OF ROS PRODUCTION WAS DEPENDENT ON 12/15-LOX
75
3.2.7 12/15-LOX MEDIATES DEX-INDUCED UPREGULATION OF ANTI-INFLAMMATORY
GENES 78
3.2.8 12/15-LOX WAS INVOLVED IN REGULATION OF SPONTANEOUS AND DRUG
INDUCED APOPTOTIC
PROCESSES 79
3.3 12/15-LOX COUNTERACTED THE ACTIVITY OF NLRP3 INFLAMMASOME 85
3.3.1 LPS STIMULATION INDUCED HIGHER AMOUNTS OF IL-LP IN SUPERNATANTS OF
12/15-LOX
MONOCYTES IN COMPARISON TO WT CELL AND THIS COULD BE FURTHER ENHANCED BY
ADDITION
OF ATP 85
3.3.2 EXPRESSION OF INFLAMMASOME ASSOCIATED MOLECULES WERE NOT ALTERED
IN 12/15-LOX
7
MONOCYTES 87
3.3.3 THE NLRP3 INFLAMMASOME EXHIBITED HIGH ACTIVATION IN 12/15-LOX
7
MONOCYTES 88
3.3.4 AIM2 INFLAMMASOME ACTIVATION DID NOT ALTER IL-1(3 SECRETION IN
12/15-LOX
MONOCYTES 90
3.3.5 12/15-LOX MONOCYTES PRODUCED HIGH LEVELS OF NO IN RESPONSE TO
LPS 91
3.3.6 THE FPR ANTAGONIST BOC-2 SLIGHTLY ENHANCED IL-IP PRODUCTION IN
12/15-LOX
MONOCYTES 91
3.3.7 ADDITION OF LIPOXINS DIMINISHED IL-LP IN 12/15-LOX DEFICIENT
MONOCYTES 93
CONTENTS
3.3.8 12/15-LOX MONOCYTES SHOWED ELEVATED EXPRESSION OF 5-LOX AND
COX2 IN RESPONSE
TO LPS 94
3.3.9 ELEVATED CASPASE-11 LEVELS HIGHLY INFLUENCE LL-IP PRODUCTION IN
12/15-LOX
MONOCYTES 97
3.4 INFLUENZA INFECTION OF 12/15-LOX AND WT MONOCYTES 99
3.4.1 VIRAL GENE EXPRESSION WAS HIGHER IN WT THAN 12/15-LOX MONOCYTES
100
3.4.2 FPV INFECTION CAUSED HIGH AMOUNTS OF VIRAL PROTEINS IN LYSATES OF
12/15-LOX
7
MONOCYTES 104
3.4.3 12/15-LOX MONOCYTES EXHIBITED HIGH TITERS OF INFLUENZA A VIRUS
FPV DURING 24 H OF
INFECTION 105
4 DISCUSSION 107
4.1 PATTERN OF 12/15-LOX EXPRESSION IN MURINE BONE MARROW MONOCYTES
REFLECTED
THAT OBSERVED IN HUMAN MONOCYTES 107
4.2 INVOLVEMENT OF 12/15-LOX IN DEX-INDUCED MECHANISMS IN MURINE
MONOCYTES .
108
4.2.1 DEX-INDUCED EFFECTS INDEPENDENT FROM 12/15-LOX 109
4.2.2 DEX-INDUCED EFFECTS REGULATED BY 12/15-LOX 114
4.3 12/15-LOX COUNTERACTS THE ACTIVITY OF THE NLRP3 INFLAMMASOME 118
4.4 LOSS OF 12/15-LOX
INCREASED SUSCEPTIBILITY TO INFLUENZA A INFECTION 125
5 SUMMARY 129
6 REFERENCES 131
7 CURRICULUM VITAE 142
8 DANKSAGUNG 143
|
| any_adam_object | 1 |
| author | Kusche, Yvonne 1985- |
| author_GND | (DE-588)1073722384 |
| author_facet | Kusche, Yvonne 1985- |
| author_role | aut |
| author_sort | Kusche, Yvonne 1985- |
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| building | Verbundindex |
| bvnumber | BV042939417 |
| ctrlnum | (OCoLC)922065671 (DE-599)DNB1074706285 |
| dewey-full | 610 |
| 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 | Medizin |
| format | Thesis Book |
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| physical | 144 S. Ill., graph. Darst. 21 cm |
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| spelling | Kusche, Yvonne 1985- Verfasser (DE-588)1073722384 aut 12/15-Lipoxygenase mediates GC-induced anti-inflammatory and LPS-induced pro-inflammatory mechanisms in monocytes vorgelegt von Yvonne Kusche Twelve/15-Lipoxygenase mediates GC-induced anti-inflammatory and LPS-induced pro-inflammatory mechanisms in monocytes 2015 144 S. Ill., graph. Darst. 21 cm txt rdacontent n rdamedia nc rdacarrier Münster (Westfalen), Univ., Diss., 2015 (DE-588)4113937-9 Hochschulschrift gnd-content B:DE-101 application/pdf http://d-nb.info/1074706285/04 Inhaltsverzeichnis DNB Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=028365949&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Kusche, Yvonne 1985- 12/15-Lipoxygenase mediates GC-induced anti-inflammatory and LPS-induced pro-inflammatory mechanisms in monocytes |
| subject_GND | (DE-588)4113937-9 |
| title | 12/15-Lipoxygenase mediates GC-induced anti-inflammatory and LPS-induced pro-inflammatory mechanisms in monocytes |
| title_alt | Twelve/15-Lipoxygenase mediates GC-induced anti-inflammatory and LPS-induced pro-inflammatory mechanisms in monocytes |
| title_auth | 12/15-Lipoxygenase mediates GC-induced anti-inflammatory and LPS-induced pro-inflammatory mechanisms in monocytes |
| title_exact_search | 12/15-Lipoxygenase mediates GC-induced anti-inflammatory and LPS-induced pro-inflammatory mechanisms in monocytes |
| title_full | 12/15-Lipoxygenase mediates GC-induced anti-inflammatory and LPS-induced pro-inflammatory mechanisms in monocytes vorgelegt von Yvonne Kusche |
| title_fullStr | 12/15-Lipoxygenase mediates GC-induced anti-inflammatory and LPS-induced pro-inflammatory mechanisms in monocytes vorgelegt von Yvonne Kusche |
| title_full_unstemmed | 12/15-Lipoxygenase mediates GC-induced anti-inflammatory and LPS-induced pro-inflammatory mechanisms in monocytes vorgelegt von Yvonne Kusche |
| title_short | 12/15-Lipoxygenase mediates GC-induced anti-inflammatory and LPS-induced pro-inflammatory mechanisms in monocytes |
| title_sort | 12 15 lipoxygenase mediates gc induced anti inflammatory and lps induced pro inflammatory mechanisms in monocytes |
| topic_facet | Hochschulschrift |
| url | http://d-nb.info/1074706285/04 http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=028365949&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |
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