Holdings: Role of Hypoxia-Inducible Factor (HIF) 1α in Dendritic Cells in Immune Regulation of Atherosclerosis

Role of Hypoxia-Inducible Factor (HIF) 1α in Dendritic Cells in Immune Regulation of Atherosclerosis: = Rolle von Hypoxie induziertem Faktor (HIF) 1α in dendritischen Zellen in der Immunregulation der Atherosklerose

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Bibliographic Details
Main Author:	Chaudhari, Sweena M. 1985- (Author)
Format:	Thesis Book
Language:	English
Published:	Würzburg 2013
Subjects:	Hypoxie Entzündung Arteriosklerose Dendritische Zelle Hypoxia Atherosclerosis Dendritic cells Hochschulschrift
Online Access:	Volltext http://d-nb.info/1125372370/34 Inhaltsverzeichnis
Item Description:	veröffentlicht: 2017
Physical Description:	162 Blätter Illustrationen, Diagramme
Format:	Langzeitarchivierung gewährleistet, LZA

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adam_text	ROLE OF HYPOXIA-INDUCIBLE FACTOR (HIF) 1A IN DENDRITIC CELLS IN IMMUNE REGULATION OF ATHEROSCLEROSIS ROLLE VON HYPOXIE INDUZIERTEM FAKTOR (HIF) 1A IN DENDRITISCHEN ZELLEN IN DER IMMUNREGULATION DER ATHEROSKLEROSE DOCTORAL THESIS FOR A DOCTORAL DEGREE AT THE GRADUATE SCHOOL OF LIFE SCIENCES JULIUS-MAXIMILIANS-UNIVERSITAET WUERZBURG SECTION BIOMEDICINE SUBMITTED BY SWEENA M. CHAUDHARI FROM MUMBAI, INDIA WUERZBURG, 2013 TABLE OF CONTENTS S U M M A R Y ..................................................................................................................... 10 ZUSAM M ENFASSUNG ............................................................................................... 12 ABBREVIATIONS..........................................................................................................14 1. IN TR O D U C TIO N .................................................................................................... 20 1.1 DENDRITIC CELLS......................................................................................................... 20 1.1.1 ORIGIN OF DENDRITIC CELLS...............................................................................20 1.1.2 DC MARKERS AND SUBTYPES......................................................................... 21 1.1.3 RELATION OF DCS TO MONOCYTES AND MACROPHAGES ............................... 22 1.1.4 CD11C MARKER AND THE CD11C-ERE MOUSE M ODEL ................................ 22 1.1.5 FUNCTIONS OF D CS ....................................................................... 23 1.1.6 DCS REGULATE HELPER T-CELL BALANCE.........................................................24 1.1.7 DC MIGRATION................................................................................................ 26 1.2 ATHEROSCLEROSIS....................................................................................................... 27 1.2.1 ATHEROSCLEROSIS - A GLOBAL BURDEN .......................................................... 27 1.2.2 PATHOGENESIS OF ATHEROSCLEROSIS ............................................................. 27 1.2.3 MONOCYTES AND MACROPHAGES IN ATHEROSCLEROSIS.................................28 1.2.4 DENDRITIC CELLS IN ATHEROSCLEROSIS ............................................................. 29 1.2.5 T CELLS IN ATHEROSCLEROSIS........................................................................... 30 1.2.6 MOUSE MODELS TO STUDY ATHEROGENESIS.................................................. 32 1.3 HYPOXIA-INDUCIBLE FACTOR 1 A ................................................................................. 33 1.3.1 HYPOXIA AND OXYGEN HOMEOSTASIS IN CELLS ............................................. 33 1.3.2 BIOLOGY OF THE HYPOXIA-INDUCIBLE FACTORS.................................................34 1.3.3 NON-HYPOXIC REGULATION OF H IF IA ............................................................. 36 1.3.4 H IF IA IN IM M UNITY- LYMPHOID CELLS.........................................................36 1.3.5 H IF IA IN IMMUNITY - MYELOID CELLS........................................................... 37 1.3.6 H IF IA IN IMMUNITY - DENDRITIC CELLS..........................................................38 1.3.7 H IF IA IN INNATE IMMUNITY ...... . ....................................................................39 1.3.8 H IF IA IN PHYSIOLOGY AND DISEASE .................................. 40 1.3.9 HYPOXIA IN ATHEROSCLEROSIS.........................................................................41 1.3.10 ROLE OF H IF IA IN MYELOID CELLS IN ATHEROSCLEROSIS .............................. 42 1.3.11 ROLE OF H IF IA IN T CELLS IN ATHEROSCLEROSIS ......................................... 42 1.3.12 NON-HYPOXIC TRIGGERS OF H IF IA LEADING TO ATHEROSCLEROSIS .............. 43 1.4 AIM OF THIS PROJECT................................................................................................ 43 2. M A TER IA LS..............................................................................................................45 2.1 CHEMICALS................................................................................................................ 45 2.2 KITS AND REAGENTS....................................................................................................46 2.2.1 KITS................................................................................................................. 46 2.2.2 MOLECULAR BIOLOGY REAGENTS.......................................................................46 2.2.3 CELL CULTURE REAGENTS.................................................................................. 47 2.3 CYTOKINES................................................................................................................. 47 2.4 ANTIBODIES................................................................................................................ 48 2.4.1 FLUORESCENT ANTIBODIES...............................................................................48 2.4.2 PRIMARY ANTIBODIES.......................................................................................48 2.4.3 SECONDARY ANTIBODIES AND DETECTION REAGENTS ...................................... 49 2.5 PRIMERS.....................................................................................................................49 2.5.1 QPCR PRIMERS...............................................................................................49 2.5.2 GENOTYPING PRIMERS.....................................................................................50 2.5.3 CHIP PRIMERS................................................................................................ 51 2.6 PLASMIDS...................................................................................................................51 2.7 ANIMALS.....................................................................................................................51 2.8 CELL LINES...................................................................................................................51 2.9 PLASTICWARE, GLASSWARE AND OTHER CONSUMABLES ............................................. 52 2.10 INSTRUMENTS... ........................................ 53 2.11 SOFTWARES............................................................................................................... 54 2.12 BUFFERS, SOLUTIONS AND M ED IA ............................................................................ 54 2.12.1 BUFFERS......................................................................................................... 54 2.12.2 SOLUTIONS......................................................................................................56 2.12.3 MEDIA........................................................................................................... 57 3. M ETHO DS................................................................................................................. 59 3.1 ANIMAL WORK........................................................................................................... 59 3.1.1 MICE EXPERIMENTS....................... 59 3.1.2 GENERATION OF CD11C-CRE+ HIF1ANOX/I]* LDLR1 TRANSGENIC M IC E .............. 59 3.1.3 GENERATION OF LYSM-CRE+ H/7FAFLOX/FLOX LD/RV TRANSGENIC M ICE ............... 59 3.1.4 GENERATION OF TRANSGENIC MICE WITH STATS OVEREXPRESSION IN DCS 60 3.1.5 SACRIFICING MICE AND HARVESTING ORGANS..................................................60 6 3.1.6 PROCESSING OF ORGANS ................................................................................ 60 3.2 HISTOLOGY...................................................................................................................61 3.2.1 OIL-RED O STAINING OF AORTAS FOR PLAQUE ANALYSIS.................................61 3.2.2 ANALYSIS OF AORTIC PLAQUE A RE A ................................................................. 62 3.2.3 PREPARATION OF AORTIC ROOTS FOR PARAFFIN SECTIONING ............................... 62 3.2.4 DE-PARAFFINIZATION OF AORTIC ROOTS...............................................................62 3.2.5 COLLAGEN-ELASTIN STAINING OF AORTIC ROOTS FOR PLAQUE ANALYSIS ............. 63 3.2.6 ANALYSIS OF AORTIC ROOT PLAQUE A RE A ......................................................... 63 3.2.7 ANTIGEN RETRIEVAL OF AORTIC ROOTS.................................................................64 3.2.8 MACROPHAGE-SMOOTH MUSCLE CELL STAINING OF AORTIC ROOTS ................. 64 3.2.9 T-CELL STAINING OF AORTIC ROOTS ..................................................................... 64 3.2.10 PREPARATION OF AORTIC ROOTS FOR FROZEN SECTIONING ............................... 65 3.2.11 CD11C+ CELL STAINING OF AORTIC ROOTS ...................................................... 65 3.2.12 ANALYSIS OF AORTIC ROOT PLAQUE CELL CONTENT..........................................66 3.2.13 CD11C+ CELL AND HIF1A STAINING OF AORTIC ROOTS..................................67 3.2.14 DETECTION OF HYPOXIA................................................................................. 68 3.3 CELL CULTURE TECHNIQUES......................................................................................... 69 3.3.1 BONE MARROW DERIVED DENDRITIC CELL (BMDC) CULTURE ........................... 69 3.3.2 BONE MARROW DERIVED MACROPHAGE (BMM) CULTURE .............................. 69 3.3.3 STIMULATION OF CULTURED CELLS.......................................................................70 3.3.4 ISOLATION OF SPLENIC D C S ............................................................................. 70 3.3.5 ISOLATION OF SPLENIC CD4+ T CELLS .............................................................. 70 3.3.6 ISOLATION OF NAIVE CD4+ T CELLS..................................................................71 3.3.7 ISOLATION OF LINEAGE DEPLETED BM CELLS .................................................... 71 3.3.8 ANTIGEN SPECIFIC T-CELL PROLIFERATION..........................................................71 3.3.9 ANTIGEN SPECIFIC T-CELL POLARIZATION..........................................................72 3.3.10 BMDC TRANSMIGRATION ASSAY...................................................................72 3.3.11 TRANSFECTION OF DCS WITH INHIBITION AND OVEREXPRESSION PLASMIDS.72 3.3.12 MAINTENANCE OF HEK293F CELLS ............................................................. 72 3.4 NUCLEIC ACID DETECTION TECHNIQUES......................................................................73 3.4.1 GENOMIC DNA ISOLATION ............................................................................. 73 3.4.2 TOTAL RNA ISOLATION ..................................................................................... 73 3.4.3 CDNA SYNTHESIS........................................................................................... 73 3.4.4 CONVENTIONAL PCR FOR GENOTYPING...........................................................74 3.4.5 QUANTITATIVE REAL TIME P C R ......................................................................75 3.5 PROTEIN DETECTION TECHNIQUES...............................................................................76 3.5.1 FLOW CYTOMETRY.............................................................................................76 3.5.2 ELISA .............................................................................................................77 3.6 LIPID DETECTION TECHNIQUES................................................................................... 77 3.6.1 CHOLESTEROL ASSAY........................................................................................ 77 3.6.2 TRIGLYCERIDE A S SAY.......................................................................................78 3.7 BACTERIAL TECHNIQUES..............................................................................................78 3.7.1 ELUTING PLASMIDS FROM FILTER PAPER SPOTS.................................................78 3.7.2 PREPARATION OF LB PLATES ............................................................................ 78 3.7.3 TRANSFORMATION OF BACTERIAL CELLS...............................................................78 3.7.4 GROWTH OF TRANSFORMED CELLS FOR PLASMID DNA ISOLATION ..................... 78 3.7.5 PLASMID DNA ISOLATION................................................................................79 3.7.6 LENTIVIRAL VECTOR - CRE VECTOR CO-TRANSFECTION ...................................... 79 3.8 LENTIVIRUS TECHNIQUES.............................................................................................79 3.8.1 TRANSFECTION OF HEK293F CELLS FOR LENTIVIRUS PRODUCTION ...................... 79 3.8.2 HARVESTING OF LENTIVIRUS...............................................................................79 3.8.3 VIRUS TITRATION................................................................................................ 80 3.8.4 VIRAL TRANSDUCTION OF BM CELLS FOR IN VITRO ANALYSIS ................................. 80 3.8.5 VIRAL TRANSDUCTION OF LIN- BM CELLS FOR IN VIVO TRANSPLANTATION ............... 81 3.9 TRANSCRIPTION FACTOR BINDING SITE ANALYSIS.......................................................... 81 3.9.1 TRANSCRIPTION FACTOR BINDING SITE PREDICTION ............................................ 81 3.9.2 CHROMATIN IMMUNOPRECIPITATION A SSAY...................................................81 3.10 STATISTICS................................................................................................................ 83 4. R ESU LTS................................................................................................................ 84 4.1 PRESENCE OF HYPOXIA AND HIF1A IN ATHEROSCLEROTIC M ICE .............................. 84 4.2 GENERATION OF MICE DEFICIENT FOR HIF1A IN CD11C+ CELLS .............................. 86 4.3 ANALYSIS OF ATHEROSCLEROSIS PROGRESSION IN MICE WITH DCS DEFICIENT FOR HIF1A AND FED A HIGH FAT DIET................................................................................ 88 4.4 ANALYSIS OF IMMUNE CELL PHENOTYPE IN MICE WITH DCS DEFICIENT FOR HIF1A AND FED A HIGH FAT DIET...................................................................... 92 4.5 ANALYSIS OF IMMUNE CELL PHENOTYPE IN MICE WITH DCS DEFICIENT FOR HIF1A AT THE BASELINE LEVEL......................................................................................................97 4.6 CHARACTERIZATION OF MATURE BMDCS DEFICIENT FOR H IF 1 A ................................ 99 4.7 CHARACTERIZATION OF SPLENIC DCS ISOLATED FROM ATHEROSCLEROTIC W T LDLRA AND CKO LDLF - MICE...................................................................................................... 104 4.8 EXPRESSION OF STAT PROTEINS IN DCS DEFICIENT IN H IF 1 A ............................. 108 4.9 A HIF1 A-STATS-IL-12-IFNY PATHWAY IN ATHEROSCLEROSIS ............................. 110 4.10 DETERMINATION OF HIF1A BINDING SITES ON GENE PROMOTERS AND EVIDENCE OF DIRECT TRANSCRIPTION FACTOR ACTIVITY BY CHIP......................................................... 113 5. D IS C U S S IO N ..........................................................................................................116 6. R E FE R E N C E S ....................................................................................................... 131 ACKNOW LEDG EM ENTS ......................................................................................... 156 CURRICULUM V IT A E ............................................................................................... 158 LIST OF PUBLICATIONS ......................................................................................... 159 LIST OF ORAL AND POSTER PRESENTATIO NS ............................................ 160 A FFID A V IT...................................................................................................................161
any_adam_object	1
author	Chaudhari, Sweena M. 1985-
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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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owner	DE-384 DE-473 DE-BY-UBG DE-703 DE-1051 DE-824 DE-29 DE-12 DE-91 DE-BY-TUM DE-19 DE-BY-UBM DE-1049 DE-92 DE-739 DE-898 DE-BY-UBR DE-355 DE-BY-UBR DE-706 DE-20 DE-1102 DE-860 DE-2174
owner_facet	DE-384 DE-473 DE-BY-UBG DE-703 DE-1051 DE-824 DE-29 DE-12 DE-91 DE-BY-TUM DE-19 DE-BY-UBM DE-1049 DE-92 DE-739 DE-898 DE-BY-UBR DE-355 DE-BY-UBR DE-706 DE-20 DE-1102 DE-860 DE-2174
physical	162 Blätter Illustrationen, Diagramme
psigel	ebook
publishDate	2013
publishDateSearch	2013
publishDateSort	2013
record_format	marc
spelling	Chaudhari, Sweena M. 1985- Verfasser (DE-588)1127356283 aut Role of Hypoxia-Inducible Factor (HIF) 1α in Dendritic Cells in Immune Regulation of Atherosclerosis = Rolle von Hypoxie induziertem Faktor (HIF) 1α in dendritischen Zellen in der Immunregulation der Atherosklerose submitted by Sweena M. Chaudhari from Mumbai, India ; Primary supervisor: Prof. Dr. Alma Zernecke ... Rolle von Hypoxie induziertem Faktor (HIF) 1α in dendritischen Zellen in der Immunregulation der Atherosklerose Würzburg 2013 162 Blätter Illustrationen, Diagramme txt rdacontent n rdamedia nc rdacarrier veröffentlicht: 2017 Dissertation Julius-Maximilians-Universität Würzburg 2014 Langzeitarchivierung gewährleistet, LZA Zusammenfassung in deutscher und englischer Sprache Hypoxie (DE-588)4133549-1 gnd rswk-swf Entzündung (DE-588)4014975-4 gnd rswk-swf Arteriosklerose (DE-588)4068866-5 gnd rswk-swf Dendritische Zelle (DE-588)4313846-9 gnd rswk-swf Hypoxia Atherosclerosis Dendritic cells (DE-588)4113937-9 Hochschulschrift gnd-content Arteriosklerose (DE-588)4068866-5 s Dendritische Zelle (DE-588)4313846-9 s Entzündung (DE-588)4014975-4 s Hypoxie (DE-588)4133549-1 s DE-604 Zernecke-Madsen, Alma 1976- (DE-588)129581607 dgs Erscheint auch als Online-Ausgabe, PDF urn:nbn:de:bvb:20-opus-91853 https://nbn-resolving.org/urn:nbn:de:bvb:20-opus-91853 Resolving-System kostenfrei Volltext http://d-nb.info/1125372370/34 Langzeitarchivierung Nationalbibliothek DNB Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=029625650&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis
spellingShingle	Chaudhari, Sweena M. 1985- Role of Hypoxia-Inducible Factor (HIF) 1α in Dendritic Cells in Immune Regulation of Atherosclerosis = Rolle von Hypoxie induziertem Faktor (HIF) 1α in dendritischen Zellen in der Immunregulation der Atherosklerose Hypoxie (DE-588)4133549-1 gnd Entzündung (DE-588)4014975-4 gnd Arteriosklerose (DE-588)4068866-5 gnd Dendritische Zelle (DE-588)4313846-9 gnd
subject_GND	(DE-588)4133549-1 (DE-588)4014975-4 (DE-588)4068866-5 (DE-588)4313846-9 (DE-588)4113937-9
title	Role of Hypoxia-Inducible Factor (HIF) 1α in Dendritic Cells in Immune Regulation of Atherosclerosis = Rolle von Hypoxie induziertem Faktor (HIF) 1α in dendritischen Zellen in der Immunregulation der Atherosklerose
title_alt	Rolle von Hypoxie induziertem Faktor (HIF) 1α in dendritischen Zellen in der Immunregulation der Atherosklerose
title_auth	Role of Hypoxia-Inducible Factor (HIF) 1α in Dendritic Cells in Immune Regulation of Atherosclerosis = Rolle von Hypoxie induziertem Faktor (HIF) 1α in dendritischen Zellen in der Immunregulation der Atherosklerose
title_exact_search	Role of Hypoxia-Inducible Factor (HIF) 1α in Dendritic Cells in Immune Regulation of Atherosclerosis = Rolle von Hypoxie induziertem Faktor (HIF) 1α in dendritischen Zellen in der Immunregulation der Atherosklerose
title_full	Role of Hypoxia-Inducible Factor (HIF) 1α in Dendritic Cells in Immune Regulation of Atherosclerosis = Rolle von Hypoxie induziertem Faktor (HIF) 1α in dendritischen Zellen in der Immunregulation der Atherosklerose submitted by Sweena M. Chaudhari from Mumbai, India ; Primary supervisor: Prof. Dr. Alma Zernecke ...
title_fullStr	Role of Hypoxia-Inducible Factor (HIF) 1α in Dendritic Cells in Immune Regulation of Atherosclerosis = Rolle von Hypoxie induziertem Faktor (HIF) 1α in dendritischen Zellen in der Immunregulation der Atherosklerose submitted by Sweena M. Chaudhari from Mumbai, India ; Primary supervisor: Prof. Dr. Alma Zernecke ...
title_full_unstemmed	Role of Hypoxia-Inducible Factor (HIF) 1α in Dendritic Cells in Immune Regulation of Atherosclerosis = Rolle von Hypoxie induziertem Faktor (HIF) 1α in dendritischen Zellen in der Immunregulation der Atherosklerose submitted by Sweena M. Chaudhari from Mumbai, India ; Primary supervisor: Prof. Dr. Alma Zernecke ...
title_short	Role of Hypoxia-Inducible Factor (HIF) 1α in Dendritic Cells in Immune Regulation of Atherosclerosis
title_sort	role of hypoxia inducible factor hif 1α in dendritic cells in immune regulation of atherosclerosis rolle von hypoxie induziertem faktor hif 1α in dendritischen zellen in der immunregulation der atherosklerose
title_sub	= Rolle von Hypoxie induziertem Faktor (HIF) 1α in dendritischen Zellen in der Immunregulation der Atherosklerose
topic	Hypoxie (DE-588)4133549-1 gnd Entzündung (DE-588)4014975-4 gnd Arteriosklerose (DE-588)4068866-5 gnd Dendritische Zelle (DE-588)4313846-9 gnd
topic_facet	Hypoxie Entzündung Arteriosklerose Dendritische Zelle Hochschulschrift
url	https://nbn-resolving.org/urn:nbn:de:bvb:20-opus-91853 http://d-nb.info/1125372370/34 http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=029625650&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA
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