Tissue-specific differentiation signals in endothelial cells:
Gespeichert in:
| 1. Verfasser: | |
|---|---|
| Format: | Abschlussarbeit Buch |
| Sprache: | English |
| Veröffentlicht: |
2003
|
| Schlagworte: | |
| Online-Zugang: | Inhaltsverzeichnis |
| Beschreibung: | 139 S. Ill., graph. Darst. |
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| 100 | 1 | |a Krüger, Bettina |e Verfasser |4 aut | |
| 245 | 1 | 0 | |a Tissue-specific differentiation signals in endothelial cells |c von Bettina Krüger |
| 264 | 1 | |c 2003 | |
| 300 | |a 139 S. |b Ill., graph. Darst. | ||
| 336 | |b txt |2 rdacontent | ||
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| 338 | |b nc |2 rdacarrier | ||
| 502 | |a Erlangen-Nürnberg, Univ., Diss., 2003 | ||
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4
-------------------------------------------------------------------------------------------
TABLE
OF
CONTENTS
TABLE
OF
CONTENTS
TABLE
OF
CONTENTS
.
4
1.
ZUSAMMENFASSUNG
.
7
1.
SUMMARY
.
11
2.
INTRODUCTION
.
14
2.1
FORMATION
OF
BLOOD
VESSELS
AND
DIFFERENTIATION
OF
THE
ENDOTHELIUM.
.
14
2.2
CELLULAR
COMPOSITION
OF
THE
GLOMERULUS
.
17
2.3
ROLE
OF
EXTRACELLULAR
MATRIX
IN
THE
GLOMERULUS
.
18
2.4
CYTOKINES
.
19
2.5
MECA-32,
A
MARKER
FOR
DIFFERENTIATED
ENDOTHELIUM
IN
THE
GLOMERULUS
.
20
2.7
CAVEOLAE
.
22
2.8
CALCIUM
SIGNALLING
FROM
CAVEOLAE
.
25
3.
HYPOTHESIS
AND
CONCEPTUAL
FORMULATION
.
28
4.
RESULTS
.
30
4.1
INFLUENCE
OF
CYTOKINES
ON
THE
REGULATION
OF
MECA-32
IN
SEND.L,
MLEND.L
AND
GLEND.2
.
30
4.1
1
STIMULATORY
EFFECT
OF
FCS
IN
THE
CELL
CULTURE
MEDIUM
.
30
4.1
2
VEGF
.
32
4.1.3
TGF-GL
.
33
4.1.4
CYTOKINE
STIMULATED
PROLIFERATION
ASSAYS
.
34
4.1.5
OTHER
CYTOKINES
.
35
4.1.
6TWO
CYTOKINES
IN
INTERACTION
.
36
4.2
INFLUENCE
OF
MATRIX
COMPONENTS
.
37
4.3
CHARACTERIZATION
OF
THE
THREE
GLOMERULAR
CELL
TYPES:
ENDOTHELIAL
CELLS,
MESANGIAL
CELLS
AND
PODOCYTES
IN
VITRO
.
38
4.3.1
THE
GLOMERULAR
ENDOTHELIAL
CELL
LINE
GLEND.2
.
38
4.3.1.1
INDUCTION
OF
E-SELECTIN
AFTER
IL-1
P
ACTIVATION
IN
GLEND.2
.
40
4.3.2
PRIMARY
MOUSE
MESANGIAL
CELLS
.
41
4.3.3
CONDITIONALLY
IMMORTALIZED
PODOCYTES
.
42
4.4
INTERACTION
OF
THE
THREE
GLOMERULAR
CELL
TYPES:
ENDOTHELIAL
CELLS,
MESANGIAL
CELLS
AND
PODOCYTES
IN
COCULTURE
EXPERIMENTS
.
44
4.4.1
THE
ROLE
OF
MESANGIAL
CELL
CULTURE
SUPERNATANT
ON
THE
MECA-32
REGULATION
IN
SEND.L
AND
MLEND.L
.
44
4.4.2
COCULTURE
EXPERIMENTS
OF
ENDOTHELIAL
AND
MESANGIAL
CELLS
.
45
4.4.3
COCULTURE
EXPERIMENTS
OF
ENDOTHELIAL
AND
PODOCYTES
.
.49
4.5
CAVEOLAE
ARE
COMPONENTS
OF
THE
PLASMA
MEMBRANE
IN
GLEND.2
AND
RAT2
FIBROBLAST
.
51
4.6
OVEREXPRESSION
OF
MECA-32
IN
GLEND.2
.
52
4.7
ISOLATION
OF
CAVEOLAR
MEMBRANES
FROM
GLEND.2
AND
GLEND/MECA-32
.
54
4.8
IDENTIFICATION
OF
MECA-32
IN
CAVEOLAE
VESICLES
AND
INVAGINATIONS
OF
GLEND.2
AND
GLEND/MECA-32
.
57
4.9
MECA-32
DISTRIBUTION
IN
CAVEOLAE
AFTER
COCULTURE
EXPERIMENTS
.
58
4.10
RNAI
IN
GLEND.2
.
60
4.11
COMPARISON
OF
GLEND.2,
GLEND/MECA-32
AND
GLEND.2
SILENCING
MECA-32
.
65
4.11.1
IMMUNOGOLD
ELECTRON
MICROSCOPY
OF
GLEND.2
SILENCING
MECA-32
.
65
4.11.2
COLOCALIZATION
OF
MECA-32
WITH
CAVEOLIN-1
.
66
4.12
THE
ROLE
OF
MECA-32
IN
A
SUBCLASS
OF
CAVEOLAE
.
67
4.12.1
RELOCATION
OF
CAVEOLAE
TO
THE
TRAILING
EDGE
.
68
4.12.2
CALCIUM
WAVE
INITIATION
IN
MIGRATING
GLEND
2
.
69
4.12.3
INITIATION
OF
CALCIUM
WAVES
IN
MIGRATING
GLEND.2
UNDER
CA
2
*
FREE
CULTURE
CONDITIONS
.
74
4.
1
2.4
CALCIUM
WAVE
INITIATION
IN
MIGRATING
GLEND.2
AFTER
DEPLETION
OF
INTRACELLULAR
CALCIUM
STORES
.
78
5.
DISCUSSION
.
81
5.1
INFLUENCE
OF
KNOWN
CYTOKINES
ON
THE
MECA-32
EXPRESSION
IN
VITRO
.
82
TABLE
OF
CONTENTS
5
5.2
COCULTURE
EXPERIMENTS
WITH
THE
THREE
GLOMERULAR
CELL
TYPES:
ENDOTHELIAL
CELLS,
MESANGIAL
CELLS
AND
PODOCYTES
.
83
5.3
LOCALIZATION
OF
MECA-32
IN
CAVEOLAE
.
86
5.4
GLEND.2
SILENCING
MECA-32
.
90
5.5
CALCIUM
SIGNALLING
IN
GLEND.2
.
91
5.5
FUTURE
EXPERIMENTS
.
98
6.
MATERIALS
AND
METHODS
.
100
6.1
MATERIALS
.
100
6.1.1
CHEMICAL
REAGENTS
.
100
6.1.2
GENERAL
BUFFERS
AND
SOLUTIONS
.
100
6.13
MOLECULAR
BIOLOGY
KITS
.
100
6.1.4
BACTERIAL
STRAIN
.
100
6.1.5
VECTORS
.
101
6.1.6
MOLECULAR
WEIGHT
AND
LENGTH
STANDARDS
AND
RESTRICTION
ENZYMES
.
101
6.1.7
BACTERIAL
MEDIA
.
101
6.1.8
CELL
LINES
AND
PRIMARY
CELLS
.
101
6.1.9
MEDIA
AND
SOLUTIONS
FOR
EUKARYOTIC
CELL
CULTURE
.
101
6.1.10
CYTOKINES
.
102
6.1.11
TRANSFECTION
REAGENTS
.
102
6.1.12
ANTIBODIES
.
103
6.1
12.1
CONJUGATES
.
103
6.1.13
OLIGONUCLEOTIDES
(PRIMER)
FOR
PCR
.
103
6.1.14
SIRNAS
.
104
6.1.15
SOFTWARE
.
104
6.2
METHODS
.
105
6.2.1
METHODS
IN
MOLECULAR
BIOLOGY
.
105
6.2.1.1
ISOLATION
OF
TOTAL
RNA
FROM
CELL
LINES
.
105
6.2.1.2
AGAROSE
GEL
ELECTROPHORESIS
.
105
6.2.1.3
FIRST
STRAND
CDNA
SYNTHESIS
.
106
6.2.1.4
POLYMERASE
CHAIN
REACTION
(PCR)
.
106
6.2.1.5
ELUTION
OF
DNA
.
106
6.2.1.6
DIGESTION
OF
DNA
WITH
RESTRICTION
ENZYMES
.
107
6.2.1.7
ISOLATION
OF
PLASMID
DNA
.
107
6.2.1.7.1
MINI-PREPARATION
OF
PLASMID
DNA
.
107
6.2.1.7.2
MIDI-PREPARATION
OF
PLASMID
DNA
.
107
6.2.1.8
NORTHERN
BLOT
ANALYSIS
.
108
6.2.2
BIOCHEMICAL METHODS
.
109
6.2.2.1
PREPARATION
OF
TOTAL
PROTEIN
FROM
CELL
LYSATES
.
109
6.2.2.2
BCA
PROTEIN
ASSAY
.
110
6.2.23
SDS-PAGE
.
110
6.2.2.4
WESTERN
BLOT
TRANSFER
.
111
6.2.2.5
IMMUNOPRECIPITATION
.
112
6.2.2.6
ENZYME
LINKED
IMMUNOSORBENT
ASSAY
(ELISA)
.
112
6.2.2.7
IMMUNHISTOCHEMISTRY
.
113
6.2.2.8.
ISOLATION
OF
CAVEOLAR
MEMBRANES
.
113
6.2.2.8.1
ISOLATION
OF
CAVEOLAR
MEMBRANES
BY
SUCROSE
GRADIENT
CENTRIFUGATION
.
113
6.2.2.8.2
DETERGENT-FREE
ISOLATION
OF
CAVEOLAR
MEMBRANES
.
114
6.2.2.9
INTRACELLULAR
CALCIUM
IMAGING
.
115
6.2.2.10
IMMUNOGOLD
ELECTRONMICROSCOPY
.
115
6.2.3 CELL
BIOLOGICAL
METHODS
.
116
6.23.1
CULTIVATION
OF
CELLS
.
116
6.23.2
TRYPSINIZATION
OF
CELLS
.
117
6.233
CELL
TRACKER
.
117
6.23.4
RNA
INTERFERENCE
.
117
6.23.4.1
SIRNAS
.
118
6.23.4.2
TRANSFECTION
OF
SIRNAS
WITH
TRANSMESSENGER
TRANSFECTION
REAGENTYY
(QIAGEN)
.
118
6.23.43
TRANSFECTION
OF
SIRNAS
WITH
METAFECTENETM
(BIONTEX)
.
119
6.23.4.4
TRANSFECTION
OF
SIRNAS
WITH
OLIGOFECTAMINETM
(INVITROGEN)
.
119
6.23.4.5
ELECTROPORATION
BY
AMAXA
NUCLEOFECTORTM
.
119
6.23.5
FLOW
CYTOMETRY
.
120
6
_
TABLE
OF
CONTENTS
6.2.3.7
TRANSFECTION
OF
CELL
LINES
.
120
6.2.3.8
RETROVIRAL
GENE
TRANSFER
.
120
6
2.3.9
COCULTURE
EXPERIMENTS
.
121
6.2.3.10
PROLIFERATION
ASSAY
.
121
7.
BIBLIOGRAPHY
.
123
8.
ABBREVIATIONS
.
129
9.
APPENDIX
.
131
A)
CDNA
SEQUENCE
MECA-32
.
131
B)
PROTEIN
SEQUENCE
MECA-32
.
131
C)
INTRACELLULAR
CA
2+
WAVES
AFTER
DEPLETION
OF
INTRACELLULAR
CA
2+
STORES
WITH
THAPSIGARGIN
.
132
DANKSAGUNGEN
.
140
LEBENSLAUF
.
141 |
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| author | Krüger, Bettina |
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| author_sort | Krüger, Bettina |
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| genre_facet | Hochschulschrift |
| id | DE-604.BV017499501 |
| illustrated | Illustrated |
| indexdate | 2024-08-19T00:07:10Z |
| institution | BVB |
| language | English |
| oai_aleph_id | oai:aleph.bib-bvb.de:BVB01-010543339 |
| oclc_num | 249496614 |
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| owner_facet | DE-29 DE-29T DE-12 DE-355 DE-BY-UBR DE-83 DE-188 |
| physical | 139 S. Ill., graph. Darst. |
| publishDate | 2003 |
| publishDateSearch | 2003 |
| publishDateSort | 2003 |
| record_format | marc |
| spelling | Krüger, Bettina Verfasser aut Tissue-specific differentiation signals in endothelial cells von Bettina Krüger 2003 139 S. Ill., graph. Darst. txt rdacontent n rdamedia nc rdacarrier Erlangen-Nürnberg, Univ., Diss., 2003 Zelldifferenzierung (DE-588)4067536-1 gnd rswk-swf Endothelzelle (DE-588)4152192-4 gnd rswk-swf Glomerulus (DE-588)4157648-2 gnd rswk-swf Embryonalentwicklung (DE-588)4070792-1 gnd rswk-swf Signaltransduktion (DE-588)4318717-1 gnd rswk-swf (DE-588)4113937-9 Hochschulschrift gnd-content Glomerulus (DE-588)4157648-2 s Endothelzelle (DE-588)4152192-4 s Zelldifferenzierung (DE-588)4067536-1 s Signaltransduktion (DE-588)4318717-1 s DE-604 Embryonalentwicklung (DE-588)4070792-1 s DE-188 DNB Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=010543339&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Krüger, Bettina Tissue-specific differentiation signals in endothelial cells Zelldifferenzierung (DE-588)4067536-1 gnd Endothelzelle (DE-588)4152192-4 gnd Glomerulus (DE-588)4157648-2 gnd Embryonalentwicklung (DE-588)4070792-1 gnd Signaltransduktion (DE-588)4318717-1 gnd |
| subject_GND | (DE-588)4067536-1 (DE-588)4152192-4 (DE-588)4157648-2 (DE-588)4070792-1 (DE-588)4318717-1 (DE-588)4113937-9 |
| title | Tissue-specific differentiation signals in endothelial cells |
| title_auth | Tissue-specific differentiation signals in endothelial cells |
| title_exact_search | Tissue-specific differentiation signals in endothelial cells |
| title_full | Tissue-specific differentiation signals in endothelial cells von Bettina Krüger |
| title_fullStr | Tissue-specific differentiation signals in endothelial cells von Bettina Krüger |
| title_full_unstemmed | Tissue-specific differentiation signals in endothelial cells von Bettina Krüger |
| title_short | Tissue-specific differentiation signals in endothelial cells |
| title_sort | tissue specific differentiation signals in endothelial cells |
| topic | Zelldifferenzierung (DE-588)4067536-1 gnd Endothelzelle (DE-588)4152192-4 gnd Glomerulus (DE-588)4157648-2 gnd Embryonalentwicklung (DE-588)4070792-1 gnd Signaltransduktion (DE-588)4318717-1 gnd |
| topic_facet | Zelldifferenzierung Endothelzelle Glomerulus Embryonalentwicklung Signaltransduktion Hochschulschrift |
| url | http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=010543339&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |
| work_keys_str_mv | AT krugerbettina tissuespecificdifferentiationsignalsinendothelialcells |