Role of the autonomic renal innervation in experimental glomerulonephritis [Elektronische Ressource] = Bedeutung der autonomen renalen Innervation für Entzündungsvorgänge bei experimentellen Nephritiden / vorgelegt von Eva-Maria Vogel

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Role of the autonomic renal innervation in experimental glomerulonephritis Bedeutung der autonomen renalen Innervation für Entzündungsvorgänge bei experimentellen Nephritiden Der Naturwissenschaftlichen Fakultät der Friedrich-Alexander-Universität Erlangen-Nürnberg zur Erlangung des Doktorgrades vorgelegt von Eva-Maria Vogel aus Fürth Als Dissertation genehmigt von der Naturwissenschaftlichen Fakultät der Universität Erlangen-Nürnberg Tag der mündlichen Prüfung: 29.09.2008 Vorsitzender der Prüfungskommission: Prof. Dr. E. Bänsch Erstberichterstatter: Prof. Dr. G. Lee Zweitberichterstatter: R. Veelken CONTENTS Publication List.......................................................................................1 Abbreviations .........................................................................................2 1 Introduction ......................................................................................4 1.1 The Kidney.......................................................................................................4 1.2 Mesangioproliferative Glomerulonephritis........................................................5 1.3 Animal model of mesangioproliferative glomerulonephritis (the anti-Thy-1.1 model).................................................................................................
Publié le : mardi 1 janvier 2008
Lecture(s) : 68
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Source : WWW.OPUS.UB.UNI-ERLANGEN.DE/OPUS/VOLLTEXTE/2008/1216/PDF/EVA_MARIAVOGELDISSERTATION.PDF
Nombre de pages : 97
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Role of the autonomic renal innervation in
experimental glomerulonephritis



Bedeutung der autonomen renalen Innervation für
Entzündungsvorgänge bei experimentellen Nephritiden












Der Naturwissenschaftlichen Fakultät
der Friedrich-Alexander-Universität Erlangen-Nürnberg
zur
Erlangung des Doktorgrades



vorgelegt von
Eva-Maria Vogel
aus Fürth


Als Dissertation genehmigt von der Naturwissenschaftlichen Fakultät der Universität
Erlangen-Nürnberg

































Tag der mündlichen Prüfung: 29.09.2008

Vorsitzender der Prüfungskommission: Prof. Dr. E. Bänsch
Erstberichterstatter: Prof. Dr. G. Lee
Zweitberichterstatter: R. Veelken
CONTENTS


Publication List.......................................................................................1
Abbreviations .........................................................................................2
1 Introduction ......................................................................................4
1.1 The Kidney.......................................................................................................4
1.2 Mesangioproliferative Glomerulonephritis........................................................5
1.3 Animal model of mesangioproliferative glomerulonephritis (the anti-
Thy-1.1 model).................................................................................................7
1.4 Immune cells in glomerulonephritis..................................................................9
1.5 Innervation of the kidney................................................................................10
1.6 Neuroimmunomodulation...............................................................................12
1.6.1 The neuropeptide Substance P ......................................................... 15
1.6.2 The neuropeptide Calcitonin Gene-related peptide ........................... 16
1.7 Aim of the study.............................................................................................19
2 Materials and Methods ..................................................................21
2.1 Animals..........................................................................................................21
2.2 Animal treatment............................................................................................21
2.2.1 Rat model of Anti-Thy-1.1 Nephritis................................................... 21
2.3 Renal denervation..........................................................................................21
2.3.1 Depletion of primary sensory neurons ............................................... 22
2.3.2 Administration of antagonists............................................................. 22
2.3.3 Blood pressure measurements.......................................................... 23
2.3.4 Sampling of material.......................................................................... 23 Contents ii
2.4 Mesangial cell culture ....................................................................................23
2.5 Determination of urinary albumin excretion by ELISA....................................24
2.6 Determination of urinary protein excretion .....................................................24
2.7 CGRP Immunoassay .....................................................................................24
2.8 Immunohistochemistry...................................................................................25
2.9 Kidney histology.............................................................................................26
2.10 Immunocytochemistry....................................................................................26
2.10.1 NK-1R immunoreactivity.................................................................... 26
2.10.2 CRLR immunoreactivity..................................................................... 27
2.11 Isolation of lymphocytes from kidneys ...........................................................27
2.12 Flow-cytometric analysis................................................................................28
2.13 Real Time RT-PCR........................................................................................28
2.14 Sequence analysis.........................................................................................30
2.15 Western blot analysis.....................................................................................30
2.16 Statistical analysis31
3 Results ............................................................................................32
3.1 Autonomic renal denervation ameliorate glomerulonephritis .........................32
3.2 Influence of primary sensory neurons on anti-Thy-1.1 nephritis. ...................37
3.3 CGRP Receptor blockade influenced anti-Thy-1.1 nephritis..........................40
3.3.1 CGRP Receptor expression in the kidney ......................................... 40
3.3.2 pression in anti-Thy-1.1 Nephritis........................ 43
3.3.3 Blockade of CGRP receptor .............................................................. 44
3.4 Blockade of NK-1R ameliorates glomerulonephritis.......................................48
3.4.1 Neurokinin-1 Receptor Expression in the kidney ............................... 48
3.4.2 1 receptor Expression in anti-Thy-1.1 Nephritis............... 50
3.4.3 Blockade of NK-1R ameliorates glomerulonephritis .......................... 53 Contents iii
4 Discussion ......................................................................................59
4.1 Protective role of the autonomic nervous system in glomerulonephritis ........59
4.2 Depletion of primary sensory neurons influenced renal inflammation............61
4.3 CGRP in anti-Thy-1.1 nephritis – differential effects......................................62
4.4 Blockade of NK-1R ameliorates kidney inflammation during
glomerulonephritis .........................................................................................65
4.5 Perspective....................................................................................................67
5 Summary68
6 References......................................................................................70
7 Deutschsprachige Zusammenfassung ........................................87
8 Danksagung....................................................................................90
9 Lebenslauf92


Publication List 1
Publication List

abstracts:

Veelken R, Vogel EM, Cordasic N, Hilgers KF, Amann K, Tiegs G. Tachykinin
receptor blockade ameliorates kidney inflammation during anti-Thy 1.1 nephritis in
rats. Possible role of renal afferent nerves. Annual Meeting of the ASN
Philadelphia 2005

Vogel EM, Cordasic N, Hilgers KF, Amann K, Veelken R, Tiegs G. The neurokinin
receptor-1 antagonist aprepitant ameliorates anti-Thy-1.1 nephritis in rats.
Naunyn-Schmiedberg’s Arch Pharmacol 2006; 372 (Suppl 1); Abstr. 261

Vogel EM, Cordasic N, Veelken R, Tiegs G. Neuropeptides in kidney inflammation
– the role of substance P and CGRP. International SFB 423 Symposium
“Molecular Targets in Renal Disease” 2006

Veelken R, Vogel EM, Hilgers KF, Amann K, Peters H, Tiegs G. CGRP receptor
blockade aggravates kidney inflammation during anti-Thy 1.1 nephritis in rats.
Annual Meeting of the ASN San Diego 2007

Vogel EM, Veelken R, Tiegs G. The substance P receptor antagonist aprepitant
prevents experimental glomerulonephritis DPhG Jahrestagung Erlangen 2007
(Abstr. F07)



journal:

Veelken R, Vogel EM, Hilgers K, Amann K, Hartner A, Sass G, Neuhuber W and
Tiegs G. Autonomic renal denervation ameliorates experimental
glomerulonephritis in the rat. J Am Soc Nephrol 2008: Apr 9. (Epub ahead of print)

Abbreviations 2
Abbreviations
cDNA copy DNA
CGRP calcitonin gene-related peptide
CRLR calcitonin receptor-like receptor
Cy3 indocarbocyanin
DC dendritic cell
DNA desoxyribonucleic acid
EIA enzyme-linked immunoassay
ELISA enzyme-linked immunosorbent assa
FACS fluorescence activated cell sorter
FCS fetal calf serum
FITC fluoresceinisothiocyanat
HRP horseradish peroxidaase
IgA immunoglobulin A
IgAN IgA nephropathy
IL-1 interleukin 1
IL-6 6
MAP mean arterial blood pressure
NE norepinephrine
NF-κB nuclear factor-kappa B
NKA neurokinin A
NKB B
NK-1R neurokinin-1 receptor
NPγ neuropeptide γ
NPK K Abbreviations 3
RAMP1 receptor activity modifying protein 1
RCP receptor component protein
RNA ribonucleic acid
RSNA renal sympathetic nerve activity
SP substance P
TGFβ transforming growth factor beta
TNFα tumor necrosis factor alpha
TRPV1 transient receptor potential vanilloid 1 Introduction 4
1 Introduction
1.1 The Kidney
The kidneys have to perform numerous functions in the body. They filter waste
products from the blood, are responsible for homeostasis of electrolytes and blood
volume and they play a role in acid-base balance (Rang, H.P. et al. 2003). The
functional unit of the kidney is the nephron, which consists of glomerulus, proximal
convoluted tubule, loop of Henle, distale convoluted tubule and collecting duct.
6Most of the approximately 1.3 x 10 nephrons are located in


Fig. 1-1: Structure and different cell types of the glomeruli
(http://herkules.oulu.fi/isbn9514264290/html/x782.html)

Introduction 5
the renal cortex. The main function of the kidney, the excretion of low-molecular-
weight waste from the blood, takes place at the glomeruli. A glomerulus consists of
a capillary tuft located inside the Bowman’s capsule and projects into a dilated end
of renal tubule (Tryggvason, K. et al. 2005). About 1.2 litre blood per minute
enters the glomeruli through afferent arterioles and all constitutes except large
plasma proteins pass the capillary wall. The wall is composed of three different
layers: the capillary endothelium, the basement membrane and the epithelial cell
layer of the capsule (Fig. 1-1). The filtrate, about 180 litres of primary urine, then
passes the tubulus system and approximately 99% of water as well as electrolytes
are reabsorbed and the concentrated filtrate excretes as urine. In contrast filtered
blood leaves the glomeruli through efferent arterioles (Rang, H.P. et al. 2003).
Hence, the glomeruli are predisposed for deposition of immune complexes due to
their central position in renal excretion.

1.2 Mesangioproliferative Glomerulonephritis
Glomerulonephritis, an immune-mediated disorder characterized by inflammation
within the glomerulus and other compartments in the kidney, remains a prevalent
cause for end-stage kidney failure worldwide with the necessity of dialysis and
transplantation (Chadban, S. J. et al. 2005). Here, the most common form of
mesangioproliferative glomerulonephritis is IgA nephropathy (IgAN), which is
characterized by deposition of polymeric IgA within the glomeruli as well as
mesangial hypercellularity (Fig. 1-2). The problem of IgAN is the invisibility and the
different clinical outcomes. Neither serum IgA nor other circulating factors like IgA-
immune complex are sufficient to identify this renal disease. A renal biopsy is the
only way to clarify diagnosis (Floege, J. et al. 2000). Interestingly, in patients with
IgA nephropathy the pattern of O-glycosylation of circulating IgA1 molecules
seems to differ from patients without kidney diseases and the abnormality of O-
glycosylation leads to a higher lectin binding (Allen, A. C. et al. 2001). The
interaction of mannose binding lectin (MBL) with IgA presents one possibility to
activate complement system via lectin pathway (Fig. 1-3) and in patients with IgAN
about 25% of biopsies showed MBL deposition (Roos, A. et al. 2006; Berger, S. P.
et al. 2007). As a consequence of IgA or IgA-containing immune complex
deposition in the mesangium two further possibilities exist how renal damage

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