clinicalweb.marionegri.it · Abbate et al., J Am Soc Nephrol, 2006. Sham RMR RMR+lisinopril...
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Transcript of clinicalweb.marionegri.it · Abbate et al., J Am Soc Nephrol, 2006. Sham RMR RMR+lisinopril...
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TREATMENT WITH ANTI-C6 ANTIBODY TO BLOCK GLOMERULAR C5b-9 FORMATION PREVENTS PROTEINURIA OF PHN Sprague Dawley rats given anti-C6 IgG to induce stable C6 depletion before injection of anti-Fx1A (sacrifice at day 4)
Baker et al., Am J Pathol, 1989
C5b-9 staining
Uri
nary
prot
ein
excr
etio
n(m
g/24
h)
30
20
10
0PHN PHN +
Anti-C6AbPHN PHN +
Anti-C6Ab
PODOCYTE DYSFUNCTION RESPONSIBLE FOR GLOMERULAR PERMSELECTIVE DEFECT IN MEMBRANOUS NEPHROPATHY
Downregulation of nephrin gene in severe PHN can be partly corrected by earlyinstitution (since day 7) of ACEi or AngII blocker treatment
Control PHN 8 mo
PHN 8 mo + ACEi PHN 8 mo + Ang II RA
Benigni et al., J Am Soc Nephrol, 2001
(mg/
day)
900
0Control
Proteinuria
600
300
PHN PHN +ACEi
PHN +AngII RA
COMPLEMENT MEDIATES NEPHRIN REDISTRIBUTION IN EXPERIMENTAL MEMBRANOUS NEPHROPATHY
Saran et al., Kidney Int, 2003
Proteinuria
PHN rats sacrificed at 4 daysComplement depletion with cobra venom factor after PHN induction
The onset of proteinuria is coincident with complement-dependent alterations in the interaction of nephrin with the actin cytoskeleton and loss of integrity of the podocyte slit diaphragm
Total glomerular nephrin
Control PHN C-deplete
Urin
e pr
otei
nex
cret
ion
(mg/
24h)
12
10
8
6
4
2
0
6000
5000
4000
3000
2000
0
Tot
al g
lom
erul
arne
phrin
(den
sito
met
ricun
its)
1000
Control PHN C-deplete
*
*
Control kidney
IMN
TRPC6 Dysfunctional cation permeable ion channel in foot processes causes a familialform of glomerulosclerosis and may contribute to glomerular disease pathogenesisvia dysregulated Ca influx Winn et al, Science, 2005
Moller et al., J Am Soc Nephrol, 2007
TRPC6 GENE DELIVERY IN MICE CAUSES TRPC6 OVEREXPRESSION AND PROTEINURIAi.v. injection of FLAG-TRCP6 fusion protein construct
Control TRPC6 gene delivery
Moller et al., J Am Soc Nephrol, 2007
Factor H
(C5 convertase) (C5 convertase)
CRIT CRIT mRNA
Moll et al., Kidney Int, 2006
Normalkidney
MN
UPREGULATION OF A NOVEL COMPLEMENT REGULATORY CELL SURFACE RECEPTOR IN MEMBRANOUS NEPHROPATHYCRIT, C2 receptor inhibitor trispanning
Predominance of non classical-pathway activating IgG4 in subepithelial deposits of IMN
Little C1q and C4 in deposits
A ROLE FOR ALTERNATIVE PATHWAY IN HUMANS?
Alternative pathway activation may amplify injury induced through the classical pathway
Abbate et al., J Am Soc Nephrol, 2006
Sham RMR RMR+lisinopril
C3-MHCII
LISINOPRIL TREATMENT LIMITS FILTERED PROTEIN LOAD AND RENAL DAMAGE IN PROTEIN OVERLOAD NEPHROPATHY
Pro
tein
uria
(mg/
day) 40
20
0
BSABSA + lisinopril
p < 0.05 vs BSA p < 0.01 vs BSA*
**
*
Tubu
lar
dam
age
(sco
re)
2.5
1.0
0
0.5
1.5
2.0
**Tubu
lar
stai
ning
(sco
re) 2.5
1.0
0
0.5
1.5
2.0
C3 IgG 60
20
0
40
F4/8
0+ce
lls(n
umbe
r/HP
F)
**** **
WT + saline C3-/- + BSAWT + BSA
F4/80
C3-DEPENDENT INTERSTITIAL INFLAMMATORY AND FIBROGENIC RESPONSE TO PROTEIN OVERLOAD OF TUBULAR CELLS
F4/8
0(c
ells
/HP
F)60
40
20
0
*
saline
WT C3-/-
BSA
WT C3-/-
° 2
1.5
1
0.5
0
αα ααSM
Ast
aini
ng(s
core
)
saline
WT C3-/-
BSA
WT C3-/-
°
*
Abbate et al, JASN, 2004
Available data support the rationale for complement inhibitory approaches to block pathways of glomerular and tubulointerstitial injury
More information is needed on the role of complement regulators in MN
C3-/- + saline
C3-/- + BSAWT + BSA
WT + saline
Apical challenge by complement occurs only in proteinuric renal disease and can be favored by the fact that apical surface of tubular epithelial cells constitutively lack complement inhibitors
Rangan et al., Kidney Int, 2005
C3 deposition
*°
°
*°
0
4,000
8,000
12,000
16,000
C3
depo
sit
area
(pix
el 2
)
alb+HS
transf+HS
control -HS
MAC deposition
0
2,000
4,000
6,000
8,000 *°
°
10,000
MA
Cde
posi
tar
ea (p
ixel
2)
alb+HS
transf+HS
control -HS
°
EFFECT OF PLASMA PROTEINS ON SERUM-INDUCED C3 AND MAC DEPOSITION
•Complement activation occured via alternative pathway since serum treatment with MgCl2-EGTA (aninhibitor of the classic pathway) had no effect on HS-induced C3 deposition in the presence of proteins.
°P<0.01 vs control, *P<0.01 vs HS
PROTEIN OVERLOAD INHIBITS FACTOR H BINDING ON HK-2 CELL SURFACE
control+ factor Hcontrol alb+ factor H transf+ factor H
Buelli et al, JASN, 2006
PROXIMAL TUBULAR EPITHELIAL CELLS EXPRESS C3a RECEPTOR AND ARE ACTIVATED BY C3a
In cultured human proximal tubular cells the C3a receptor mediates cell activation in response to C3 triggering inositoltriphosphate generation
Immunohistochemistry
Expression both in human and mouse tubuli and glomeruli, limited to epithelial cell types
In situ hybridization
Braun et al., J Immunol, 2004
RAT GLOMERULAR EPITHELIAL CELLS PRODUCE AND BEAR FACTOR H ON THEIR SURFACE THAT IS UP REGULATED UNDER COMPLEMENT ATTACK
Ren et al., Kidney Int, 2003
BEFORE RITUXIMAB AFTER RITUXIMAB
IMMUNOFLUORESCENCE STAINING FOR IgG4
COURSE OF TRANSPLANTED HEYMANN NEPHRITIS KIDNEY IN NORMAL HOSTRemission and residual proteinuria resembling the response to rituximab in humans
Makker et al., J Immunol, 1989
Active HN(12-16 w)
Nephrectomizedsyngenic recipientTransplant kidney
Pro
tein
uria
(mg/
24 h
rs)
Weeks post-transplant
0 4 8 12 16 200
70
*** *
* *
2 24 28
60
50
40
30
20
10
Normal range
***
*
< 1.7 > 1.7
Tubulo-Interstitial score
Cybulsky et al., Am J Physiol Renal, 2005
SIGNALING PATHWAYS ACTIVATED BY C5b-9 IN CULTURED GLOMERULAR EPITHELIAL CELLS
DOES A RELATIVE DYSFUNCTION OF COMPLEMENT REGULATORY PROTEINS IN PODOCYTES LEAD TO COMPLEMENT ACTIVATION AND INJURY?
Fx1A that is used to induce active Heymann nephritis contains Crry and CD59
Antibodies to Crry and CD59 of Fx1A neutralize their complement regulatory activities in glomerular epithelial cells in vitro
Neutralizing autoantibodies to Crry and CD59 render the podocyte susceptible to complement-mediated injury in vivo
CR1 protein is reduced on podocytes in human MN
Quigg et al, J Immunol 1991
Schiller et al, J Exp Med 1998Cunningham et al, KI 2001
Kazatchkine et al, J Clin Invest 1982Moll et al, KI 2001
