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Pharmacological Nephroprotection in a Novel Mouse Model of Hereditary Nephrotic Syndrome
Ivana Simic, Mansoureh Tabatabaeifar, Helga Denc, Tanja Wlodkowski, Geraldine Mollet, Corinne Antignac, Franz Schaefer
Division of Pediatric Nephrology, University of Heidelberg Department of Human Genetics, Hopital Necker, Paris
25th European Congress of Pathology, Lisbon, 31 August – 4 September 2013
The NPHS2 Gene and Hereditary Nephrotic Syndrome • Mutations in the NPHS2 gene, encoding podocin, cause autosomal recessive steroid-
resistant nephrotic syndrome• R138Q, the most common podocin mutation in Europeans, causes early disease onset
and rapid progression to end-stage renal disease
• Recently, we generated and characterized an inducible knock-in mouse carrying the R140Q podocin mutation, the murine analogue of the most common human mutation R138Q
N-term
C-term
R138Q
Creation of a Conditional Knock-in Mouse Model of R140Q Mutation
Cre
Cre
Cre
Cre
Nphs2R140Q/+ Nphs2lox2/lox2
Cre+/+
Nphs2lox2/R140Q
Cre+
Nphs2R140Q/-
X
Tamoxifen induction
Bl6 Bl6
Prophylactic RAS Blockade in a Conditional Knock-in Mouse Model of R140Q Mutation
Tamoxifen-induced mice received 10 mg/kg/day of:
•ACE inhibitor Ramipril (R)
•AT1 receptor blocker Candesartan (C)
•the combination of Ramipril and Candesartan (R+C)
•non-RAS antihypertensive amlodipine (A)
Control groups:
•Tamoxifen induction (sick controls) •Vehicle injections (healthy controls)
Blood Pressure Effect of RAS Antagonists vs. Amlodipine
0
20
40
60
80
100
120
1 2 3 4Week of observation
MA
BP
(m
mH
g)
healthy untreated Ramipril Candesartan C+R Amlodipine
***
***
*** ******
***
*** ***
***
***
****
***
* p<0.05; ** p<0.01; *** p<0.001
Markedly Attenuated Proteinuria in Podocin R140Q Knock-In Mice Treated with RAS Antagonists
Normoalbuminemia in Podocin R140Q Knock-In Mice Treated with RAS Antagonists
* p<0.05; ** p<0.01; *** p<0.0001
Reduced Plasma Creatinine Levels in All Treated Animals
Subtotal Loss of Podocin Protein in All Induced Animals Irrespective of Pharmacological Treatment
Podocin
GAPDH
37
37
kDa 4 WKS Control R C R+C A
The Loss of Podocin Signal at Immunofluorescence Analysis
healthy untreated
R+C Amlodipine
Preserved Podocin mRNA Expression in All Induced Animals
0
500
1000
1500
2000
2500
3000
healthy sick Ramipril Candesartan R+C Amlodipine
Type of treatment
Po
do
cin
exp
ress
ion
(%
18S
)
*
* p>0.05 comparing to both sick untreated and healthy animals
Attenuated Glomerulosclerosis in Animals Treated with RAS Antagonists
0,00
0,20
0,40
0,60
0,80
1,00
1,20
1,40
1,60
1,80
healthy untreated Ramipril Candesartan R+C Amlodipine
Type of treatment
Glo
mer
ula
r sc
lero
sis
ind
ex
****** ***
*
* p>0.05; ** p<0.05; *** p<0.01; **** p<0.001
A.
B.
PAS staining of renal tissue in induced animals: A. Untreated (notable GS); B. Mice treated with R+C.
Insignificantly Attenuated Tubulointerstitial Fibrosis in Animals Treated with RAS Antagonists
0,00
1,00
2,00
3,00
4,00
5,00
6,00
7,00
8,00
9,00
10,00
healthy untreated Ramipril Candesartan R+C Amlodipine
Type of treatment
Are
a af
fect
ed b
y fi
bro
sis
(% o
f to
tal
kid
ney
are
a)
p>0.05
Sirius-Red-staining: A. Untreated animals; B. Mice treated with R+C.
A.
B.
Improved Podocyte Survival in Animals Treated with RAS Antagonists
0
20
40
60
80
100
120
healthy untreated Ramipril Candesartan R+C Amlodipine
Type of treatment
Po
do
cyte
nu
mb
er (
per
glo
mer
ulu
s)
***
**
***
* p<0.05; ** p<0.01; *** p<0.001
A.
B.
WT1 immunostaining of glomeruli of induced mice: A. Untreated (reduced podocyte number); B. Treated with R+C.
Summary
• RAS antagonists markedly attenuate proteinuria in mice hemizygous for podocin R140Q mutation
• After 4 weeks, mice receiving R+C were normo-albuminemic and serum creatinine was increased less than in untreated or animals treated with Amlodipine
• Reduced glomerulosclerosis and better podocyte survival in animals treated with RAS antagonists
• RAS blockade provides effective pharmacological nephroprotection in this hereditary podocytopathy model
Dr. Ivana Simic
Dr. Mansoureh
Tabatabaeifar
Tanja Wlodkowski
Dr. Helga Denc
Prof. Dr. Franz Schaefer
Hopital Necker, France
Dr. Geraldine Mollet
Prof. Dr. Corinne Antignac