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 knockin 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
Bl6
X
Nphs2R140Q/+
Cre
Bl6
Cre
Nphs2lox2/lox2
Cre+/+
Cre
Nphs2lox2/R140Q
Cre+
Tamoxifen induction
Nphs2R140Q/Cre
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
120
MABP (mmHg)
100
80
**
*
**
***
*** ***
**
***
***
***
*** ***
***
***
60
40
20
0
1
2
3
Week of observation
healthy
* p<0.05; ** p<0.01; *** p<0.001
untreated
Ramipril
Candesartan
4
C+R
Amlodipine
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
4 WKS
Control
R
C
R+C
A
kDa
37
37
Podocin
GAPDH
The Loss of Podocin Signal at Immunofluorescence
Analysis
healthy
untreated
R+C
Amlodipine
Preserved Podocin mRNA Expression
in All Induced Animals
Podocin expression (% 18S)
3000
*
2500
2000
1500
1000
500
0
healthy
sick
Ramipril
Candesartan
Type of treatment
* p>0.05 comparing to both sick untreated and healthy animals
R+C
Amlodipine
Attenuated Glomerulosclerosis in Animals
Treated with RAS Antagonists
A.
Glomerular sclerosis index
1,80
1,60
*
1,40
**
1,20
***
****
1,00
0,80
B.
0,60
0,40
0,20
0,00
healthy
untreated
Ramipril
Candesartan
R+C
Amlodipine
Type of treatment
* p>0.05; ** p<0.05; *** p<0.01; **** p<0.001
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
A.
Area affected by fibrosis
(% of total kidney area)
10,00
p>0.05
9,00
8,00
7,00
6,00
5,00
B.
4,00
3,00
2,00
1,00
0,00
healthy
untreated
Ramipril
Candesartan
R+C
Amlodipine
Type of treatment
Sirius-Red-staining: A. Untreated
animals; B. Mice treated with R+C.
Improved Podocyte Survival in Animals
Treated with RAS Antagonists
Podocyte number (per glomerulus)
A.
120
*
100
**
***
**
80
B.
60
40
20
0
healthy
untreated
Ramipril
Candesartan
R+C
Amlodipine
Type of treatment
* p<0.05; ** p<0.01; *** p<0.001
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