Survival Rate

1. Helga Denc1 ,Tanja Wlodkowski1, Mansoureh Tabatabaeifar1,Ivana Simic1, Geraldine Mollet2, Corinne Antignac2, Franz Schaefer1 1 Pediatric Nephrology, Center for Pediatrics and Adolescent Medicine, University of Heidelberg, Germany 2 Inserm U983 and Dept. of Genetics, Hopital Necker, France Background NPHS2 mutations cause hereditary nephrotic syndrome and progressive renal failure. We recently generated an inducible knock-in mouse model carrying R140Q, the analogue of the most common human mutation R138Q. These mice develop focal-segmental glomerulosclerosis with nephrotic syndrome and progressive renal failure. Here we tested the efficacy of early and delayed RAS blockade in this model of hereditary podocyte disease. Model In C57BL/6 mice with Nphs2Flox/R140Q/Cre+ genotype, hemizygosity for mutant podocin was induced by tamoxifeninjection (i.p. 33 mg/kg/day). Pharmacologicaltreatment Animals received combined high-dose ACE inhibition and AT1 receptor blockade (ramipril+candesartan 10 mg/kg each, R+C) or remained untreated. Treatment was started either prophylactically (P) at time of induction or with a 4-week delay (D). Animals were either sacrificed after 5 wks (9 wks in D) or observed open-end and the glomerular filtration rate, biochemical parameters and histopathological changes were examined (glomerulosclerosis, tubulointerstitialchanges, podocyte loss).Proteinuria and blood pressure were monitored weekly. Plasma was obtained via cardiac puncture at the time of sacrificing. (***p<0.001 vs. untreated, Student´s t-Test) Proteinuria In the P animals, RAS blockade persistently inhibited proteinuria (13% of untreated animals at 4 wks). In the D group, proteinuria sharply decreased upon RAS blockade. Podocinprotein abundance was almost totally lost in both untreated and treated animals, despite preserved or even increased mRNA expression. Control Wk 4 untreated Wk 8 untreated Wk 4 prophylacticRx Wk8 delayedRx week 4 week 8 Control Wk 8 untreated Wk 4 prophylacticRx Wk 4 untreated Wk8 delayedRx control control untreated delayedRx untreated prophylacticRx p>0.05 Nephroprotective Efficacy of RAS Blockade in Mice Carrying R140Q Podocin Mutation Week 4 prophylacticRx Week 8 delayedRx untreatedweek 4 Control Podocin protein & gene expression Number of podocytes per glomerulus Methods Survival Rate WT1 immunostainingofglomerularcross-sections (4µm & 10 µm). Numberofpodocytes per glomerulus was preserved in P animals (***p<0.001).Quantificationwas donewithImageJaccording to Animal Models of Diabetic Complications Consortium protocol. Glomerular sclerosis Results RAS Blockade Preserves / Restitutes Normoalbuminemia Sclerosis within the glomerular tuft examined by PAS staining of 3µm paraffin sections (glomerular sclerosis index – GSI, 50 glomeruli/animal. *p<0.05 **p<0.01) Tubulointerstitial fibrosis prophylacticRx Three-month survival was 31% in the untreated, 67% in the D and 100% in the P group respectively (p<0.05) *** *** Tubulointerstitialfibrosisdeterminedby Sirus Redstaining. A slight, non-significantreductionwas noted in treatedanimals. Quantification was donewith Image ProPremierSoftware. delayedRx untreated Conclusion In micecarryingthemostcommon human podocinmutation, RAS blockadeattenatuesproteinuria, podocyteloss andglomerulosclerosisdespitepersistentlyincreaseddegradationofmutantpodocinprotein. Renal failureisdelayedandsurvivalprolonged. Treatment ismoreeffectivewhenappliedprophylacticallythanwhenstarted in establishednephropathy. Ourfindingsindicatethatearly RAS blockademayprovideeffectivenephroprotection in thishereditarypodocytopathy. The research leading to these results has received funding from the European Community’s Seventh Framework Programme under grant agreement n° 2012-305608 “European Consortium for High-Throughput Research in Rare Kidney Diseases (EURenOmics)”