Abstract: TH-PO1060
Knockdown of Podocyte Nephronectin by Glomerular Endothelial Cell-Derived MicroRNA-192 Leads to Alterations in GBM
Session Information
- Glomerular Diseases: Podocyte Biology - I
November 07, 2019 | Location: Exhibit Hall, Walter E. Washington Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Glomerular Diseases
- 1204 Podocyte Biology
Authors
- Müller-Deile, Janina, University of Erlangen, Erlangen, Germany
- Sopel, Nina, Universitätsklinikum Erlangen, Erlangen, Germany
- Hiremath, Chitkale, UTSouthwestern, Dallas, Texas, United States
- Ohs, Alexandra, Universitätsklinikum Erlangen, Erlangen, Germany
- Marciano, Denise K., University of Texas Southwestern Medical Center, Dallas, Texas, United States
- Daehn, Ilse S., Mount Sinai School of Medicine, New York, New York, United States
- Schiffer, Mario, University Hospital Erlangen, Erlangen, Germany
Background
Nephronectin (NPNT) is an extracellular matrix protein downregulated by either TGF-β or miR-378a in podocytes. Knockdown of npnt leads to proteinuria, podocyte effacement and splitting of the glomerular basement membrane (GBM) in zebrafish larvae. Here we investigated the regulation von NPNT by TGF-β and miRs as well as GBM phenotype after knockdown of NPNT more in detail by using cell culture, zebrafish and mice models.
Methods
By using mouse SMAD2/3 knockout podocytes as well as selective inhibitors of the TGF-β pathway we analyzed the TGF-β-NPNT axis more in detail. We overexpressed this miR in zebrafish larvae and analyzed the glomerular phenotype. Finally podocyte specific knockdown of Npnt was investigated in Npnt -/-;Six2-cre mice.
Results
TGF-β regulation of NPNT is mediated by the canonical TGF-β pathway in a SMAD-dependent manner. We identified glomerular endothelial cell-derived miR-192 as a regulator of NPNT in podocytes. Transfection of cultured podocytes with a miR-192 mimic down regulated NPNT expression. Overexpression of miR-192 in zebrafish larvae induced edema, proteinuria and GBM thickening similar to the phenotype after morpholino induced npnt knockdown (Fig. 1A). We characterized the phenotype of npnt knockdown in more detail by using SBF-SEM that allowed a three dimensional on the zebrafish glomerular filtration barrier in zebrafish (Fig. 1B). The unique GBM pathology was further confirmed in a mouse model with specific knockout of Npnt in podocyte progenitor cells (Npnt -/-;Six2-cre mice) (Fig. 1C).
Conclusion
The results confirm the role of podocytic npnt for proper GBM function and suggests its regulation by podocyte- and glomerular endothelial cell-derived miRs.