Abstract: SA-PO311
Differences in Podocyte Regeneration Between Cortical and Juxtamedullary Nephrons Defined by 3D Analysis and STED Imaging
Session Information
- Cellular Crosstalk in Glomerular Diseases - II
October 27, 2018 | Location: Exhibit Hall, San Diego Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Glomerular Diseases
- 1201 Glomerular Diseases: Fibrosis and Extracellular Matrix
Authors
- Angelotti, Maria Lucia, Excellence Center for Research, Transfer and High Education Denothe, University of Florence, Florence, Italy
- Antonelli, Giulia, University of Florence, Florence, Italy
- Conte, Carolina, University of Florence, Florence, Italy
- Melica, Maria elena, Università di Firenze, Florence, Italy
- Anders, Hans J., Klinikum der Universitat Munchen, Munchen, Germany
- Romagnani, Paola, University of Florence, Florence, Italy
Background
Podocyte injury is a central pathomechanism of glomerulosclerosis. Regeneration of lost podocytes from podocyte progenitors along Bowman`s capsule can occur but the number of new podocytes seemed to be rather low. We hypothesized that traditional 2D tissue analysis underestimates podocyte regeneration and that injury and regeneration are not equally distributed between cortical and juxtamedullary nephrons.
Methods
To test this concept we employed Nphs2.rtTA;TetO.Cre;mT/mG and Pax2.rtTA;TetO.Cre;mT/mG mice to quantify adriamycin (ADR)-related podocyte loss and de novo podocyte formation from Pax2+ podocyte progenitors by lineage tracing, using Z-stack confocal microscopy for 3D quantitative analysis and STED microscopy to assure integration of new podocytes into the glomerular filtration barrier. CXCL12 inhibition was used to assess drug-related enhancement of podocyte regeneration.
Results
2D analysis underestimated the percentage of glomeruli showing podocyte regeneration in comparison to 3D (8.05% vs 25.23% p<0.05). Doxycyclin-induced podocyte labeling in Nphs2.iCreER;mT/mG mice revealed a higher number of podocytes in juxtamedullary glomeruli vs. cortical glomeruli (52.56±1.5 vs 81.96±2.85, p<0.001). In contrast, doxycyclin-induced Pax2+ progenitor labeling in Pax2.rtTA;TetO.Cre;mT/mG mice revealed that less juxtamedullary glomeruli had progenitors along Bowman`s capsule vs. cortical glomeruli (79.03±0.15% vs 46.15±2.2%, p<0.05). Upon ADR exposure, Nphs2.iCreER;mT/mG mice juxtamedullary glomeruli lost a higher percentage of podocytes as compared to cortical glomeruli (13.79±3.18% vs 34.11±1.65%, p<0.05). Pax2.rtTA;TetO.Cre;mT/mG mice revealed that podocyte regeneration was limited in juxtamedullary glomeruli, while cortical glomeruli revealed new progenitor-derived podocytes, a difference that increased with CXCL12 inhibition (24.03±4.8 vs 75.96±4.8, p<0.05). STED microscopy revealed the ultrastructure of new podocytes, including secondary and tertiary foot processes. CXCL12 blockade also resulted in proteinuria improvement (p<0.05 at 28 days).
Conclusion
These results show that juxtamedullary and cortical nephrons differ in their capacity to regenerate podocytes upon injury and explain the clinical observation that FSGS starts and is more severe in juxtamedullary nephrons.
Funding
- Government Support - Non-U.S.