Abstract: FR-PO949
Podocyte Cell Cycle Manipulation as a Potential Tool in Treating Glomerular Disease
Session Information
- Glomerular Diseases: Podocyte Biology - II
November 08, 2019 | Location: Exhibit Hall, Walter E. Washington Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Glomerular Diseases
- 1204 Podocyte Biology
Authors
- Perin, Laura, Childrens Hospital Los Angeles, Los Angeles, California, United States
- Nicolas frank, Camille H., The Saban Research Institute, Los Angeles, California, United States
- Lemley, Kevin V., Childrens Hospital Los Angeles, Los Angeles, California, United States
- De Filippo, Roger E., Childrens Hospital Los Angeles, Los Angeles, California, United States
Background
Loss of podocytes is a hallmark of most progressive kidney disease. Podocytes do not replicate in situ; hence, hypertrophy is the only mechanism they have to compensate for cell loss by allowing a smaller number of podocytes to effectively cover the glomerular tuft. To undergo hypertrophy, podocytes re-enter the cell cycle, moving from quiescent G0 to G1. Regulation of the G1/S checkpoint is critical. If podocytes exit G1 and continue toward mitosis, they detach from the GBM during cytokinesis and are lost in the urine. Understanding cell cycle regulation may be pivotal in developing novel therapies to prevent podocyte loss.
Methods
To the study the podocyte cell cycle, we established colonies of FUCCI2aR mice. These mice have ubiquitylation-mediated fluorescent protein expression, which reflects the cell cycle in vivo: red in G1/S, green in S/G2, yellow for S phase, no fluorescence in G0. These FUCCI mice were bred with our model of glomerular injury (Alport Syndrome, AS) and with a podocyte-specific Cre-mouse, yielding a model that allows real-time studies of the cell cycle specifically in podocytes.
Results
Podocytes isolated from glomeruli of wild type FUCCI2aR mice were 23% of total glomerular cells: 93% were in G0, 6% in G1 and none in S/G2. In late-stage proteinuric AS FUCCI2aR mice, 95% of the podocytes were in G1. We found that rapamycin (an mTOR inhibitor) was protective by supporting podocyte survival in vitro. Podocytes isolated from cultured glomeruli exposed to rapamycin for 3 days had increased survival (16%-20%) compared to podocytes not exposed to rapamycin (7%-9%). Furthermore, in PAN-exposed podocytes in vitro rapamycin increased the number of podocytes in G1 [from 4% to 13.5%]. The G1 podocytes were also larger by 41.5% (p<0.05) than those in G0 as assessed by flow cytometry forward scatter, confirming that the G1 podocytes are in a hypertrophic state.
Conclusion
Interventions that support podocyte hypertrophy, while limiting progression to mitosis or cytokinesis, may stabilize glomeruli against sclerosis following a loss of podocytes. Rapamycin presents a potential novel therapy in glomerular diseases such as AS by enhancing stable hypertrophy of podocytes in G1, while preventing progression through the cell cycle to S and mitosis.
Funding
- Private Foundation Support