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Abstract: SA-OR26

Podocyte-Specific Loss of Klf4 Induces the Formation of Extracellular Matrix Extensions

Session Information

Category: Glomerular Diseases

  • 1301 Glomerular Diseases: Fibrosis and Extracellular Matrix

Authors

  • Cintron Pregosin, Nina, Stony Brook Medicine, Stony Brook, New York, United States
  • Bronstein, Robert, Stony Brook Medicine, Stony Brook, New York, United States
  • Mallipattu, Sandeep K., Stony Brook Medicine, Stony Brook, New York, United States
Background

Podocyte loss is characteristic of multiple glomerular diseases, including focal segmental glomerulosclerosis (FSGS) and rapidly progressive glomerulonephritis (RPGN). In these diseases, podocyte loss triggers the activation and proliferation of neighboring parietal epithelial cells (PECs), leading to crescent formation in the bowman’s space and eventual glomerular injury. We previously identified potential key ligand-receptor interactions between injured podocytes that trigger the activation and proliferation of quiescent PECs in the setting of podocyte-specific knockdown of the pro-differentiation transcription factor, Krüppel-like factor 4 (KLF4). Recent evidence also suggests that injured podocytes might be capable of physically interacting with neighboring cells by extending their extracellular matrices to form bridges between cells. These physical extensions are unique from foot processes and filopodia of podocytes, yet their function is not known.

Methods

We investigated bridge formation in two models of proliferative glomerulopathy: mice with podocyte podocyte-specific loss of Klf4 (Klf4ΔPod) and dual reporter (RFP labeling podocytes, eGFP labeling PECs) mice treated with nephrotoxic serum (NTS). Immunohistochemistry, immunofluorescence staining, and electron microscopy was used to identify extracellular matrix extensions in glomeruli. Single nucleus (sn)RNA-seq was performed on both injury models.

Results

Periodic-acid schiff staining and electron microscopy revealed de novo bridges in mice with podocyte-specific loss of Klf4 and in mice treated with NTS. Staining NTS-treated dual reporter mice for CD44, a marker of activated PECs, revealed both RFP+ and CD44+ extensions. We also observed colocalization of RFP and CD44, suggesting that both podocytes and PECs are capable of forming bridges. Enrichment analysis of differentially expressed genes between wild type and Klf4ΔPod mice and IgG and NTS treatment showed common upregulated pathways including focal adhesion (ACTN1, ITGA6, BIRC3), axon guidance (ROCK2, MYL12A, MYL21B), and actin cytoskeleton regulation (ACTN1, FGFR2, MYH9, MSN) in the podocyte and PEC clusters.

Conclusion

This is the first study to demonstrate that glomerular bridges in proliferative glomerulopathy originate from both injured podocytes and activated PECs.

Funding

  • NIDDK Support