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Abstract: FR-OR103

Paracrine Effects of Injured Podocytes on the Transcriptome of Neighboring Cells

Session Information

Category: Glomerular Diseases

  • 1403 Podocyte Biology

Authors

  • Zeng, Yuting, University of Washington, Seattle, Washington, United States
  • Schweickart, Robert A., Cleveland Clinic, Cleveland, Ohio, United States
  • Pippin, Jeffrey W., University of Washington, Seattle, Washington, United States
  • Wessely, Oliver, Cleveland Clinic, Cleveland, Ohio, United States
  • Theberge, Ashleigh B., University of Washington, Seattle, Washington, United States
  • Shankland, Stuart J., University of Washington, Seattle, Washington, United States
Background

Knowledge gaps in glomerular diseases include how podocytes respond to different forms of injury and the crosstalk between injured podocytes and neighboring cells (healthy podocytes or parietal epithelial cells (PECs)). The goal of this study was to establish an in vitro microfluidic coculture model to distinguish podocyte's responses to different injuries, and to determine what mediators from injured podocytes cause paracrine damage to neighboring cells in the glomeruli.

Methods

We engineered an open microfluidic coculture device consisting of two separate but interconnectible chambers. The outer chamber was seeded with primary human podocytes and the inner chamber was seeded with human podocytes or PECs in different series of experiments. Following the induction of podocyte injury in the outer chamber by cytotoxic sheep anti-podocyte IgG (IgG), puromycin (PAN), Adriamycin (ADR), chambers were connected to study the paracrine effects between injured podocytes and naïve podocytes or PECs. Imaging, cell viability and RNA-sequencing were temporally measured in cells of both chambers.

Results

Injured human podocytes displayed foot process effacement, cell body shrinkage, and decreased cell viability. In the podocyte-podocyte coculture, gene set enrichment analysis showed several common pathways for injury by IgG, PAN, and ADR, including apoptosis, complement, p53, TNFα, IFN, Wnt/β-catenin, and reactive oxygen species (ROS). IgG and ADR but not PAN increased TGFβ or inflammatory response (IL2, IL6, KRAS). ADR and PAN but not IgG increased DNA repair. The naïve podocytes cocultured with injured ones showed similar enrichments. Among the 65 ligand-receptor pairs shared by all three injuries were CSF1 & CSF1 receptor, BMP2 & BMP type I receptors, IL1A & IL1 receptor, and GDNF & RET receptor. The transcriptomic results from podocyte-PEC experiments are pending.

Conclusion

The in vitro open microfluidic coculture device represents a new model for studying autocrine and paracrine signaling between injured podocytes and healthy neighboring cells. The paracrine injury response of podocytes is similar to the direct injury response. Mechanistic studies are currently ongoing to identify which ligands from the injured podocytes and which receptors on naïve podocytes mediate these paracrine effects.

Funding

  • NIDDK Support