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Abstract: FR-PO740

Extracellular Vesicles Rebalance Glomerular Endothelial Lipid Metabolism in CKD

Session Information

Category: Glomerular Diseases

  • 1401 Glomerular Diseases: Mechanisms, including Podocyte Biology

Authors

  • Sedrakyan, Sargis, Children's Hospital Los Angeles, Los Angeles, California, United States
  • Soloyan, Hasmik, Children's Hospital Los Angeles, Los Angeles, California, United States
  • Clair, Geremy, Pacific Northwest National Laboratory, Richland, Washington, United States
  • Cuala, Janielle M., University of Southern California Keck School of Medicine, Los Angeles, California, United States
  • Chomoyan, Hripsime, Children's Hospital Los Angeles, Los Angeles, California, United States
  • Zhang, Qi, Children's Hospital Los Angeles, Los Angeles, California, United States
  • Thornton, Matthew Edward, University of Southern California Keck School of Medicine, Los Angeles, California, United States
  • Georgia, Senta K., Children's Hospital Los Angeles, Los Angeles, California, United States
  • Cravedi, Paolo, Icahn School of Medicine at Mount Sinai, New York, New York, United States
  • Angeletti, Andrea, Istituto Giannina Gaslini, Genova, Liguria, Italy
  • De Filippo, Roger E., Children's Hospital Los Angeles, Los Angeles, California, United States
  • Da Sacco, Stefano, Children's Hospital Los Angeles, Los Angeles, California, United States
  • Perin, Laura, Children's Hospital Los Angeles, Los Angeles, California, United States
Background

Chronic kidney disease (CKD) affects more than 10% of the population worldwide, and our understanding of the mechanisms in many forms of CKD are poorly understood. Alport syndrome (AS), is a form of CKD caused by mutations in the collagen IVα3, α4, or α5 gene. Even though injury to glomerular endothelial cells (GEC) in AS is well established, the role of GEC in Alport progression has not been elucidated. Here, we describe the role of mitochondria and lipid metabolism in GEC injury in an animal model of AS, and the potential of using amniotic fluid stem cell (AFSC) derived extracellular vesicles, (EVs) as a rescue strategy to restore glomerular homeostasis.

Methods

The phasor approach to fluorescent lifetime imaging microscopy (FLIM) and flow cytometry was applied to evaluate the mitochondria and metabolic changes in GEC in AS vs WT mice. GEC isolated by FACS from Tek-tdT reporter AS and WT mice were compared by bulk RNA-seq, lipidomic and flow cytometric analysis. In vitro, silencing experiments on primary human primary GEC were performed to study the role of fatty acid synthase (FASN) in mitochondrial dysfunction and GEC damage. FASN-carrying AFSC-EVs and control EVs were applied both in vitro and in vivo to restore lipid homeostasis in GEC.

Results

FLIM studies showed strong correlation between the metabolic state of GEC and the age and severity of disease in AS mice. RNA-seq analysis revealed changes in the pathways associated with lipid metabolism and mitochondria function. Mitochondrial dysfunction was confirmed using flow cytometric analysis of the MitoTracker signal in tdTomato expressing GEC. Lipidome analysis revealed high abundance of triglycerides in AS GEC. We confirmed accumulation of lipid droplets in the glomeruli of AS mice and in FASN KO human primary GEC, in vitro. These results suggest potential mitochondrial dysfunction in GEC. AFSC derived EV treatment restored lipid homeostasis in GEC, both in vitro and in vivo.

Conclusion

We report for the first-time mitochondrial dysfunction in Alport GEC, and the ability of AFSC-derived EVs to rescue this phenotype. Better understanding of the metabolic changes in AS GEC could lead to the development of targeted new therapies also for other forms of CKD.

Funding

  • NIDDK Support