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Abstract: SA-PO777

Tuberous Sclerosis Complex (TSC) Renal Cyst Extracellular Vesicles (EVs) Have Unique miRNA and Protein Profile

Session Information

Category: Genetic Diseases of the Kidneys

  • 1201 Genetic Diseases of the Kidneys: Cystic

Author

  • Bissler, John J., The University of Tennessee Health Science Center College of Medicine, Memphis, Tennessee, United States
Background

Over one million people worldwide have tuberous sclerosis complex (TSC), and half exhibit renal cystic disease. There is an unmet clinical need as there are no approved therapies. We identified that TSC renal cystogenesis is a cell non-autonomous disease process. To address this, we investigated the role of extracellular vesicles (EVs) as a novel mechanism of renal cystic disease to find new therapeutic options. Understanding the mechanism(s) for TSC-associated renal cystogenesis has significant ramifications. Similar to the renal cystic disease-associated renal cell carcinoma seen in von Hippel Lindau disease, TSC-associated renal cystic disease also is associated with renal cell carcinoma in 4-5% of the patient. An in-depth understanding of renal cystogenesis and the abnormal proliferation that leads to renal cell carcinoma will lead to better prevention strategies and therapies.

Methods

After Institutional Review Board approval, we isolated TSC patient derived EVs and performed tunable resistive pulse sensing, electron microscopy, dynamic light scatteringm and western blot analysis to characterize the EVs. We also performed mass spectroscopy for protein analysis, and RNASeq to better understand the miRNA and lncRNA associated with this diease state. For comparison, we used TSC serum derived EVs. For the analysis we used unsupervised clustering with hierarchical clustering and Pearson's correlation coefficients as well as a principal component analysis.

Results

The EVs from the human cyst fluid showed the expected cup shape morphology. Western blot analyses revealed a signature for small EVs, which also contained aquaporin 2, supporting their origin from principal cells. Sizing by TRPS and DLS revealed the EVs were approximately 140 nm in diameter at a concentration of 109 EVs per ml. There was strong agreement among DLS, TRPS, and TEM sizing methods. The protein analysis revealed three major groups of proteins that clearly distinguish the EVs isolated from the renal cyst from the EVs isolated from plasma. These experiments clearly reveal the the cyst EVs are unique when compared to EVs derived from patients with TSC. Furthermore, there were unique miRNA and lncRNA signatures for the EVs isolated from the cysts.

Conclusion

EVs in TSC patient cyst fluid continue unique protein and RNA signatures that may offer new avenues into the biology of the diease, and more importantly, into possible therapeutic approaches.

Funding

  • Private Foundation Support