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Kidney Week

Abstract: PO0307

Renal Cell-Derived Extracellular Vesicles Improve Functional Phenotype of Kidney Tubuloids

Session Information

  • Bioengineering
    October 22, 2020 | Location: On-Demand
    Abstract Time: 10:00 AM - 12:00 PM

Category: Bioengineering

  • 300 Bioengineering

Authors

  • Lindoso, Rafael Soares, Utrecht Institute for Pharmaceutical Sciences, Utrecht, Netherlands
  • Mancheño Juncosa, Estela, Universiteit Utrecht, Utrecht, Utrecht, Netherlands
  • Yousef Yengej, Fjodor, University Medical Center Utrecht, Utrecht, Netherlands
  • Rookmaaker, Maarten B., University Medical Center Utrecht, Utrecht, Netherlands
  • Verhaar, Marianne C., University Medical Center Utrecht, Utrecht, Netherlands
  • Masereeuw, Rosalinde, Utrecht Institute for Pharmaceutical Sciences, Utrecht, Netherlands
Background

Kidney tubuloids (KT) are adult stem cell-derived organoid that hold great potential in therapeutic applications as cell source for bioartificial kidney (BAK). However, reproducing complete cell maturation and function remains a challenge. Extracellular vesicles (EVs) are cell-derived structures that regulate several cellular processes. In the kidney, EVs mediate intranephron communication through the transfer of bioactive molecules. This study aimed to investigate the use of renal EVs as modulator of KT phenotype by increasing the levels of organic anion transport 1 (OAT1), involved in renal waste handling, and explore their use for BAK engineering.

Methods

EVs from conditionally immortalized proximal tubule epithelial cells overexpressing OAT1 (ciPTEC-OAT1) were isolated with 100 K filters, quantified via nanoparticle tracking analysis and incubated with KT. The 24 h conditioned media (CM) of ciPTEC-OAT1, depleted or not of EVs, were used as controls. Gene expression was determined by QPCR and Western blotting. For renal tubule engineering, KT were seeded on hollow fibers and were exposed to EVs or CM. Monolayer integrity and cell polarity were analyzed by immunofluorescence and confocal microscopy.

Results

KT exposed to CM showed increased OAT1 levels (protein: 2.0±0.3-fold and mRNA: 2.8±0.5-fold). Moreover, EVs mimicked CM effects (2.6±0.4-fold), while CM EV depleted didn't induce OAT1. EVs were shown to contain OAT1 protein and mRNA as cargo. Visual observations of KT seeded on hollow fibers with CM containing EVs, presented slightly improvement in 3D tubular structure organization with the expression of tight junction protein (ZO-1) and cell polarity (apical cilia formation and Na+/K+-ATPase presence at the basolateral side) when compared with the control condition.

Conclusion

KT phenotype can be directed by renal EVs obtained from ciPTEC-OAT1. In addition, renal EVs can support KT to form tight monolayers on hollow fiber membranes. Further research is aimed at a full functional characterization of these bioengineered proximal tubules for application in BAK.

ACKNOWLEDGEMENTS
Work supported by Regenerative Medicine Crossing Borders (www.regmedxb.com). Powered by Health~Holland, Top Sector Life Sciences & Health. We acknowledge Prof. Hans Clevers for providing the kidney tubuloids.

Funding

  • Government Support - Non-U.S.