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Abstract: FR-PO989

Inhibition of TGF-β Unlocks Multiple Proximal Tubule Functions In Vitro

Session Information

  • CKD: Pathobiology - I
    November 04, 2022 | Location: Exhibit Hall, Orange County Convention Center‚ West Building
    Abstract Time: 10:00 AM - 12:00 PM

Category: CKD (Non-Dialysis)

  • 2203 CKD (Non-Dialysis): Mechanisms

Authors

  • Fissell, William Henry, Vanderbilt University Medical Center, Nashville, Tennessee, United States
  • Evans, Rachel C., Vanderbilt University Medical Center, Nashville, Tennessee, United States
  • Hunter, Kuniko, Vanderbilt University, Nashville, Tennessee, United States
  • Love, Harold D., Vanderbilt University Medical Center, Nashville, Tennessee, United States
  • Roy, Shuvo, University of California San Francisco, San Francisco, California, United States

Group or Team Name

  • The Kidney Project
Background

Renal proximal tubule cells are both essential to renal function and vulnerable to toxins and ischemia. In vitro, renal tubule cells quickly lose structural, biochemical, and fucntional features characteristic of their identity. This dedifferentiation of cell culture stress limits the use of tubule cell culture in drug screening and tiissue engineering. We examined the role of TGF-β in the dedifferentiation of cell culture stress.

Methods

Renal proximal tubule epithelial cells were grown to confluence in 50:50 DMEM/F12 on permeable supports under constant fluid shear stress. Half the wells (n=12) were treated with an inhibitor of TGF-b receptor 1, SB431542. Another 12 wells served as controls. Cells were cultured over 300 days. Confluence of the monolayer was assessed by dextran leak and all experiments were performed only on confluent monolayers. Apicobasal fluid transport, organic anion excretion, and glucose transport were measured. Inhibitors of NHE3 (tenapanor), OAT3 (probenecid), and SGLT2 (phlorizin) were used to examine the specificity of response.

Results

Renal tubule cells in control wells did not transport fluid volume, glucose, or organic anions; concentrations in apical and basolateral compartments were identical. Cells treated with SB431542 showed tenapanor-inhibitable volume transport, probenecid-inhibitable para-amino hippurate excretion, and phlorizin-inhibitable glucose uptake.

Conclusion

Startlingly, inhibition of a single cytokine receptor, TGF-bR1, was necessary and sufficient to unlock a wide range of differentiated tubule cell functions over a very prolonged period of time. It may be possible to extend the medical use of cell culture to applications requiring stable quantitative fidelity between the in vitro and in vivo phenotypes.

Funding

  • Private Foundation Support