Kidney Week

Abstract: FR-PO536

Cell Volume-Sensitive Transcription Factor(s) Regulate Claudin-2 Expression in Epithelial Cells

Session Information

• Fluid, Electrolyte, Acid-Base Disorders: Basic
  November 03, 2023 | Location: Exhibit Hall, Pennsylvania Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: Fluid, Electrolytes, and Acid-Base Disorders

• 1101 Fluid, Electrolyte, and Acid-Base Disorders: Basic

Authors

• Koumangoye, Rainelli, Vanderbilt University Medical Center, Nashville, Tennessee, United States

Rainelli Koumangoye,

• Delpire, Eric J., Vanderbilt University Medical Center, Nashville, Tennessee, United States

Eric J. Delpire,

Background

The basolateral Na⁺-K⁺-2Cl^- cotransporter (NKCC1) plays a critical role in maintaining the intracellular concentrations of Na⁺, K⁺, and Cl^- ions, and cell volume. While it is well recognized that NKCC1 is critically important for vectorial transport of Na⁺, K⁺, and Cl^- across epithelia, its role in transcellular ions transport and epithelial barrier function is not well understood. Here we ask whether loss of NKCC1 function affect kidney and intestinal epithelial tight junction permeability.

Methods

Wild-type MDCK I and NKCCC1-deficient clones LKA3 and LKC1 were plated and polarized on transwell filters, and their TER was measured. Tight junction protein expression and localization was assessed by western blotting and immunofluorescence.

Results

The TER values of NKCC1-deficient MDCK I cells decreased by more than 20-fold, from 8000 Ω.cm² in parental MDCK I to 300-500 Ω.cm² in LKA3 and LKC1 clones. Western blot analysis showed that genetic ablation or pharmacologic of NKCC1 function increases the expression of the cation selective claudin-2. This is consistent with the observed decreased TER. In addition, growing MDCK I cells in Cl^--free, Na⁺-free or both Cl^- and Na⁺-free media upregulates claudin-2 expression.

Conclusion

Our data indicate that loss of NKCC1 function affects tight junction protein expression without affecting their localization. Particularly, claudin-2 was significantly upregulated. This led to an increase in paracellular flux of positively charged ions across the MDCK epithelial cell monolayers. Notably, this phenomenon is not unique to kidney epithelial cells cells, as loss of NKCC1 function in intestinal HT29 cells also led to significant decrease in TER and upregulation of claudin-2 expression.

Funding

• NIDDK Support