Kidney Week

Abstract: SA-PO472

Pannexin-1 Mediates Fluid Shear Stress-Sensitive Purinergic Signaling and Cyst Growth in Polycystic Kidney Disease

Session Information

• Cystic Kidney Diseases: Basic/Translational
  November 09, 2019 | Location: Exhibit Hall, Walter E. Washington Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: Genetic Diseases of the Kidneys

• 1001 Genetic Diseases of the Kidneys: Cystic

Authors

• Verschuren, Eric, Radboud university medical center, Nijmegen, Netherlands

Eric Verschuren,

• Rigalli, Juan pablo, Radboud university medical center, Nijmegen, Netherlands

Juan pablo Rigalli,

• Castenmiller, Charlotte, Radboud university medical center, Nijmegen, Netherlands

Charlotte Castenmiller,

• Bindels, René J., Radboud university medical center, Nijmegen, Netherlands

René J. Bindels,

• Peters, Dorien J.M., Leiden University Medical Center, Leiden, Netherlands

Dorien J.M. Peters,

• Arjona, Francisco J., Radboud university medical center, Nijmegen, Netherlands

Francisco J. Arjona,

• Hoenderop, Joost, Radboud university medical center, Nijmegen, Netherlands

Joost Hoenderop,

Background

Tubular ATP release is regulated by mechanosensation of fluid shear stress (FSS), but the molecular mechanism mediating this process is poorly understood. Extracellular ATP is implicated in polycystic kidney disease (PKD), where polycystin-1/polycystin-2 (PC1/PC2) is dysfunctional. In health, PC1/PC2 functions as a mechanosensory complex in the kidney. This study aims to provide new insights into renal ATP signaling under physiological conditions and PKD.

Methods

Microfluidic setups, pharmacologic inhibition of mTORC1 and pannexin-1 by rapamycin and brilliant blue FCF, respectively, and CRISPR/Cas9 loss-of-function approaches were combined to assess the ATP release by renal cells. Acute water loading by healthy human subjects was used to evaluate the ATP release under variable urinary flow. PKD models, using inducible kidney-specific Pkd1 knockout mice (iKsp-Pkd1^lox/lox) and zebrafish with translation blocking morpholino targeting the ortholog of human PKD2 (pkd2), were employed to study in vivo the relevance of the mechanisms disclosed in vitro.

Results

Immortalized renal mouse distal convoluted tubule 15 (mDCT15) cells subjected to FSS displayed an increased ATP release. Furthermore, inhibition of mTORC1 amplified this FSS-modulated ATP release. Inhibition of pannexin-1 decreased the FSS-modulated ATP release by these renal cells. To translate this in vitro phenomenon to the in vivo situation, healthy human subjects were exposed to acute water loading resulting in an increased urine production and ATP excretion. In precystic iKsp-Pkd1^-/- mice, increased renal pannexin-1 mRNA expression and urinary ATP were observed. Similarly, in renal Pkd1^-/- mDCT15 cells, elevated ATP release was observed upon FSS mechanosensation. In these cells, inhibition of mTORC1 failed to enhance ATP extrusion, but an increase in pannexin-1 mRNA expression was observed compared to renal Pkd1^+/+ mDCT15 cells. Importantly, inhibition of pannexin-1 in a zebrafish PKD model decreased renal cyst growth.

Conclusion

Our results suggest that renal pannexin-1 channels mediate ATP release from epithelial cells towards the tubular lumen. The redundancy of this mechanism in PKD, where urinary ATP levels are elevated, presents pannexin-1 as a new therapeutic target to prevent cyst growth in PKD.

Funding

• Government Support - Non-U.S.