ASN's Mission

To create a world without kidney diseases, the ASN Alliance for Kidney Health elevates care by educating and informing, driving breakthroughs and innovation, and advocating for policies that create transformative changes in kidney medicine throughout the world.

learn more

Contact ASN

1401 H St, NW, Ste 900, Washington, DC 20005

email@asn-online.org

202-640-4660

The Latest on X

Kidney Week

Abstract: FR-PO524

Inhibition of Calcium-Sensing Receptor in Mice Proximal Tubule Leads to Calcium Phosphate Crystalluria

Session Information

Category: Fluid, Electrolytes, and Acid-Base Disorders

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

Authors

  • Shin, Samuel, Washington DC VA Medical Center, Washington, District of Columbia, United States
  • Asico, Laureano D., The George Washington University School of Medicine and Health Sciences, Washington, District of Columbia, United States
  • Jose, Pedro A., The George Washington University School of Medicine and Health Sciences, Washington, District of Columbia, United States
  • Bandyopadhyay, Bidhan C., Washington DC VA Medical Center, Washington, District of Columbia, United States
Background

The proximal tubular (PT) Ca2+ transport is crucial for maintaining tubular Ca2+ levels to prevent downstream calcium phosphate (CaP) crystal formation. While PT Ca2+ transport is often described as paracellular, we found a transcellular Ca2+ transport pathway in the PT that is mediated by the interaction of calcium-sensing receptor (CaSR) and transient receptor potential canonical type 3 (TRPC3) channel. TRPC3 knockout mice exhibited hypercalciuria and microcalcifications arguing for a protective role of TRPC3 in preventing nephrocalcinosis. However, the role of CaSR in CaP crystal formation is unclear. Thus, we used a renal-selective knockdown of CaSR by the renal subcapsular infusion of CaSR siRNA to sort out the CaSR-mediated Ca2+ transport in the PT and its contribution to urinary CaP formation.

Methods

Adult male C57BL/6J mice were uninephrectomized 1 week before the renal implantation of osmotic minipumps. Osmotic minipumps (100 μL; flow rate 0.5 μL/h for 7 days) were filled with previously validated CaSR or non-silencing siRNA as control. siRNAs were dissolved in an in vivo transfection reagent under sterile conditions and was delivered via the osmotic minipumps into the renal cortex of each mouse. 24h urine collection and ion measurements were performed and collected urine were stained with AR pH 4.3 to detect CaP. Real time intracellular Ca2+ was measured in siRNA- or scramble-transfected isolated mice PT cells.

Results

In mice that underwent renal subcapsular infusion with siRNA for two weeks, 24h urine showed notable CaP crystal formations compared with that of control and scramble siRNA- infused mice. Renal cortical knockdown of CaSR did not significantly affect Na+, Cl-, and K+ excretion (normalized by creatinine) except for Ca2+. Fura-2 intracellular Ca2+ measurements in CaSR siRNA-transfected PT cells revealed decreased or nearly complete absence of Ca2+ entry after application with L-phenylalanine (CaSR agonist), confirming the effects of the siRNA.

Conclusion

The decreased Ca2+ influx due to CaSR inhibition suggest diminished tubular Ca2+ reabsorption confirming CaSR’s involvement in PT Ca2+ regulation. This study thus enhances our understanding about transcellular regulation of PT Ca2+ and its role in regulating urinary Ca2+ levels and subsequently CaP stone formation.

Funding

  • NIDDK Support