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Abstract: TH-OR046

Physiologic Regulation of Systemic Klotho Levels by Renal CaSR Signaling in Response to CaSR Ligands and Extracellular pH

Session Information

Category: Bone and Mineral Metabolism

  • 401 Bone and Mineral Metabolism: Basic

Authors

  • Miller, R. Tyler, University of Texas Southwestern Medical School, Dallas, Texas, United States
  • Yoon, Joonho, UTsouthwestern medical center, Dallas, Texas, United States
  • Liu, Zhenan, UT southwestern medical center, DALLAS, Texas, United States
  • Lee, Eunyoung, UT Southwestern Medical Center, Dallas, Texas, United States
  • Chang, Audrey N., UTSouthwestern Medical Center, Dallas, Texas, United States
  • Moe, Orson W., University of Texas Southwestern Medical Center, Dallas, Texas, United States
Background

Chronic kidney disease (CKD) progresses to end-stage renal disease accompanied by complications resembling the premature multi-organ failure akin to the Klotho-hypomorphic - mice (Kl/Kl). The kidney is the source of soluble Klotho (sKlotho), and as renal disease progresses serum and urine sKlotho levels fall, and patients acquire characteristics resembling the of Kl/Kl mice. Pharmacologic or dietary alkaline supplementation slows progression of CKD even in stages 3 and 4. The mechanism(s) by which HCO3 supplementation or alkaline diets work and physiologic mechanisms by which sKlotho levels might be regulated are unknown.

Methods

We measured: 1) urine and serum Kotho in mice treated with calcimimetics or alkali: 2) Klotho release in medium from minced mouse kidneys, and 3) medium Klotho from HEK-293 cells treated with calcimimetics, HCO3, and ADAM protease inhibitors and expressing the calcium-sensing receptor (CaSR) and Klotho. The CaSR, Klotho, and ADAM10 were co-localized in mouse kidneys and in cells expressing the CaSR and Klotho using differential centrifugation, co-IP, and immunofluorescent staining visualized by confocal microscopy.

Results

In intact mice, minced kidneys, and cultured cells, (CaSR) activation with high Ca or calcimimetics increases sKlotho levels via ADAM10-mediated shedding. Alkaline pH values increase, and acid pH values decrease CaSR signaling. Alkali treatment increases serum and urine sKlotho in mice, Klotho shedding in mouse kidney homogenates and cultured cells in a CaSR-dependent manner. Oral K citrate for 72 hrs increases serum and urine Klotho in human volunteers. ADAM10-dependence was demonstrated using the ADAM10 inhibitor GI 254023X and siRNA. In HEK-293 cells the CaSR, Klotho, and ADAM10 form cell surface aggregates that disperse following CaSR activation, but not with ADAM10 inhibition.

Conclusion

We define a novel physiologic mechanism for regulation of sKlotho levels by the renal CaSR-ADAM10-Klotho pathway that can be modified by pH. We predict that acidosis accelerates, and alkalinization slows the rate of loss of renal function in CKD partly because acid decreases, and alkaline increases renal CaSR-stimulated Klotho shedding from the kidney.

Funding

  • NIDDK Support