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Abstract: TH-PO517

The New RANKL Receptor LGR4 Regulates Vascular Smooth Muscle Cell Calcification in CKD

Session Information

Category: Bone and Mineral Metabolism

  • 401 Bone and Mineral Metabolism: Basic

Authors

  • Carrillo-Lopez, Natalia, Bone and Mineral Research Unit. Hospital Universitario Central de Asturias. Instituto de Investigación Sanitaria del Principado de Asturias (ISPA), REDinREN-ISCIII, Oviedo, Spain
  • Martinez-Arias, Laura, Bone and Mineral Research Unit. Hospital Universitario Central de Asturias. Instituto de Investigación Sanitaria del Principado de Asturias (ISPA), REDinREN-ISCIII, Oviedo, Spain
  • Alonso-Montes, Cristina, Bone and Mineral Research Unit. Hospital Universitario Central de Asturias. Instituto de Investigación Sanitaria del Principado de Asturias (ISPA), REDinREN-ISCIII, Oviedo, Spain
  • Martin-Carro, Beatriz, Bone and Mineral Research Unit. Hospital Universitario Central de Asturias. Instituto de Investigación Sanitaria del Principado de Asturias (ISPA), REDinREN-ISCIII, Oviedo, Spain
  • Martin-Virgala, Julia, Bone and Mineral Research Unit. Hospital Universitario Central de Asturias. Instituto de Investigación Sanitaria del Principado de Asturias (ISPA), REDinREN-ISCIII, Oviedo, Spain
  • Palomo, Carmen, Department of Internal Medicine. Hospital Universitario Central de Asturias, Oviedo, Spain
  • Dusso, Adriana S., Bone and Mineral Research Unit. Hospital Universitario Central de Asturias. Instituto de Investigación Sanitaria del Principado de Asturias (ISPA), REDinREN-ISCIII, Oviedo, Spain
  • Naves, Manuel, Bone and Mineral Research Unit. Hospital Universitario Central de Asturias. Instituto de Investigación Sanitaria del Principado de Asturias (ISPA), REDinREN-ISCIII, Oviedo, Spain
  • Cannata-Andia, Jorge B., Bone and Mineral Research Unit. Hospital Universitario Central de Asturias. Instituto de Investigación Sanitaria del Principado de Asturias (ISPA), REDinREN-ISCIII, Oviedo, Spain
  • Panizo, Sara, Bone and Mineral Research Unit. Hospital Universitario Central de Asturias. Instituto de Investigación Sanitaria del Principado de Asturias (ISPA), REDinREN-ISCIII, Oviedo, Spain
Background

PTH is a main regulator of RANKL and OPG in bone remodeling. These 3 molecules have a well-known role in vascular calcification (VC). Due to controversies in the direct role of PTH in VC and the discovery of leucine-rich repeat-containing G-protein-coupled receptor 4 (LGR4) as a new RANKL receptor, we analyzed LGR4 involvement in VC in chronic kidney disease (CKD).

Methods

In vivo: We analyzed LGR4, RANK, RANKL and OPG gene expression and aortic calcium content ijn CKD rats fed for 14 weeks a normal phosphorus (0.6% -NP-) or a high phosphorus(0.9% -HP-) diet with or without parathyroidectomy (PTX) and PTH 1-34 supplementation aiming to normalize PTH. Rats with normal renal function fed a NP diet were used as reference. In vitro: We analyzed LGR4, RANK, RANKL and OPG expression and calcium content in vascular smooth muscle cells (VSMC) from rat aortas exposed to 3 different culture media (control, calcifying or calcifying plus 10-7/10-9M PTH). To characterize the pathways involved in calcification, the same parameters were examined in VSMC upon silencing of LGR4 or PTH1R (main PTH receptor) or after incubation with specific protein kinase (PK) A or PKC inhibitors, or with forskolin, a specific PKA agonist.

Results

CKD significantly increased aortic LGR4 and RANKL mRNA expression and decreased OPG, increasing aortic calcium content. These changes were greater in the group with higher PTH (CKD-HP) and were prevented by PTX. There were no changes in RANK expression under any of the experimental conditions. In VSMC, 10-7M PTH, but not 10-9M PTH, increased LGR4 and RANKL, reduced OPG expression with increases in calcium content. LGR4 and PTH1R silencing significantly attenuated the increases in calcium content induced by 10-7M PTH. Furthermore, silencing of PTH1R and PKA inhibition, but not PKC inhibition, prevented the increases in RANKL and LGR4 and OPG reduction. Exposure to forskolin corroborated these results.

Conclusion

In CKD, high PTH increases the aortic expression of LGR4 receptor and its ligand RANKL and decreases OPG inducing VC. These PTH actions in VSMC involve binding to PTH1R and PKA activation. Thus, LGR4 was identified for the first time as a pro-calcifying factor of VSMC in CKD.

Funding

  • Government Support - Non-U.S.