Hyperphosphatemia-induced nanocrystals upregulate the expression of bone morphogenetic protein-2 and osteopontin genes in mouse smooth muscle cells in vitro

Kidney Int. 2011 Feb;79(4):414-22. doi: 10.1038/ki.2010.390. Epub 2010 Oct 13.


Vascular calcification, which contributes to cardiovascular disease in patients with uremic hyperphosphatemia, is associated with vascular cell expression of osteogenic genes, including bone morphogenetic protein (BMP)-2 and osteopontin (OPN). High inorganic phosphate levels in vitro stimulate the osteogenic conversion of smooth muscle cells; however, the mechanism governing this is not clear. We found that high-phosphate medium increased the expression of BMP-2 and OPN in mouse smooth muscle cells in culture. However, this effect was lost in the presence of the mineralization inhibitor, pyrophosphate, suggesting a contribution of calcium phosphate crystals. Addition of 1-2 mmol/l phosphate alone to growth medium was sufficient to induce nanosized crystals after 1 day at 37 °C. Isolated crystals were about 160 nm in diameter and had a calcium to phosphate ratio of 1.35, consistent with the hydroxyapatite precursor octacalcium phosphate. Nanocrystal formation increased fourfold in the absence of serum, was blocked by fetuin-A, and was dependent on time and on the concentrations of phosphate and calcium. Purified synthetic hydroxyapatite nanocrystals and isolated high-phosphate-induced nanocrystals, but not nanocrystal-free high-phosphate medium, also induced BMP-2 and OPN. Thus, our results suggest that BMP-2 and OPN are induced by calcium phosphate nanocrystals, rather than soluble phosphate. This mechanism may contribute, in part, to hyperphosphatemia-related vascular cell differentiation and calcification.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Base Sequence
  • Bone Morphogenetic Protein 2 / genetics*
  • Calcinosis / etiology
  • Calcinosis / metabolism
  • Calcinosis / pathology
  • Calcium Phosphates / metabolism
  • Calcium Phosphates / pharmacology
  • Cell Differentiation
  • Cell Line
  • DNA Primers / genetics
  • Hyperphosphatemia / complications
  • Hyperphosphatemia / genetics*
  • Hyperphosphatemia / metabolism*
  • In Vitro Techniques
  • Mice
  • Microscopy, Electron, Scanning
  • Myocytes, Smooth Muscle / drug effects
  • Myocytes, Smooth Muscle / metabolism*
  • Myocytes, Smooth Muscle / pathology
  • Nanoparticles / chemistry*
  • Nanoparticles / ultrastructure
  • Osteogenesis / genetics
  • Osteopontin / genetics*
  • Up-Regulation / drug effects


  • Bmp2 protein, mouse
  • Bone Morphogenetic Protein 2
  • Calcium Phosphates
  • DNA Primers
  • Spp1 protein, mouse
  • Osteopontin
  • calcium phosphate