Klotho/FGF23 axis mediates high phosphate-induced vascular calcification in vascular smooth muscle cells via Wnt7b/β-catenin pathway

Kaohsiung J Med Sci. 2019 Jul;35(7):393-400. doi: 10.1002/kjm2.12072. Epub 2019 Apr 18.

Abstract

Vascular calcification (VC) plays as a critical role on cardiovascular disease (CVD) and acts as a notable risk factor in cardiovascular system. Vascular smooth muscle cells (VSMCs) calcification can be triggered by high phosphate treatment; however, the explicit mechanism remains unclear. In the present study, we isolated VSMCs from primary rat artery, applied β-GP (β-glycerophosphate) for inducing VSMCs calcification in vitro to explore the mechanism of phosphate-induced calcification in VSMCs. Alizarin red staining was performed to assess the mineralization in VSMCs. Calcium deposition experiment was taken to evaluate the calcium content. ALP staining was determined to assess the ALP activity. The recombinant adenoviruses were constructed for the overexpression of Klotho and FGF23, respectively. qRT-PCR and western blot analysis were subjected to measure the expression of Klotho/FGF23 and correlated genes among Wnt7b/β-catenin pathway. We found that the calcium content was obviously increased and Alizarin red staining was positive in calcification group exposure with high phosphate in a time-dependent manner. The expression of Klotho and FGF23 was significantly decreased in the calcification group. However, overexpression of Klotho and FGF23 markedly reversed VSMCs calcification stimulating with high phosphate treatment. Moreover, Wnt7b/β-catenin inhibitor DKK1 could partly attenuate the effect of high phosphate on calcified VSMCs. These findings demonstrated that Klotho/FGF23 axis could modulate high phosphate-induced VSMCs calcification via Wnt7b/β-catenin signaling pathway. Our findings unravel that Klotho/FGF23- Wnt7b/β-catenin axis functions as a crucial role in the VSMCs calcification.

Keywords: FGF23; Klotho; Wnt7b/β-catenin; high phosphate; vascular calcification.

MeSH terms

  • Alkaline Phosphatase / genetics
  • Alkaline Phosphatase / metabolism
  • Animals
  • Aorta / cytology
  • Aorta / metabolism
  • Calcium / metabolism
  • Fibroblast Growth Factor-23
  • Fibroblast Growth Factors / genetics*
  • Fibroblast Growth Factors / metabolism
  • Gene Expression Regulation
  • Glucuronidase / genetics*
  • Glucuronidase / metabolism
  • Glycerophosphates / pharmacology*
  • Humans
  • Intercellular Signaling Peptides and Proteins / biosynthesis
  • Intercellular Signaling Peptides and Proteins / pharmacology
  • Klotho Proteins
  • Male
  • Myocytes, Smooth Muscle / cytology
  • Myocytes, Smooth Muscle / drug effects*
  • Myocytes, Smooth Muscle / metabolism
  • Primary Cell Culture
  • Rats
  • Rats, Sprague-Dawley
  • Recombinant Proteins / biosynthesis
  • Recombinant Proteins / pharmacology
  • Signal Transduction
  • Vascular Calcification / genetics*
  • Wnt Proteins / antagonists & inhibitors
  • Wnt Proteins / genetics*
  • Wnt Proteins / metabolism
  • beta Catenin / antagonists & inhibitors
  • beta Catenin / genetics*
  • beta Catenin / metabolism

Substances

  • Ctnnb1 protein, rat
  • DKK1 protein, human
  • FGF23 protein, human
  • Fgf23 protein, rat
  • Glycerophosphates
  • Intercellular Signaling Peptides and Proteins
  • Recombinant Proteins
  • Wnt Proteins
  • Wnt7b protein, rat
  • beta Catenin
  • Fibroblast Growth Factors
  • Fibroblast Growth Factor-23
  • Alkaline Phosphatase
  • Glucuronidase
  • Klotho Proteins
  • Calcium
  • beta-glycerophosphoric acid