APE1/Ref-1 Inhibits Phosphate-Induced Calcification and Osteoblastic Phenotype Changes in Vascular Smooth Muscle Cells

Int J Mol Sci. 2017 Sep 25;18(10):2053. doi: 10.3390/ijms18102053.


Vascular calcification plays a role in the pathogenesis of atherosclerosis, diabetes, and chronic kidney disease; however, the role of apurinic/apyrimidinic endonuclease 1/redox factor-1 (APE1/Ref-1) in inorganic phosphate (Pi)-induced vascular smooth muscle cell (VSMC) calcification remains unknown. In this study, we investigated the possible role of APE1/Ref-1 in Pi-induced VSMC calcification. We observed that Pi decreased endogenous APE1/Ref-1 expression and promoter activity in VSMCs, and that adenoviral overexpression of APE1/Ref-1 inhibited Pi-induced calcification in VSMCs and in an ex vivo organ culture of a rat aorta. However, a redox mutant of APE1/Ref-1(C65A/C93A) did not reduce Pi-induced calcification in VSMCs, suggesting APE1/Ref-1-mediated redox function against vascular calcification. Additionally, APE1/Ref-1 overexpression inhibited Pi-induced intracellular and mitochondrial reactive oxygen species production, and APE1/Ref-1 overexpression resulted in decreased Pi-induced lactate dehydrogenase activity, pro-apoptotic Bax levels, and increased anti-apoptotic Bcl-2 protein levels. Furthermore, APE1/Ref-1 inhibited Pi-induced osteoblastic differentiation associated with alkaline phosphatase activity and inhibited Pi-exposure-induced loss of the smooth muscle phenotype. Our findings provided valuable insights into the redox function of APE1/Ref-1 in preventing Pi-induced VSMC calcification by inhibiting oxidative stress and osteoblastic differentiation, resulting in prevention of altered osteoblastic phenotypes in VSMCs.

Keywords: APE1/Ref-1; inorganic phosphate; vascular calcification; vascular smooth muscle cells.

MeSH terms

  • Animals
  • Apoptosis / drug effects
  • Apoptosis / genetics
  • Calcification, Physiologic / drug effects
  • Calcification, Physiologic / genetics*
  • Cell Differentiation / drug effects
  • Cell Differentiation / genetics
  • Cells, Cultured
  • DNA-(Apurinic or Apyrimidinic Site) Lyase / genetics*
  • DNA-(Apurinic or Apyrimidinic Site) Lyase / metabolism
  • Gene Expression Regulation / drug effects
  • Gene Silencing
  • Male
  • Mitochondria / metabolism
  • Muscle, Smooth, Vascular / cytology*
  • Muscle, Smooth, Vascular / pathology
  • Mutation
  • Myocytes, Smooth Muscle / drug effects
  • Myocytes, Smooth Muscle / metabolism*
  • Osteoblasts / drug effects
  • Osteoblasts / metabolism*
  • Oxidation-Reduction
  • Phenotype*
  • Phosphates / metabolism
  • Phosphates / pharmacology
  • RNA Interference
  • Rats
  • Reactive Oxygen Species / metabolism
  • Vascular Calcification / genetics
  • Vascular Calcification / metabolism
  • Vascular Calcification / pathology


  • Phosphates
  • Reactive Oxygen Species
  • Apex1 protein, rat
  • DNA-(Apurinic or Apyrimidinic Site) Lyase