Pharmacological Inhibition of PHOSPHO1 Suppresses Vascular Smooth Muscle Cell Calcification

Tina Kiffer-Moreira, Manisha C. Yadav, Dongxing Zhu, Sonoko Narisawa, Campbell Sheen, Boguslaw Stec, Nicholas D. Cosford, Russell Dahl, Colin Farquharson, Marc F. Hoylaerts, Vicky E. MacRae, Jose Luis Millan*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

Abstract

Medial vascular calcification (MVC) is common in patients with chronic kidney disease, obesity, and aging. MVC is an actively regulated process that resembles skeletal mineralization, resulting from chondro-osteogenic transformation of vascular smooth muscle cells (VSMCs). Here, we used mineralizing murine VSMCs to study the expression of PHOSPHO1, a phosphatase that participates in the first step of matrix vesicles-mediated initiation of mineralization during endochondral ossification. Wild-type (WT) VSMCs cultured under calcifying conditions exhibited increased Phospho1 gene expression and Phospho1(-/-) VSMCs failed to mineralize in vitro. Using natural PHOSPHO1 substrates, potent and specific inhibitors of PHOSPHO1 were identified via high-throughput screening and mechanistic analysis and two of these inhibitors, designated MLS-0390838 and MLS-0263839, were selected for further analysis. Their effectiveness in preventing VSMC calcification by targeting PHOSPHO1 function was assessed, alone and in combination with a potent tissue-nonspecific alkaline phosphatase (TNAP) inhibitor MLS-0038949. PHOSPHO1 inhibition by MLS-0263839 in mineralizing WT cells (cultured with added inorganic phosphate) reduced calcification in culture to 41.8% +/- 2.0% of control. Combined inhibition of PHOSPHO1 by MLS-0263839 and TNAP by MLS-0038949 significantly reduced calcification to 20.9% +/- 0.74% of control. Furthermore, the dual inhibition strategy affected the expression of several mineralization-related enzymes while increasing expression of the smooth muscle cell marker Acta2. We conclude that PHOSPHO1 plays a critical role in VSMC mineralization and that "phosphatase inhibition" may be a useful therapeutic strategy to reduce MVC. (C) 2013 American Society for Bone and Mineral Research.

Original languageEnglish
Pages (from-to)81-91
Number of pages11
JournalJournal of Bone and Mineral Research
Volume28
Issue number1
Early online date14 Aug 2012
DOIs
Publication statusPublished - Jan 2013

Keywords

  • SKELETAL MINERALIZATION
  • SMALL-MOLECULES
  • ARTERIAL CALCIFICATION
  • ALKALINE PHOSPHATASE
  • MATRIX VESICLES
  • EADOCK DSS
  • KINETIC STUDIES
  • PYROPHOSPHATE
  • PHARMACOLOGICAL INHIBITORS
  • MODEL
  • HIGH-THROUGHPUT SCREENING
  • NONSPECIFIC ALKALINE-PHOSPHATASE
  • IDENTIFICATION
  • AORTIC CALCIFICATION
  • CHRONIC KIDNEY-DISEASE

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