Extracellular pyrophosphate metabolism and calcification in vascular smooth muscle

• Authors: Ricardo Villa-Bellosta, University of Zaragoza; Xiaonan Wang, Emory University; Jose Luis Millan, Sanford-Burnham Medical Research Institute; George R. Dubyak, Case Western Reserve University; W Charles O'Neill, Emory University
• Journal Title: AJP - Heart and Circulatory Physiology
• Volume: Volume 301, Number 1
• Publisher: American Physiological Society | 2011-07, Pages H61-H68
• Type of Work: Article | Post-print: After Peer Review
• Publisher DOI: http://doi.org/10.1152/ajpheart.01020.2010
• Subjects: Biophysics, Medical; Health Sciences, Toxicology; Health Sciences, Medicine and Surgery
• Keywords: medial calcification; alkaline phosphatase; ectonucleotide pyrophosphorylase; adenosine 5′-triphosphate; ankyrin

Abstract

Extracellular inorganic pyrophosphate (ePPi) is an important endogenous inhibitor of vascular calcification, but it is not known whether systemic or local vascular PPi metabolism controls calcification. To determine the role of ePPi in vascular smooth muscle, we identified the pathways responsible for ePPi production and hydrolysis in rat and mouse aortas and manipulated them to demonstrate their role in the calcification of isolated aortas in culture. Rat and mouse aortas contained mRNA for ectonucleotide pyrophosphatase/phosphodiesterases (NPP1–3), the putative PPi transporter ANK, and tissue-nonspecific alkaline phosphatase (TNAP). Synthesis of PPi from ATP in aortas was blocked by β,γ-methylene-ATP, an inhibitor of NPPs. Aortas from mice lacking NPP1 (Enpp1−/−) did not synthesize PPi from ATP and exhibited increased calcification in culture. Although ANK-mediated transport of PPi could not be demonstrated in aortas, aortas from mutant (ank/ank) mice calcified more in culture than did aortas from normal (ANK/ANK) mice. Hydrolysis of PPi was reduced 25% by β,γ-methylene-ATP and 50% by inhibition of TNAP. Hydrolysis of PPi was increased in cells overexpressing TNAP or NPP3 but not NPP1 and was not reduced in Enpp1−/− aortas. Overexpression of TNAP increased calcification of cultured aortas. The results show that smooth muscle NPP1 and TNAP control vascular calcification through effects on synthesis and hydrolysis of ePPi, indicating an important inhibitory role of locally produced PPi. Smooth muscle ANK also affects calcification, but this may not be mediated through transport of PPi. NPP3 is identified as an additional pyrophosphatase that could influence vascular calcification.