Research ArticleVascular Disease

Increased activity of TNAP compensates for reduced adenosine production and promotes ectopic calcification in the genetic disease ACDC

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Sci. Signal.  13 Dec 2016:
Vol. 9, Issue 458, pp. ra121
DOI: 10.1126/scisignal.aaf9109

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Understanding vascular calcification

ACDC is a rare genetic vascular calcification disease caused by loss of CD73, a secreted enzyme that converts adenosine monophosphate (AMP) to adenosine. Cells from ACDC patients have a compensatory increase in the phosphatase TNAP, which primarily catalyzes the conversion of pyrophosphate to inorganic phosphate but can also convert AMP to adenosine. Jin et al. generated induced pluripotent stem cells (iPSCs) from ACDC patients. Although in culture, these cells generated adenosine from AMP, the cells had decreased amounts of pyrophosphate, which inhibits calcification. ACDC patient–derived cells showed increased activation of the mTOR pathway, which promotes calcification. When injected into mice, the ACDC patient–derived iPSCs formed calcified teratomas. Treating mice bearing these teratomas with an adenosine receptor agonist, the mTOR inhibitor rapamycin, or etidronate (a drug that is structurally similar to pyrophosphate) reduced calcification in the teratomas, suggesting multiple potential strategies for treating ectopic calcification in ACDC patients and thereby alleviating the pain and peripheral ischemia associated with the disease.

Abstract

ACDC (arterial calcification due to deficiency of CD73) is an autosomal recessive disease resulting from loss-of-function mutations in NT5E, which encodes CD73, a 5′-ectonucleotidase that converts extracellular adenosine monophosphate to adenosine. ACDC patients display progressive calcification of lower extremity arteries, causing limb ischemia. Tissue-nonspecific alkaline phosphatase (TNAP), which converts pyrophosphate (PPi) to inorganic phosphate (Pi), and extracellular purine metabolism play important roles in other inherited forms of vascular calcification. Compared to cells from healthy subjects, induced pluripotent stem cell–derived mesenchymal stromal cells (iMSCs) from ACDC patients displayed accelerated calcification and increased TNAP activity when cultured under conditions that promote osteogenesis. TNAP activity generated adenosine in iMSCs derived from ACDC patients but not in iMSCs from control subjects, which have CD73. In response to osteogenic stimulation, ACDC patient–derived iMSCs had decreased amounts of the TNAP substrate PPi, an inhibitor of extracellular matrix calcification, and exhibited increased activation of AKT, mechanistic target of rapamycin (mTOR), and the 70-kDa ribosomal protein S6 kinase (p70S6K), a pathway that promotes calcification. In vivo, teratomas derived from ACDC patient cells showed extensive calcification and increased TNAP activity. Treating mice bearing these teratomas with an A2b adenosine receptor agonist, the mTOR inhibitor rapamycin, or the bisphosphonate etidronate reduced calcification. These results show that an increase of TNAP activity in ACDC contributes to ectopic calcification by disrupting the extracellular balance of PPi and Pi and identify potential therapeutic targets for ACDC.

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