title: Nuclear factor of activated T cells mediates oxidised LDL-induced calcification of vascular smooth muscle cells.
abstract: OBJECTIVE
  Vascular calcification is a prominent feature of both atherosclerosis and diabetes, and is clinically associated with osteoporosis. The expression of bone-regulatory factors and the impact of oxidative stress in aortic calcification are well-documented. Recently, nuclear factor of activated T cells (NFAT) cytoplasmic, calcineurin-dependent 1 (NFATc1) was identified in calcified aortic valves and has been implicated in vascular calcification. Therefore, we assessed the mechanisms of osteogenic transdifferentiation of vascular smooth muscle cells induced by oxidised LDL (oxLDL) and evaluated the role of NFAT in this process.
  
  
  METHODS
  Human coronary artery smooth muscle cells (HCASMCs) were cultured for 21 days in medium supplemented with oxLDL. NFAT was inhibited using the NFAT inhibitor VIVIT, or by knockdown with small interfering RNA (siRNA). Osteogenic transdifferentiation was assessed by gene expression, matrix mineralisation and alkaline phosphatase activity.
  
  
  RESULTS
  Exposure to oxLDL caused the transformation of HCASMCs towards an osteoblast-like phenotype based on increased mineral matrix formation and RUNX2 expression. NFATc1 blockade completely prevented oxLDL-induced osteogenic transformation of HCASMCs as well as oxLDL-induced stimulation of osteoblast differentiation. In contrast, matrix mineralisation induced by osteogenic medium was independent of the NFAT pathway. Of note, oxLDL-conditioned medium from HCASMCs transferred to bone cells promoted osteoblast mineralisation. Consistent with these in vitro findings, diabetic rats with a twofold increase in oxidised lipid levels displayed higher aortic calcium concentrations and increased expression of osteogenic markers and production of NFATc1.
  
  
  CONCLUSIONS
  Our results identify the NFAT signalling pathway as a novel regulator of oxLDL-induced transdifferentiation of vascular smooth muscle cells towards an osteoblast-like phenotype.
authors.affiliation: Division of Endocrinology, Diabetes and Metabolic Bone Diseases, Department of Medicine III, Technical University Medical Center, Fetscherstraße 74, 01307 Dresden, Germany.
authors.forename: C
authors.initials: C
authors.lastname: Goettsch
authors.forename: M
authors.initials: M
authors.lastname: Rauner
authors.forename: C
authors.initials: C
authors.lastname: Hamann
authors.forename: K
authors.initials: K
authors.lastname: Sinningen
authors.forename: U
authors.initials: U
authors.lastname: Hempel
authors.forename: S R
authors.initials: SR
authors.lastname: Bornstein
authors.forename: L C
authors.initials: LC
authors.lastname: Hofbauer
mesh_terms: D000818:Animals; D002114:Calcinosis / Q000139:chemically induced / Q000378:metabolism*; D002454:Cell Differentiation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D050659:Core Binding Factor Alpha 1 Subunit / Q000235:genetics / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D006801:Humans; D008077:Lipoproteins, LDL / Q000494:pharmacology*; D009131:Muscle, Smooth, Vascular / Q000166:cytology*; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D050778:NFATC Transcription Factors / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D009842:Oligopeptides / Q000494:pharmacology; D010006:Osteoblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D018384:Oxidative Stress / Q000187:drug effects; D051381:Rats; D011924:Rats, Zucker; D060888:Real-Time Polymerase Chain Reaction
chemical_list: D050659:Core Binding Factor Alpha 1 Subunit; D008077:Lipoproteins, LDL; D050778:NFATC Transcription Factors; D009842:Oligopeptides; C121611:VIVIT peptide; C099252:oxidized low density lipoprotein