title: Magnesium inhibits Wnt/β-catenin activity and reverses the osteogenic transformation of vascular smooth muscle cells.
abstract: Magnesium reduces vascular smooth muscle cell (VSMC) calcification in vitro but the mechanism has not been revealed so far. This work used only slightly increased magnesium levels and aimed at determining: a) whether inhibition of magnesium transport into the cell influences VSMC calcification, b) whether Wnt/β-catenin signaling, a key mediator of osteogenic differentiation, is modified by magnesium and c) whether magnesium can influence already established vascular calcification. Human VSMC incubated with high phosphate (3.3 mM) and moderately elevated magnesium (1.4 mM) significantly reduced VSMC calcification and expression of the osteogenic transcription factors Cbfa-1 and osterix, and up-regulated expression of the natural calcification inhibitors matrix Gla protein (MGP) and osteoprotegerin (OPG). The protective effects of magnesium on calcification and expression of osteogenic markers were no longer observed in VSMC cultured with an inhibitor of cellular magnesium transport (2-aminoethoxy-diphenylborate [2-APB]). High phosphate induced activation of Wnt/β-catenin pathway as demonstrated by the translocation of β-catenin into the nucleus, increased expression of the frizzled-3 gene, and downregulation of Dkk-1 gene, a specific antagonist of the Wnt/β-catenin signaling pathway. The addition of magnesium however inhibited phosphate-induced activation of Wnt/β-catenin signaling pathway. Furthermore, TRPM7 silencing using siRNA resulted in activation of Wnt/β-catenin signaling pathway. Additional experiments were performed to test the ability of magnesium to halt the progression of already established VSMC calcification in vitro. The delayed addition of magnesium decreased calcium content, down-regulated Cbfa-1 and osterix and up-regulated MGP and OPG, when compared with a control group. This effect was not observed when 2-APB was added. In conclusion, magnesium transport through the cell membrane is important to inhibit VSMC calcification in vitro. Inhibition of Wnt/β-catenin by magnesium is one potential intracellular mechanism by which this anti-calcifying effect is achieved.
authors.affiliation: IMIBIC, Hospital Univ Reina Sofia, REDInRen, Cordoba, Spain ; Depto. Medicina y Cirugía Animal, University of Cordoba, Cordoba, Spain.
authors.forename: Addy
authors.initials: A
authors.lastname: Montes de Oca
authors.affiliation: IMIBIC, Hospital Univ Reina Sofia, REDInRen, Cordoba, Spain ; Depto. Medicina y Cirugía Animal, University of Cordoba, Cordoba, Spain.
authors.forename: Fatima
authors.initials: F
authors.lastname: Guerrero
authors.affiliation: IMIBIC, Hospital Univ Reina Sofia, REDInRen, Cordoba, Spain.
authors.forename: Julio M
authors.initials: JM
authors.lastname: Martinez-Moreno
authors.affiliation: IMIBIC, Hospital Univ Reina Sofia, REDInRen, Cordoba, Spain.
authors.forename: Juan A
authors.initials: JA
authors.lastname: Madueño
authors.affiliation: IMIBIC, Hospital Univ Reina Sofia, REDInRen, Cordoba, Spain.
authors.forename: Carmen
authors.initials: C
authors.lastname: Herencia
authors.affiliation: Depto. Medicina y Cirugía Animal, University of Cordoba, Cordoba, Spain.
authors.forename: Alan
authors.initials: A
authors.lastname: Peralta
authors.affiliation: IMIBIC, Hospital Univ Reina Sofia, REDInRen, Cordoba, Spain ; Lipid and Atherosclerosis Unit, IMIBIC/Reina Sofia University Hospital/University of Cordoba, and CIBER Fisiopatologia Obesidad y Nutricion (CIBEROBN), Instituto de Salud Carlos III, Cordoba, Spain.
authors.forename: Yolanda
authors.initials: Y
authors.lastname: Almaden
authors.affiliation: Depto. Medicina y Cirugía Animal, University of Cordoba, Cordoba, Spain.
authors.forename: Ignacio
authors.initials: I
authors.lastname: Lopez
authors.affiliation: Depto. Medicina y Cirugía Animal, University of Cordoba, Cordoba, Spain.
authors.forename: Escolastico
authors.initials: E
authors.lastname: Aguilera-Tejero
authors.affiliation: Fresenius Medical Care Deutschland GmbH, Bad Homburg, Germany.
authors.forename: Kristina
authors.initials: K
authors.lastname: Gundlach
authors.affiliation: Fresenius Medical Care Deutschland GmbH, Bad Homburg, Germany.
authors.forename: Janine
authors.initials: J
authors.lastname: Büchel
authors.affiliation: Fresenius Medical Care Deutschland GmbH, Bad Homburg, Germany.
authors.forename: Mirjam E
authors.initials: ME
authors.lastname: Peter
authors.affiliation: Department of Nephrology, University of Duesseldorf, Dusseldorf, Germany.
authors.forename: Jutta
authors.initials: J
authors.lastname: Passlick-Deetjen
authors.affiliation: IMIBIC, Hospital Univ Reina Sofia, REDInRen, Cordoba, Spain ; Servicio de Nefrologia, Hospital Univ Reina Sofia, REDInRen, Cordoba, Spain.
authors.forename: Mariano
authors.initials: M
authors.lastname: Rodriguez
authors.affiliation: IMIBIC, Hospital Univ Reina Sofia, REDInRen, Cordoba, Spain.
authors.forename: Juan R
authors.initials: JR
authors.lastname: Muñoz-Castañeda
mesh_terms: D001896:Boron Compounds / Q000494:pharmacology; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism; D002478:Cells, Cultured; D016326:Extracellular Matrix Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008274:Magnesium / Q000494:pharmacology*; D009131:Muscle, Smooth, Vascular / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism; D010012:Osteogenesis / Q000187:drug effects*; D053244:Osteoprotegerin / Q000235:genetics / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects; D050053:TRPM Cation Channels / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D061205:Vascular Calcification / Q000188:drug therapy* / Q000378:metabolism; D051153:Wnt Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D051176:beta Catenin / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D000097568:Matrix Gla Protein
chemical_list: D001896:Boron Compounds; D002135:Calcium-Binding Proteins; D016326:Extracellular Matrix Proteins; D008274:Magnesium; D053244:Osteoprotegerin; D017346:Protein Serine-Threonine Kinases; D012333:RNA, Messenger; D050053:TRPM Cation Channels; D051153:Wnt Proteins; D051176:beta Catenin; D000097568:Matrix Gla Protein; C506030:TNFRSF11B protein, human; C109986:2-aminoethoxydiphenyl borate; C486583:TRPM7 protein, human