title: Role of Noncanonical Wnt Signaling Pathway in Human Aortic Valve Calcification.
abstract: OBJECTIVE
  The mechanisms underlying the pathogenesis of aortic valve calcification remain unclear. With accumulating evidence demonstrating that valve calcification recapitulates bone development, the crucial roles of noncanonical Wnt ligands WNT5a, WNT5b, and WNT11 in osteogenesis make them critical targets in the study of aortic valve calcification.
  
  
  RESULTS
  Using immunohistochemistry, real-time qPCR, Western blotting, and tissue culture, we examined the tissue distribution of WNT5a, WNT5b, and WNT11 in noncalcified and calcified aortic valves and their effects on human aortic valve interstitial cells (HAVICs). Only focal strong immunostaining for WNT5a was seen in and around areas of calcification. Abundant immunostaining for WNT5b and WNT11 was seen in inflammatory cells, fibrosis, and activated myofibroblasts in areas of calcified foci. There was significant correlation between WNT5b and WNT11 overall staining and presence of calcification, lipid score, fibrosis, and microvessels (P<0.05). Real-time qPCR and Western blotting revealed abundant expression of both Wnts in stenotic aortic valves, particularly in bicuspid valves. Incubation of HAVICs from noncalcified valves with the 3 noncanonical Wnts significantly increased cell apoptosis and calcification (P<0.05). Treatment of HAVICs with the mitogen-activated protein kinase-38β and GSK3β inhibitors significantly reduced their mineralization (P<0.01). Raman spectroscopy identified the inorganic phosphate deposits as hydroxyapatite and showed a significant increase in hydroxyapatite deposition in HAVICs in response to WNT5a and WNT11 (P<0.05). Similar crystallinity was seen in the deposits found in HAVICs treated with Wnts and in calcified human aortic valves.
  
  
  CONCLUSIONS
  These findings suggest a potential role for noncanonical Wnt signaling in the pathogenesis of aortic valve calcification.
authors.affiliation: From the Division of Cardiology and Division of Cardiac Surgery, McGill University Health Centre, Montreal, Quebec, Canada (I.A., B.Y., H.A.-K., B.B., L.O., M.A.-R., B.d.V., D.S.-T., A.S.); Department of Material Engineering, McGill University, Montreal, Quebec, Canada (M.C., O.G.); and Montreal Heart Institute, Université de Montréal, Montreal, Quebec, Canada (E.R., J.C.T.).
authors.forename: Isabella
authors.initials: I
authors.lastname: Albanese
authors.affiliation: From the Division of Cardiology and Division of Cardiac Surgery, McGill University Health Centre, Montreal, Quebec, Canada (I.A., B.Y., H.A.-K., B.B., L.O., M.A.-R., B.d.V., D.S.-T., A.S.); Department of Material Engineering, McGill University, Montreal, Quebec, Canada (M.C., O.G.); and Montreal Heart Institute, Université de Montréal, Montreal, Quebec, Canada (E.R., J.C.T.).
authors.forename: Bin
authors.initials: B
authors.lastname: Yu
authors.affiliation: From the Division of Cardiology and Division of Cardiac Surgery, McGill University Health Centre, Montreal, Quebec, Canada (I.A., B.Y., H.A.-K., B.B., L.O., M.A.-R., B.d.V., D.S.-T., A.S.); Department of Material Engineering, McGill University, Montreal, Quebec, Canada (M.C., O.G.); and Montreal Heart Institute, Université de Montréal, Montreal, Quebec, Canada (E.R., J.C.T.).
authors.forename: Hamood
authors.initials: H
authors.lastname: Al-Kindi
authors.affiliation: From the Division of Cardiology and Division of Cardiac Surgery, McGill University Health Centre, Montreal, Quebec, Canada (I.A., B.Y., H.A.-K., B.B., L.O., M.A.-R., B.d.V., D.S.-T., A.S.); Department of Material Engineering, McGill University, Montreal, Quebec, Canada (M.C., O.G.); and Montreal Heart Institute, Université de Montréal, Montreal, Quebec, Canada (E.R., J.C.T.).
authors.forename: Bianca
authors.initials: B
authors.lastname: Barratt
authors.affiliation: From the Division of Cardiology and Division of Cardiac Surgery, McGill University Health Centre, Montreal, Quebec, Canada (I.A., B.Y., H.A.-K., B.B., L.O., M.A.-R., B.d.V., D.S.-T., A.S.); Department of Material Engineering, McGill University, Montreal, Quebec, Canada (M.C., O.G.); and Montreal Heart Institute, Université de Montréal, Montreal, Quebec, Canada (E.R., J.C.T.).
authors.forename: Leah
authors.initials: L
authors.lastname: Ott
authors.affiliation: From the Division of Cardiology and Division of Cardiac Surgery, McGill University Health Centre, Montreal, Quebec, Canada (I.A., B.Y., H.A.-K., B.B., L.O., M.A.-R., B.d.V., D.S.-T., A.S.); Department of Material Engineering, McGill University, Montreal, Quebec, Canada (M.C., O.G.); and Montreal Heart Institute, Université de Montréal, Montreal, Quebec, Canada (E.R., J.C.T.).
authors.forename: Mohammad
authors.initials: M
authors.lastname: Al-Refai
authors.affiliation: From the Division of Cardiology and Division of Cardiac Surgery, McGill University Health Centre, Montreal, Quebec, Canada (I.A., B.Y., H.A.-K., B.B., L.O., M.A.-R., B.d.V., D.S.-T., A.S.); Department of Material Engineering, McGill University, Montreal, Quebec, Canada (M.C., O.G.); and Montreal Heart Institute, Université de Montréal, Montreal, Quebec, Canada (E.R., J.C.T.).
authors.forename: Benoit
authors.initials: B
authors.lastname: de Varennes
authors.affiliation: From the Division of Cardiology and Division of Cardiac Surgery, McGill University Health Centre, Montreal, Quebec, Canada (I.A., B.Y., H.A.-K., B.B., L.O., M.A.-R., B.d.V., D.S.-T., A.S.); Department of Material Engineering, McGill University, Montreal, Quebec, Canada (M.C., O.G.); and Montreal Heart Institute, Université de Montréal, Montreal, Quebec, Canada (E.R., J.C.T.).
authors.forename: Dominique
authors.initials: D
authors.lastname: Shum-Tim
authors.affiliation: From the Division of Cardiology and Division of Cardiac Surgery, McGill University Health Centre, Montreal, Quebec, Canada (I.A., B.Y., H.A.-K., B.B., L.O., M.A.-R., B.d.V., D.S.-T., A.S.); Department of Material Engineering, McGill University, Montreal, Quebec, Canada (M.C., O.G.); and Montreal Heart Institute, Université de Montréal, Montreal, Quebec, Canada (E.R., J.C.T.).
authors.forename: Marta
authors.initials: M
authors.lastname: Cerruti
authors.affiliation: From the Division of Cardiology and Division of Cardiac Surgery, McGill University Health Centre, Montreal, Quebec, Canada (I.A., B.Y., H.A.-K., B.B., L.O., M.A.-R., B.d.V., D.S.-T., A.S.); Department of Material Engineering, McGill University, Montreal, Quebec, Canada (M.C., O.G.); and Montreal Heart Institute, Université de Montréal, Montreal, Quebec, Canada (E.R., J.C.T.).
authors.forename: Ophélie
authors.initials: O
authors.lastname: Gourgas
authors.affiliation: From the Division of Cardiology and Division of Cardiac Surgery, McGill University Health Centre, Montreal, Quebec, Canada (I.A., B.Y., H.A.-K., B.B., L.O., M.A.-R., B.d.V., D.S.-T., A.S.); Department of Material Engineering, McGill University, Montreal, Quebec, Canada (M.C., O.G.); and Montreal Heart Institute, Université de Montréal, Montreal, Quebec, Canada (E.R., J.C.T.).
authors.forename: Eric
authors.initials: E
authors.lastname: Rhéaume
authors.affiliation: From the Division of Cardiology and Division of Cardiac Surgery, McGill University Health Centre, Montreal, Quebec, Canada (I.A., B.Y., H.A.-K., B.B., L.O., M.A.-R., B.d.V., D.S.-T., A.S.); Department of Material Engineering, McGill University, Montreal, Quebec, Canada (M.C., O.G.); and Montreal Heart Institute, Université de Montréal, Montreal, Quebec, Canada (E.R., J.C.T.).
authors.forename: Jean-Claude
authors.initials: JC
authors.lastname: Tardif
authors.affiliation: From the Division of Cardiology and Division of Cardiac Surgery, McGill University Health Centre, Montreal, Quebec, Canada (I.A., B.Y., H.A.-K., B.B., L.O., M.A.-R., B.d.V., D.S.-T., A.S.); Department of Material Engineering, McGill University, Montreal, Quebec, Canada (M.C., O.G.); and Montreal Heart Institute, Université de Montréal, Montreal, Quebec, Canada (E.R., J.C.T.). adel.schwertani@mcgill.ca.
authors.forename: Adel
authors.initials: A
authors.lastname: Schwertani
mesh_terms: D000368:Aged; D000369:Aged, 80 and over; D000469:Alkaline Phosphatase / Q000378:metabolism; D001021:Aortic Valve / Q000378:metabolism* / Q000473:pathology*; D001024:Aortic Valve Stenosis / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D017209:Apoptosis; D002114:Calcinosis / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D002478:Cells, Cultured; D017886:Durapatite / Q000378:metabolism; D005260:Female; D005355:Fibrosis; D005786:Gene Expression Regulation; D000071679:Glycogen Synthase Kinase 3 beta / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D008875:Middle Aged; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D010012:Osteogenesis / Q000235:genetics; D010766:Phosphorylation; D013059:Spectrum Analysis, Raman; D051153:Wnt Proteins / Q000235:genetics / Q000378:metabolism*; D060449:Wnt Signaling Pathway; D000071818:Wnt-5a Protein / Q000235:genetics / Q000378:metabolism*
chemical_list: C085389:WNT5A protein, human; C435831:WNT5B protein, human; D051153:Wnt Proteins; D000071818:Wnt-5a Protein; C540523:Wnt11 protein, human; D017886:Durapatite; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; D020928:Mitogen-Activated Protein Kinases; D000469:Alkaline Phosphatase
keywords: aortic valve; bone marrow; calcium; human; lipids