title: MFAP4 Promotes Vascular Smooth Muscle Migration, Proliferation and Accelerates Neointima Formation.
abstract: OBJECTIVE
  Arterial injury stimulates remodeling responses that, when excessive, lead to stenosis. These responses are influenced by integrin signaling in vascular smooth muscle cells (VSMCs). Microfibrillar-associated protein 4 (MFAP4) is an integrin ligand localized to extracellular matrix fibers in the vascular wall. The role of MFAP4 in vascular biology is unknown. We aimed to test the hypothesis that MFAP4 would enhance integrin-dependent VSMC activation.
  
  
  RESULTS
  We produced Mfap4-deficient (Mfap4(-/-)) mice and performed carotid artery ligation to explore the role of MFAP4 in vascular biology in vivo. Furthermore, we investigated the effects of MFAP4 in neointimal formation ex vivo and in primary VSMC and monocyte cultures in vitro. When challenged with carotid artery ligation, Mfap4(-/-) mice exhibited delayed neointimal formation, accompanied by early reduction in the number of proliferating medial and neointimal cells, as well as infiltrating leukocytes. Delayed neointimal formation was associated with decreased cross-sectional area of ligated Mfap4(-/-) carotid arteries resulting in lumen narrowing 28 days after ligation. MFAP4 blockade prohibited the formation of neointimal hyperplasia ex vivo. Moreover, we demonstrated that MFAP4 is a ligand for integrin αVβ3 and mediates VSMC phosphorylation of focal adhesion kinase, migration, and proliferation in vitro. MFAP4-dependent VSMC activation was reversible by treatment with MFAP4-blocking antibodies and inhibitors of focal adhesion kinase and downstream kinases. In addition, we showed that MFAP4 promotes monocyte chemotaxis in integrin αVβ3-dependent manner.
  
  
  CONCLUSIONS
  MFAP4 regulates integrin αVβ3-induced VSMC proliferation and migration, as well as monocyte chemotaxis, and accelerates neointimal hyperplasia after vascular injury.
authors.affiliation: From the Institute of Molecular Medicine, University of Southern Denmark, Odense, Denmark (A.S., B.P., L.E.H., K.K., G.B.K., H.W.-J., J.B.M., K.K.-M., L.K.D., P.B.L.H., J.S., U.H, G.L.S.); Department of Molecular Biology and Genetics, Aarhus University, Aarhus, Denmark (E.-M.F.); Department of Pathology, Odense University Hospital, Odense, Denmark (O.N.); Institute of Functional and Applied Anatomy, Hannover Medical School, Hannover, Germany (C.W., J.H., M.O.); Biomedical Research in Endstage and Obstructive Lung Disease Hannover (BREATH), Member of the German Center for Lung Research (DZL), Hannover, Germany (C.W., J.H., M.O.); REBIRTH Cluster of Excellence, Hannover, Germany (C.W., J.H., M.O.); German Mouse Clinic, Institute of Experimental Genetics, Helmholtz Zentrum München, German Research Center for Environmental Health, Munich, Germany (B.R., A.S., V.G.-D., H.F., M.H.d.A.); Institute of Molecular Animal Breeding and Biotechnology, Gene Center, Ludwig-Maximilians-University München, Munich, Germany (B.R, E.W.); Division of Cardiology, Department of Medicine III, University of Heidelberg, Heidelberg, Germany (A.S., R.B.); Chair of Experimental Genetics, Center of Life and Food Sciences Weihenstephan, Technische Universität München, Freising-Weihenstephan, Munich, Germany (M.H.d.A.); Cardiovascular Research Unit, Viborg Hospital, Viborg, Denmark (J.S.L.); and Department of Cardiothoracic and Vascular Surgery, Center of Individualized Medicine in Arterial Diseases (CIMA), Odense University Hospital, Odense, Denmark (J.S.L.).
authors.forename: Anders
authors.initials: A
authors.lastname: Schlosser
authors.affiliation: From the Institute of Molecular Medicine, University of Southern Denmark, Odense, Denmark (A.S., B.P., L.E.H., K.K., G.B.K., H.W.-J., J.B.M., K.K.-M., L.K.D., P.B.L.H., J.S., U.H, G.L.S.); Department of Molecular Biology and Genetics, Aarhus University, Aarhus, Denmark (E.-M.F.); Department of Pathology, Odense University Hospital, Odense, Denmark (O.N.); Institute of Functional and Applied Anatomy, Hannover Medical School, Hannover, Germany (C.W., J.H., M.O.); Biomedical Research in Endstage and Obstructive Lung Disease Hannover (BREATH), Member of the German Center for Lung Research (DZL), Hannover, Germany (C.W., J.H., M.O.); REBIRTH Cluster of Excellence, Hannover, Germany (C.W., J.H., M.O.); German Mouse Clinic, Institute of Experimental Genetics, Helmholtz Zentrum München, German Research Center for Environmental Health, Munich, Germany (B.R., A.S., V.G.-D., H.F., M.H.d.A.); Institute of Molecular Animal Breeding and Biotechnology, Gene Center, Ludwig-Maximilians-University München, Munich, Germany (B.R, E.W.); Division of Cardiology, Department of Medicine III, University of Heidelberg, Heidelberg, Germany (A.S., R.B.); Chair of Experimental Genetics, Center of Life and Food Sciences Weihenstephan, Technische Universität München, Freising-Weihenstephan, Munich, Germany (M.H.d.A.); Cardiovascular Research Unit, Viborg Hospital, Viborg, Denmark (J.S.L.); and Department of Cardiothoracic and Vascular Surgery, Center of Individualized Medicine in Arterial Diseases (CIMA), Odense University Hospital, Odense, Denmark (J.S.L.).
authors.forename: Bartosz
authors.initials: B
authors.lastname: Pilecki
authors.forename: Line E
authors.initials: LE
authors.lastname: Hemstra
authors.forename: Karin
authors.initials: K
authors.lastname: Kejling
authors.forename: Gudlaug B
authors.initials: GB
authors.lastname: Kristmannsdottir
authors.forename: Helle
authors.initials: H
authors.lastname: Wulf-Johansson
authors.forename: Jesper B
authors.initials: JB
authors.lastname: Moeller
authors.forename: Ernst-Martin
authors.initials: EM
authors.lastname: Füchtbauer
authors.forename: Ole
authors.initials: O
authors.lastname: Nielsen
authors.forename: Katrine
authors.initials: K
authors.lastname: Kirketerp-Møller
authors.forename: Lalit K
authors.initials: LK
authors.lastname: Dubey
authors.forename: Pernille B L
authors.initials: PB
authors.lastname: Hansen
authors.forename: Jane
authors.initials: J
authors.lastname: Stubbe
authors.forename: Christoph
authors.initials: C
authors.lastname: Wrede
authors.forename: Jan
authors.initials: J
authors.lastname: Hegermann
authors.forename: Matthias
authors.initials: M
authors.lastname: Ochs
authors.forename: Birgit
authors.initials: B
authors.lastname: Rathkolb
authors.forename: Anja
authors.initials: A
authors.lastname: Schrewe
authors.forename: Raffi
authors.initials: R
authors.lastname: Bekeredjian
authors.forename: Eckhard
authors.initials: E
authors.lastname: Wolf
authors.forename: Valérie
authors.initials: V
authors.lastname: Gailus-Durner
authors.forename: Helmut
authors.initials: H
authors.lastname: Fuchs
authors.forename: Martin
authors.initials: M
authors.lastname: Hrabě de Angelis
authors.forename: Jes S
authors.initials: JS
authors.lastname: Lindholt
authors.forename: Uffe
authors.initials: U
authors.lastname: Holmskov
authors.forename: Grith L
authors.initials: GL
authors.lastname: Sorensen
mesh_terms: D000818:Animals; D017209:Apoptosis; D002339:Carotid Arteries / Q000378:metabolism / Q000473:pathology; D002340:Carotid Artery Diseases / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D002634:Chemotaxis, Leukocyte; D004195:Disease Models, Animal; D016326:Extracellular Matrix Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D051417:Focal Adhesion Kinase 1 / Q000037:antagonists & inhibitors / Q000378:metabolism; D005838:Genotype; D006023:Glycoproteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D006965:Hyperplasia; D039302:Integrin alphaVbeta3 / Q000378:metabolism; D008024:Ligands; D008297:Male; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D009000:Monocytes / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D058426:Neointima; D010641:Phenotype; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D015398:Signal Transduction; D013997:Time Factors; D066253:Vascular Remodeling
chemical_list: D002352:Carrier Proteins; D016326:Extracellular Matrix Proteins; D006023:Glycoproteins; D039302:Integrin alphaVbeta3; D008024:Ligands; C400752:MFAP4 protein, human; C000600072:MFAP4 protein, mouse; D047428:Protein Kinase Inhibitors; D051417:Focal Adhesion Kinase 1; C496051:Ptk2 protein, mouse
keywords: MFAP4 protein; carotid stenosis; extracellular matrix proteins; hyperplasia; integrin alphaVbeta3; mouse; muscle; smooth; vascular