Heidi Noels1, Baixue Zhou2, Pathricia V Tilstam2, Wendy Theelen2, Xiaofeng Li2, Lukas Pawig2, Corinna Schmitz2, Shamima Akhtar2, Sakine Simsekyilmaz2, Erdenechimeg Shagdarsuren2, Andreas Schober2, Ralf H Adams2, Jürgen Bernhagen2, Elisa A Liehn2, Yvonne Döring2, Christian Weber1. 1. From the Institute for Molecular Cardiovascular Research (H.N., B.Z., P.V.T., W.T., X.L., L.P., S.A., S.S., E.S., E.A.L.) and Institute of Biochemistry and Molecular Cell Biology (C.S., J.B.), University Hospital Aachen, Rheinisch-Westfälische Technische Hochschule (RWTH) Aachen University, Aachen, Germany; Institute for Cardiovascular Prevention (A.S., Y.D., C.W.) and August-Lenz-Stiftung, Institute for Cardiovascular Research (J.B.), Ludwig-Maximilians-University Munich, Munich, Germany; Max Planck Institute for Molecular Biomedicine, University of Münster, Münster, Germany (R.H.A.); Cardiovascular Research Institute Maastricht, Maastricht University, Maastricht, The Netherlands (C.W.); and German Centre for Cardiovascular Research (Deutsches Zentrum für Herz-Kreislauf-Forschung), partner site Munich Heart Alliance, Munich, Germany (C.W.). kreislaufinstitut@med.uni-muenchen.de hnoels@ukaachen.de. 2. From the Institute for Molecular Cardiovascular Research (H.N., B.Z., P.V.T., W.T., X.L., L.P., S.A., S.S., E.S., E.A.L.) and Institute of Biochemistry and Molecular Cell Biology (C.S., J.B.), University Hospital Aachen, Rheinisch-Westfälische Technische Hochschule (RWTH) Aachen University, Aachen, Germany; Institute for Cardiovascular Prevention (A.S., Y.D., C.W.) and August-Lenz-Stiftung, Institute for Cardiovascular Research (J.B.), Ludwig-Maximilians-University Munich, Munich, Germany; Max Planck Institute for Molecular Biomedicine, University of Münster, Münster, Germany (R.H.A.); Cardiovascular Research Institute Maastricht, Maastricht University, Maastricht, The Netherlands (C.W.); and German Centre for Cardiovascular Research (Deutsches Zentrum für Herz-Kreislauf-Forschung), partner site Munich Heart Alliance, Munich, Germany (C.W.).
Abstract
OBJECTIVE: The Cxcl12/Cxcr4 chemokine ligand/receptor axis mediates the mobilization of smooth muscle cell progenitors, driving injury-induced neointimal hyperplasia. This study aimed to investigate the role of endothelial Cxcr4 in neointima formation. APPROACH AND RESULTS: β-Galactosidase staining using bone marrow x kinase (Bmx)-CreER(T2) reporter mice and double immunofluorescence revealed an efficient and endothelial-specific deletion of Cxcr4 in Bmx-CreER(T2+) compared with Bmx-CreER(T2-) Cxcr4-floxed apolipoprotein E-deficient (Apoe(-/-)) mice (referred to as Cxcr4(EC-KO)ApoE(-/-) and Cxcr4(EC-WT) ApoE(-/-), respectively). Endothelial Cxcr4 deficiency significantly increased wire injury-induced neointima formation in carotid arteries from Cxcr4(EC-KO)ApoE(-/-) mice. The lesions displayed a higher number of macrophages, whereas the smooth muscle cell and collagen content were reduced. This was associated with a significant reduction in reendothelialization and endothelial cell proliferation in injured Cxcr4(EC-KO)ApoE(-/-) carotids compared with Cxcr4(EC-WT)ApoE(-/-) controls. Furthermore, stimulation of human aortic endothelial cells with chemokine (C-X-C motif) ligand 12 (CXCL12) significantly enhanced their wound-healing capacity in an in vitro scratch assay, an effect that could be reversed with the CXCR4 antagonist AMD3100. Also, flow cytometric analysis showed a reduced mobilization of Sca1(+)Flk1(+)Cd31(+) and of Lin(-)Sca1(+) progenitors in Cxcr4(EC-KO) ApoE(-/-) mice after vascular injury, although Cxcr4 surface expression was unaltered. No differences could be detected in plasma concentrations of Cxcl12, vascular endothelial growth factor, sphingosine 1-phosphate, or Flt3 (fms-related tyrosine kinase 3) ligand, all cytokines with an established role in progenitor cell mobilization. Nonetheless, double immunofluorescence revealed a significant reduction in local endothelial Cxcl12 staining in injured carotids from Cxcr4(EC-KO)ApoE(-/-) mice. CONCLUSIONS: Endothelial Cxcr4 is crucial for efficient reendothelialization after vascular injury through endothelial wound healing and proliferation, and through the mobilization of Sca1(+)Flk1(+)Cd31(+) cells, often referred to as circulating endothelial progenitor cells.
OBJECTIVE: The Cxcl12/Cxcr4 chemokine ligand/receptor axis mediates the mobilization of smooth muscle cell progenitors, driving injury-induced neointimal hyperplasia. This study aimed to investigate the role of endothelial Cxcr4 in neointima formation. APPROACH AND RESULTS: β-Galactosidase staining using bone marrow x kinase (Bmx)-CreER(T2) reporter mice and double immunofluorescence revealed an efficient and endothelial-specific deletion of Cxcr4 in Bmx-CreER(T2+) compared with Bmx-CreER(T2-) Cxcr4-floxed apolipoprotein E-deficient (Apoe(-/-)) mice (referred to as Cxcr4(EC-KO)ApoE(-/-) and Cxcr4(EC-WT) ApoE(-/-), respectively). Endothelial Cxcr4 deficiency significantly increased wire injury-induced neointima formation in carotid arteries from Cxcr4(EC-KO)ApoE(-/-) mice. The lesions displayed a higher number of macrophages, whereas the smooth muscle cell and collagen content were reduced. This was associated with a significant reduction in reendothelialization and endothelial cell proliferation in injured Cxcr4(EC-KO)ApoE(-/-) carotids compared with Cxcr4(EC-WT)ApoE(-/-) controls. Furthermore, stimulation of human aortic endothelial cells with chemokine (C-X-C motif) ligand 12 (CXCL12) significantly enhanced their wound-healing capacity in an in vitro scratch assay, an effect that could be reversed with the CXCR4 antagonist AMD3100. Also, flow cytometric analysis showed a reduced mobilization of Sca1(+)Flk1(+)Cd31(+) and of Lin(-)Sca1(+) progenitors in Cxcr4(EC-KO) ApoE(-/-) mice after vascular injury, although Cxcr4 surface expression was unaltered. No differences could be detected in plasma concentrations of Cxcl12, vascular endothelial growth factor, sphingosine 1-phosphate, or Flt3 (fms-related tyrosine kinase 3) ligand, all cytokines with an established role in progenitor cell mobilization. Nonetheless, double immunofluorescence revealed a significant reduction in local endothelial Cxcl12 staining in injured carotids from Cxcr4(EC-KO)ApoE(-/-) mice. CONCLUSIONS: Endothelial Cxcr4 is crucial for efficient reendothelialization after vascular injury through endothelial wound healing and proliferation, and through the mobilization of Sca1(+)Flk1(+)Cd31(+) cells, often referred to as circulating endothelial progenitor cells.
Authors: Yvonne Döring; Heidi Noels; Emiel P C van der Vorst; Carlos Neideck; Virginia Egea; Maik Drechsler; Manuela Mandl; Lukas Pawig; Yvonne Jansen; Katrin Schröder; Kiril Bidzhekov; Remco T A Megens; Wendy Theelen; Barbara M Klinkhammer; Peter Boor; Leon Schurgers; Rick van Gorp; Christian Ries; Pascal J H Kusters; Allard van der Wal; Tilman M Hackeng; Gabor Gäbel; Ralf P Brandes; Oliver Soehnlein; Esther Lutgens; Dietmar Vestweber; Daniel Teupser; Lesca M Holdt; Daniel J Rader; Danish Saleheen; Christian Weber Journal: Circulation Date: 2017-04-27 Impact factor: 29.690
Authors: Thomas Rathjen; Britta Kunkemoeller; Carly T Cederquist; Xuanchun Wang; Sam M Lockhart; James C Patti; Hanni Willenbrock; Grith Skytte Olsen; Gro Klitgaard Povlsen; Hans Christian Beck; Lars Melholt Rasmussen; Qian Li; Kyoungmin Park; George L King; Christian Rask-Madsen Journal: Arterioscler Thromb Vasc Biol Date: 2022-06-02 Impact factor: 10.514
Authors: Tanja Holopainen; Markus Räsänen; Andrey Anisimov; Tomi Tuomainen; Wei Zheng; Denis Tvorogov; Juha J Hulmi; Leif C Andersson; Bruno Cenni; Pasi Tavi; Eero Mervaala; Riikka Kivelä; Kari Alitalo Journal: Proc Natl Acad Sci U S A Date: 2015-10-01 Impact factor: 11.205