title: SDF-1/CXCR4 axis in Tie2-lineage cells including endothelial progenitor cells contributes to bone fracture healing.
abstract: CXC chemokine receptor 4 (CXCR4) is a specific receptor for stromal-derived-factor 1 (SDF-1). SDF-1/CXCR4 interaction is reported to play an important role in vascular development. On the other hand, the therapeutic potential of endothelial progenitor cells (EPCs) in fracture healing has been demonstrated with mechanistic insight of vasculogenesis/angiogenesis and osteogenesis enhancement at sites of fracture. The purpose of this study was to investigate the influence of the SDF-1/CXCR4 pathway in Tie2-lineage cells (including EPCs) in bone formation. We created CXCR4 gene conditional knockout mice using the Cre/loxP system and set two groups of mice: Tie2-Cre(ER) CXCR4 knockout mice (CXCR4(-/-) ) and wild-type mice (WT). We report here that in vitro, EPCs derived from of CXCR4(-/-) mouse bone marrow demonstrated severe reduction of migration activity and EPC colony-forming activity when compared with those derived from WT mouse bone marrow. In vivo, radiological and morphological examinations showed fracture healing delayed in the CXCR4(-/-) group and the relative callus area at weeks 2 and 3 was significantly smaller in CXCR4(-/-) group mice. Quantitative analysis of capillary density at perifracture sites also showed a significant decrease in the CXCR4(-/-) group. Especially, CXCR4(-/-) group mice demonstrated significant early reduction of blood flow recovery at fracture sites compared with the WT group in laser Doppler perfusion imaging analysis. Real-time RT-PCR analysis showed that the gene expressions of angiogenic markers (CD31, VE-cadherin, vascular endothelial growth factor [VEGF]) and osteogenic markers (osteocalcin, collagen 1A1, bone morphogenetic protein 2 [BMP2]) were lower in the CXCR4(-/-) group. In the gain-of-function study, the fracture in the SDF-1 intraperitoneally injected WT group healed significantly faster with enough callus formation compared with the SDF-1 injected CXCR4(-/-) group. We demonstrated that an EPC SDF-1/CXCR4 axis plays an important role in bone fracture healing using Tie2-Cre(ER) CXCR4 conditional knockout mice.
authors.affiliation: Group of Vascular Regeneration, Institute of Biomedical Research and Innovation, Kobe, Japan; Department of Orthopaedic Surgery, Kobe University Graduate School of Medicine, Kobe, Japan.
authors.forename: Yohei
authors.initials: Y
authors.lastname: Kawakami
authors.forename: Masaaki
authors.initials: M
authors.lastname: Ii
authors.forename: Tomoyuki
authors.initials: T
authors.lastname: Matsumoto
authors.forename: Ryosuke
authors.initials: R
authors.lastname: Kuroda
authors.forename: Tomoya
authors.initials: T
authors.lastname: Kuroda
authors.forename: Sang-Mo
authors.initials: SM
authors.lastname: Kwon
authors.forename: Atsuhiko
authors.initials: A
authors.lastname: Kawamoto
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Akimaru
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Mifune
authors.forename: Taro
authors.initials: T
authors.lastname: Shoji
authors.forename: Tomoaki
authors.initials: T
authors.lastname: Fukui
authors.forename: Masahiro
authors.initials: M
authors.lastname: Kurosaka
authors.forename: Takayuki
authors.initials: T
authors.lastname: Asahara
mesh_terms: D000818:Animals; D000943:Antigens, Differentiation / Q000096:biosynthesis / Q000235:genetics; D001861:Bone Regeneration; D002478:Cells, Cultured; D054377:Chemokine CXCL12 / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D066026:Endothelial Progenitor Cells / Q000378:metabolism* / Q000473:pathology; D050723:Fractures, Bone / Q000178:diet therapy / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D051379:Mice; D018345:Mice, Knockout; D042787:Receptor, TIE-2 / Q000235:genetics / Q000378:metabolism*; D019718:Receptors, CXCR4 / Q000235:genetics / Q000378:metabolism*
chemical_list: D000943:Antigens, Differentiation; C514936:CXCR4 protein, mouse; D054377:Chemokine CXCL12; C515256:Cxcl12 protein, mouse; D019718:Receptors, CXCR4; D042787:Receptor, TIE-2; C079086:Tek protein, mouse
keywords: CX CHEMOKINE RECEPTOR 4 (CXCR4); ENDOTHELIAL PROGENITOR CELL (EPC); FRACTURE HEALING; KNOCKOUT MICE; STROMAL-DERIVED FACTOR-1 (SDF-1)