Activation of fractalkine/CX3CR1 by vascular endothelial cells induces angiogenesis through VEGF-A/KDR and reverses hindlimb ischaemia.
OBJECTIVE
The present study investigated the detailed mechanism by which fractalkine (Fkn), a CX3C chemokine, induces angiogenesis and its functional implication in alleviating ischaemia in vivo.


RESULTS
Fkn induced new vessel formation on the excised rat aorta and chick chorioallantoic membrane (CAM) through CX3CR1 activation. Immunoblotting analysis, promoter assay and electrophoretic mobility shift assay showed that Fkn upregulated hypoxia-inducible factor-1 alpha (HIF-1alpha) by cultured human aortic endothelial cells (ECs), which in turn induced mRNA and protein levels of vascular endothelial growth factor (VEGF)-A through a p42/44 mitogen-activated protein kinase pathway. In vivo Fkn-induced angiogenesis on CAM was completely blocked by functional inhibition of VEGF receptor 2 kinase insert domain-containing receptor (KDR) and Rho GTPase. C57/BL6 mice with CX3CR1(-/-) bone marrow-derived cells developed angiogenesis in the implanted Fkn-mixed Matrigel plug, suggesting CX3CR1 activation in vascular ECs is sufficient for Fkn-induced angiogenesis in vivo. The condition of rat hindlimb ischaemia, which rapidly stimulated mRNA expression of both Fkn and VEGF-A, was significantly alleviated by the injection of whole-length Fkn protein.


CONCLUSIONS
Fkn-induced activation of CX3CR1 by ECs leads to in vivo angiogenesis through two sequential steps: the induction of HIF-1alpha and VEGF-A gene expression by CX3CR1 activation and the subsequent VEGF-A/KDR-induced angiogenesis. The potent induction of angiogenesis by Fkn can be used as a therapeutic strategy for alleviating peripheral ischaemia.
Ryu
Jewon
J
Division of Cardiology, Department of Internal Medicine, Asan Medical Center, University of Ulsan College of Medicine, 388-1 Pungnap-2 dong Songpa-gu 138-736, Seoul 138-736, Republic of Korea.
Lee
Cheol-Whan
CW
Hong
Kyung-Hee
KH
Shin
Jin-Ae
JA
Lim
Sun-Hee
SH
Park
Chan-Sik
CS
Shim
Jiyeon
J
Nam
Ki Byung
KB
Choi
Kee-Joon
KJ
Kim
You-Ho
YH
Han
Ki Hoon
KH
D042501:Angiogenic Proteins / Q000378:metabolism* / Q000494:pharmacology; D000818:Animals; D000073883:CX3C Chemokine Receptor 1; D002460:Cell Line; D002478:Cells, Cultured; D054428:Chemokine CX3CL1 / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D002642:Chick Embryo; D004195:Disease Models, Animal; D042783:Endothelial Cells / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D006614:Hindlimb; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000378:metabolism; D007511:Ischemia / Q000378:metabolism* / Q000503:physiopathology / Q000628:therapy; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D018482:Muscle, Skeletal / Q000098:blood supply*; D018919:Neovascularization, Physiologic / Q000187:drug effects; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D054387:Receptors, CXCR / Q000235:genetics / Q000378:metabolism*; D019707:Receptors, Chemokine / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D012039:Regional Blood Flow; D013997:Time Factors; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000378:metabolism*; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000378:metabolism*; D020741:rho GTP-Binding Proteins / Q000378:metabolism
D042501:Angiogenic Proteins; D000073883:CX3C Chemokine Receptor 1; C515538:CX3CL1 protein, human; C095235:CX3CR1 protein, human; C516305:CX3CR1 protein, rat; D054428:Chemokine CX3CL1; C515540:Cx3cl1 protein, rat; C471885:Cx3cr1 protein, mouse; C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D012333:RNA, Messenger; D054387:Receptors, CXCR; D019707:Receptors, Chemokine; D011994:Recombinant Proteins; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; C467498:vascular endothelial growth factor A, rat; D040301:Vascular Endothelial Growth Factor Receptor-2; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020741:rho GTP-Binding Proteins