title: A key role for the integrin alpha2beta1 in experimental and developmental angiogenesis.
abstract: The alpha2beta1 integrin receptor plays a key role in angiogenesis. Here we investigated the effects of small molecule inhibitors (SMIs) designed to disrupt integrin alpha2 I or beta1 I-like domain function on angiogenesis. In unchallenged endothelial cells, fibrillar collagen induced robust capillary morphogenesis. In contrast, tube formation was significantly reduced by SMI496, a beta1 I-like domain inhibitor and by function-blocking anti-alpha2beta1 but not -alpha1beta1 antibodies. Endothelial cells bound fluorescein-labeled collagen I fibrils, an interaction specifically inhibited by SMI496. Moreover, SMI496 caused cell retraction and cytoskeletal collapse of endothelial cells as well as delayed endothelial cell wound healing. SMI activities were examined in vivo by supplementing the growth medium of zebrafish embryos expressing green fluorescent protein under the control of the vascular endothelial growth factor receptor-2 promoter. SMI496, but not a control compound, interfered with angiogenesis in vivo by reversibly inhibiting sprouting from the axial vessels. We further characterized zebrafish alpha2 integrin and discovered that this integrin is highly conserved, especially the I domain. Notably, a similar vascular phenotype was induced by morpholino-mediated knockdown of the integrin alpha2 subunit. By live videomicroscopy, we confirmed that the vessels were largely nonfunctional in the absence of alpha2beta1 integrin. Collectively, our results provide strong biochemical and genetic evidence of a central role for alpha2beta1 integrin in experimental and developmental angiogenesis.
authors.affiliation: Department of Pathology, Anatomy and Cell Biology, 1020 Locust Street, Thomas Jefferson University, Philadelphia, PA 19107, USA.
authors.forename: James D
authors.initials: JD
authors.lastname: San Antonio
authors.forename: Jason J
authors.initials: JJ
authors.lastname: Zoeller
authors.forename: Kari
authors.initials: K
authors.lastname: Habursky
authors.forename: Kevin
authors.initials: K
authors.lastname: Turner
authors.forename: Wittaya
authors.initials: W
authors.lastname: Pimtong
authors.forename: Michelle
authors.initials: M
authors.lastname: Burrows
authors.forename: Sungwook
authors.initials: S
authors.lastname: Choi
authors.forename: Sandeep
authors.initials: S
authors.lastname: Basra
authors.forename: Joel S
authors.initials: JS
authors.lastname: Bennett
authors.forename: William F
authors.initials: WF
authors.lastname: DeGrado
authors.forename: Renato V
authors.initials: RV
authors.lastname: Iozzo
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D019138:Antibodies, Blocking; D002448:Cell Adhesion / Q000502:physiology; D002465:Cell Movement / Q000502:physiology; D002478:Cells, Cultured; D003094:Collagen; D004151:Dipeptides / Q000494:pharmacology*; D004730:Endothelium, Vascular / Q000166:cytology / Q000502:physiology; D006801:Humans; D038982:Integrin alpha2beta1 / Q000037:antagonists & inhibitors / Q000502:physiology*; D018919:Neovascularization, Physiologic / Q000502:physiology*; D017434:Protein Structure, Tertiary; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000502:physiology; D015027:Zebrafish
chemical_list: D019138:Antibodies, Blocking; D004151:Dipeptides; D038982:Integrin alpha2beta1; C545492:SMI496; D003094:Collagen; D040301:Vascular Endothelial Growth Factor Receptor-2
references.citation: Proc Natl Acad Sci U S A. 1997 Dec 9;94(25):13612-7
references.pmid: 9391074
references.citation: PLoS Genet. 2007 May 25;3(5):e78
references.pmid: 17530925
references.citation: Microsc Res Tech. 2008 May;71(5):339-48
references.pmid: 18300285
references.citation: J Med Chem. 2007 Nov 1;50(22):5457-62
references.pmid: 17915848
references.citation: Cell. 2000 Mar 31;101(1):47-56
references.pmid: 10778855
references.citation: Matrix Biol. 2002 Nov;21(7):547-58
references.pmid: 12475639
references.citation: J Med Chem. 2007 May 31;50(11):2742-6
references.pmid: 17447751
references.citation: Development. 2003 Nov;130(21):5281-90
references.pmid: 12954720
references.citation: Trends Cell Biol. 2005 Jan;15(1):52-60
references.pmid: 15653078
references.citation: Exp Cell Res. 1996 Apr 10;224(1):39-51
references.pmid: 8612690
references.citation: Biochemistry. 2000 Aug 15;39(32):9859-67
references.pmid: 10933804
references.citation: Science. 2000 Aug 18;289(5482):1197-202
references.pmid: 10947988
references.citation: Cancer Cell. 2002 Apr;1(3):257-67
references.pmid: 12086862
references.citation: J Invest Dermatol. 2007 Feb;127(2):467-78
references.pmid: 16977325
references.citation: Proc Natl Acad Sci U S A. 2000 Nov 21;97(24):12965-9
references.pmid: 11087852
references.citation: J Mol Biol. 2004 Jan 23;335(4):1019-28
references.pmid: 14698296
references.citation: Curr Pharm Des. 2005;11(7):867-80
references.pmid: 15777240
references.citation: J Biol Chem. 2005 Nov 4;280(44):36873-82
references.pmid: 16103112
references.citation: Curr Opin Hematol. 2008 May;15(3):197-203
references.pmid: 18391785
references.citation: Semin Oncol. 2002 Dec;29(6 Suppl 16):15-8
references.pmid: 12516034
references.citation: Bioorg Med Chem Lett. 2006 Jul 1;16(13):3380-2
references.pmid: 16678410
references.citation: Nature. 2007 Feb 15;445(7129):781-4
references.pmid: 17259972
references.citation: Am J Pathol. 2002 Jan;160(1):195-204
references.pmid: 11786413
references.citation: Proteome Sci. 2008 Feb 12;6:7
references.pmid: 18269764
references.citation: Nat Biotechnol. 2004 May;22(5):595-9
references.pmid: 15097998
references.citation: Curr Biol. 2001 Oct 2;11(19):1481-91
references.pmid: 11591315
references.citation: BMC Cell Biol. 2006 Jun 20;7:24
references.pmid: 16787535
references.citation: Am J Pathol. 2002 Jul;161(1):337-44
references.pmid: 12107118
references.citation: Development. 2002 Feb;129(4):973-82
references.pmid: 11861480
references.citation: Circ Res. 2005 Nov 25;97(11):1093-107
references.pmid: 16306453
references.citation: Biochem J. 2003 Nov 15;376(Pt 1):77-85
references.pmid: 12871211
references.citation: J Biol Chem. 2003 Aug 15;278(33):30516-24
references.pmid: 12788934
references.citation: Arterioscler Thromb Vasc Biol. 2003 May 1;23(5):911-2
references.pmid: 12740225
references.citation: J Biol Chem. 2008 Jan 25;283(4):2335-43
references.pmid: 18024432
references.citation: Blood. 2008 Apr 1;111(7):3562-70
references.pmid: 18198349
references.citation: J Natl Cancer Inst. 2006 Nov 15;98(22):1634-46
references.pmid: 17105986
references.citation: FEBS J. 2007 Mar;274(6):1481-91
references.pmid: 17489103
references.citation: J Biol Chem. 2003 Feb 7;278(6):4238-49
references.pmid: 12435733
references.citation: Proc Natl Acad Sci U S A. 2009 Jan 20;106(3):719-24
references.pmid: 19141632
references.citation: Nat Rev Mol Cell Biol. 2005 Aug;6(8):646-56
references.pmid: 16064139
references.citation: Dev Biol. 2001 Feb 15;230(2):278-301
references.pmid: 11161578
references.citation: J Cell Biol. 2008 Apr 21;181(2):381-94
references.pmid: 18426981
references.citation: Nat Genet. 2000 Oct;26(2):216-20
references.pmid: 11017081
references.citation: Blood. 2007 May 1;109(9):3745-8
references.pmid: 17197432
references.citation: J Biol Chem. 2002 Mar 29;277(13):10789-94
references.pmid: 11788609
references.citation: Development. 2006 Sep;133(18):3563-73
references.pmid: 16914496
references.citation: J Biol Chem. 1997 Nov 7;272(45):28512-7
references.pmid: 9353312
references.citation: Gene Expr Patterns. 2008 Feb;8(3):171-80
references.pmid: 18068546
references.citation: J Cell Biol. 2004 Jul 5;166(1):97-109
references.pmid: 15240572
references.citation: J Biol Chem. 2003 Jan 3;278(1):327-34
references.pmid: 12399469
references.citation: Exp Cell Res. 2008 Nov 15;314(19):3593-604
references.pmid: 18809396
references.citation: J Biol Chem. 2005 Feb 25;280(8):7080-7
references.pmid: 15591058
references.citation: Nat Rev Cancer. 2003 Jun;3(6):422-33
references.pmid: 12778132