Identification of a 'snapshot' of co-expressed angiogenic markers in laser-dissected vessels from unstable carotid plaques with targeted arrays.
OBJECTIVE
Angiogenesis is a feature of the atherogenic process, with intimal neovascularisation arising from vessels in the adventitia, adjacent to a plaque. Immature, leaky blood vessels from unstable plaques proliferate abnormally and, being poorly invested with smooth muscle cells, may contribute to instability of the plaque by facilitation of inflammatory cell infiltration and haemorrhagic complications.


METHODS
We used laser-capture microdissection to isolate angiogenic areas of the extracellular matrix (containing CD105/flt-1-positive, fragile thin-walled vessels) and non-angiogenic vascular areas (CD105-negative, with smooth muscle cell covering) of complicated endarterectomy plaques, and specifically designed angiogenesis-TaqMan real-time PCR microarrays to identify gene expression.


RESULTS
Important pro-angiogenic components, including Notch-3, delta-like-4 (DLL4), Tie-2, angiopoietin-1 (Angio-1) and receptor for advanced glycation end products (RAGE), and one anti-angiogenic factor, endostatin, were up-regulated in these regions. Immunohistochemistry demonstrated localisation within intimal, active (CD105-positive) microvessels and co-localisation of Notch-3 and DLL4/Tie-2 and Angio-1 in the same vessels indicating multiple/synergistic signalling mechanisms associated with vessel development.


CONCLUSIONS
These data, although providing only a snapshot of information, demonstrate that plaque vascularisation occurs in the presence of multiple angiogenically active factors. Knowledge of their combined effects could help in the formulation of novel therapeutics designed to stabilise or prevent their formation in the treatment of atherosclerosis.
Slevin
M
M
School of Biology, Chemistry and Health Science, Manchester Metropolitan University, Manchester M1 5GD, UK. .a.slevin@mmu.ac.uk
Turu
M M
MM
Rovira
N
N
Luque
A
A
Baldellou
M
M
Krupinski
J
J
Badimon
L
L
D048868:Adaptor Proteins, Signal Transducing; D000368:Aged; D042501:Angiogenic Proteins / Q000032:analysis / Q000235:genetics*; D042683:Angiopoietin-1 / Q000235:genetics; D015703:Antigens, CD / Q000032:analysis; D002135:Calcium-Binding Proteins; D002339:Carotid Arteries / Q000737:chemistry / Q000276:immunology / Q000601:surgery; D016893:Carotid Stenosis / Q000235:genetics* / Q000276:immunology / Q000503:physiopathology / Q000601:surgery; D004210:Dissection / Q000295:instrumentation*; D016894:Endarterectomy, Carotid; D000071063:Endoglin; D043169:Endostatins / Q000235:genetics; D020869:Gene Expression Profiling / Q000379:methods*; D005819:Genetic Markers; D006801:Humans; D007150:Immunohistochemistry; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics; D007834:Lasers; D008297:Male; D008875:Middle Aged; D018919:Neovascularization, Physiologic / Q000235:genetics*; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction; D000067759:Receptor for Advanced Glycation End Products; D000071656:Receptor, Notch3; D042787:Receptor, TIE-2 / Q000235:genetics; D011956:Receptors, Cell Surface / Q000032:analysis; D011971:Receptors, Immunologic / Q000235:genetics; D051880:Receptors, Notch / Q000235:genetics; D012421:Rupture; D040281:Vascular Endothelial Growth Factor Receptor-1 / Q000235:genetics
C467630:ANGPT1 protein, human; D048868:Adaptor Proteins, Signal Transducing; D042501:Angiogenic Proteins; D042683:Angiopoietin-1; D015703:Antigens, CD; D002135:Calcium-Binding Proteins; C424126:DLL4 protein, human; C064206:ENG protein, human; D000071063:Endoglin; D043169:Endostatins; D005819:Genetic Markers; D036341:Intercellular Signaling Peptides and Proteins; C499374:NOTCH3 protein, human; D000067759:Receptor for Advanced Glycation End Products; D000071656:Receptor, Notch3; D011956:Receptors, Cell Surface; D011971:Receptors, Immunologic; D051880:Receptors, Notch; C501162:FLT1 protein, human; D042787:Receptor, TIE-2; D040281:Vascular Endothelial Growth Factor Receptor-1