Inhibition of delta-like-4-mediated signaling impairs reparative angiogenesis after ischemia.
BACKGROUND
Notch signaling regulates vascular development. However, the implication of the Notch ligand Delta-like 4 (Dll4) in postischemic angiogenesis remains unclear.


OBJECTIVE
We investigated the role of Dll4/Notch signaling in reparative angiogenesis using a mouse model of ischemia.


RESULTS
We found Dll4 weakly expressed in microvascular endothelial cells of normoperfused muscles. Conversely, Dll4 is upregulated following ischemia and localized at the forefront of sprouting capillaries. We analyzed the effect of inhibiting endogenous Dll4 by intramuscular injection of an adenovirus encoding the soluble form of Dll4 extracellular domain (Ad-sDll4). Dll4 inhibition caused the formation of a disorganized, low-perfused capillary network in ischemic muscles. This structural abnormality was associated to delayed blood flow recovery and muscle hypoxia and degeneration. Analysis of microvasculature at early stages of repair revealed that Dll4 inhibition enhances capillary sprouting in a chaotic fashion and causes excessive leukocyte infiltration of ischemic muscles. Furthermore, Dll4 inhibition potentiated the elevation of the leukocyte chemoattractant CXCL1 (chemokine [C-X-C motif] ligand 1) following ischemia, without altering peripheral blood levels of stromal cell-derived factor-1 and monocyte chemoattractant protein-1. In cultured human monocytes, Dll4 induces the transcription of Notch target gene Hes-1 and inhibits the basal and tumor necrosis factor-alpha-stimulated production of interleukin-8, the human functional homolog of murine CXCL1. The inhibitory effect of Dll4 on interleukin-8 was abolished by DAPT, a Notch inhibitor, or by coculturing activated human monocytes with Ad-sDll4-infected endothelial cells.


CONCLUSIONS
Dll4/Notch interaction is essential for proper reparative angiogenesis. Moreover, Dll4/Notch signaling regulates sprouting angiogenesis and coordinates the interaction between inflammation and angiogenesis under ischemic conditions.
Experimental Cardiovascular Medicine Division, Bristol Heart Institute,University of Bristol, Bristol BS28HW, United Kingdom.
Ayman
A
Al Haj Zen
Atsuhiko
A
Oikawa
Miriam
M
Bazan-Peregrino
Marco
M
Meloni
Costanza
C
Emanueli
Paolo
P
Madeddu
D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D002135:Calcium-Binding Proteins; D002478:Cells, Cultured; D054360:Chemokine CXCL1 / Q000378:metabolism; D002634:Chemotaxis, Leukocyte; D018920:Coculture Techniques; D004195:Disease Models, Animal; D042783:Endothelial Cells / Q000378:metabolism*; D006614:Hindlimb; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D016209:Interleukin-8 / Q000378:metabolism; D007511:Ischemia / Q000000981:diagnostic imaging / Q000235:genetics / Q000378:metabolism* / Q000503:physiopathology; D017078:Laser-Doppler Flowmetry; D007962:Leukocytes / Q000378:metabolism; D008297:Male; D051379:Mice; D018482:Muscle, Skeletal / Q000098:blood supply*; D018919:Neovascularization, Physiologic / Q000235:genetics; D051880:Receptors, Notch / Q000378:metabolism*; D012038:Regeneration; D012039:Regional Blood Flow; D015398:Signal Transduction / Q000235:genetics; D013997:Time Factors; D014162:Transfection; D014463:Ultrasonography
D048868:Adaptor Proteins, Signal Transducing; C475829:CXCL1 protein, human; C508606:CXCL8 protein, human; D002135:Calcium-Binding Proteins; D054360:Chemokine CXCL1; C475895:Cxcl1 protein, mouse; C424126:DLL4 protein, human; D036341:Intercellular Signaling Peptides and Proteins; D016209:Interleukin-8; D051880:Receptors, Notch