NO mediates mural cell recruitment and vessel morphogenesis in murine melanomas and tissue-engineered blood vessels.
NO has been shown to mediate angiogenesis; however, its role in vessel morphogenesis and maturation is not known. Using intravital microscopy, histological analysis, alpha-smooth muscle actin and chondroitin sulfate proteoglycan 4 staining, microsensor NO measurements, and an NO synthase (NOS) inhibitor, we found that NO mediates mural cell coverage as well as vessel branching and longitudinal extension but not the circumferential growth of blood vessels in B16 murine melanomas. NO-sensitive fluorescent probe 4,5-diaminofluorescein imaging, NOS immunostaining, and the use of NOS-deficient mice revealed that eNOS in vascular endothelial cells is the predominant source of NO and induces these effects. To further dissect the role of NO in mural cell recruitment and vascular morphogenesis, we performed a series of independent analyses. Transwell and under-agarose migration assays demonstrated that endothelial cell-derived NO induces directional migration of mural cell precursors toward endothelial cells. An in vivo tissue-engineered blood vessel model revealed that NO mediates endothelial-mural cell interaction prior to vessel perfusion and also induces recruitment of mural cells to angiogenic vessels, vessel branching, and longitudinal extension and subsequent stabilization of the vessels. These data indicate that endothelial cell-derived NO induces mural cell recruitment as well as subsequent morphogenesis and stabilization of angiogenic vessels.
Kashiwagi
Satoshi
S
Edwin L. Steele Laboratory, Department of Radiation Oncology, and Cardiovascular Research Center, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA.
Izumi
Yotaro
Y
Gohongi
Takeshi
T
Demou
Zoe N
ZN
Xu
Lei
L
Huang
Paul L
PL
Buerk
Donald G
DG
Munn
Lance L
LL
Jain
Rakesh K
RK
Fukumura
Dai
D
D000818:Animals; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000473:pathology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015870:Gene Expression; D006801:Humans; D008546:Melanoma, Experimental / Q000098:blood supply* / Q000188:drug therapy / Q000473:pathology; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D016513:Mice, SCID; D009389:Neovascularization, Pathologic / Q000188:drug therapy; D018919:Neovascularization, Physiologic; D009569:Nitric Oxide / Q000502:physiology*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors / Q000172:deficiency / Q000235:genetics; D023822:Tissue Engineering; D019323:omega-N-Methylarginine / Q000494:pharmacology
D004791:Enzyme Inhibitors; D019323:omega-N-Methylarginine; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase