Angiogenic sprouting and capillary lumen formation modeled by human umbilical vein endothelial cells (HUVEC) in fibrin gels: the role of fibroblasts and Angiopoietin-1☆

Nakatsu, Martin N.; Sainson, Richard C.A.; Aoto, Jason; Taylor, K. L.; Aitkenhead, Mark; Pérez‐del‐Pulgar, Sofía; Carpenter, Philip M.; Hughes, Christopher C.W.

doi:10.1016/s0026-2862(03)00045-1

Cited by 414 publications

(390 citation statements)

References 39 publications

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“…43 Moreover, HUVECs have been used to model sprout formation, which is an important step of angiogenesis seen in vivo. 44 LfcinB did not exhibit any cytotoxic activity against HUVECs, excluding the possibility that LfcinB simply caused HUVECs to undergo apoptosis, as occurs when various cancer cell lines are exposed to LfcinB. 24,27 Because both bovine lactoferrin and LfcinB are known to bind heparin, 29,30 our findings led us to hypothesize that LfcinB competed with bFGF and VEGF 165 for heparin-like binding sites on heparan sulfate proteoglycans on the surface of HUVECs.…”

Section: Discussionmentioning

confidence: 77%

Bovine Lactoferricin Inhibits Basic Fibroblast Growth Factor- and Vascular Endothelial Growth Factor165-Induced Angiogenesis by Competing for Heparin-Like Binding Sites on Endothelial Cells

Mader

Smyth

Marshall

et al. 2006

The American Journal of Pathology

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Section: Discussionmentioning

confidence: 77%

Bovine Lactoferricin Inhibits Basic Fibroblast Growth Factor- and Vascular Endothelial Growth Factor165-Induced Angiogenesis by Competing for Heparin-Like Binding Sites on Endothelial Cells

Mader

Smyth

Marshall

et al. 2006

The American Journal of Pathology

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“…A fibrin beads sprouting assay was conducted, as described previously, using a fibrin beads assay kit (Amersham‐Pharmacia Biotech), according the supplier's instructions 31. Briefly, HUVECs were incubated with the Cytodex 3 microcarrier beads (400 cells per bead; Sigma‐Aldrich) at 37°C overnight.…”

Section: Methodsmentioning

confidence: 99%

miR‐342‐5p Is a Notch Downstream Molecule and Regulates Multiple Angiogenic Pathways Including Notch, Vascular Endothelial Growth Factor and Transforming Growth Factor β Signaling

Yan

Cao

Liang

et al. 2016

JAHA

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BackgroundEndothelial cells (ECs) form blood vessels through angiogenesis that is regulated by coordination of vascular endothelial growth factor (VEGF), Notch, transforming growth factor β, and other signals, but the detailed molecular mechanisms remain unclear.Methods and ResultsSmall RNA sequencing initially identified miR‐342‐5p as a novel downstream molecule of Notch signaling in ECs. Reporter assay, quantitative reverse transcription polymerase chain reaction and Western blot analysis indicated that miR‐342‐5p targeted endoglin and modulated transforming growth factor β signaling by repressing SMAD1/5 phosphorylation in ECs. Transfection of miR‐342‐5p inhibited EC proliferation and lumen formation and reduced angiogenesis in vitro and in vivo, as assayed by using a fibrin beads–based sprouting assay, mouse aortic ring culture, and intravitreal injection of miR‐342‐5p agomir in P3 pups. Moreover, miR‐342‐5p promoted the migration of ECs, accompanied by reduced endothelial markers and increased mesenchymal markers, indicative of increased endothelial–mesenchymal transition. Transfection of endoglin at least partially reversed endothelial–mesenchymal transition induced by miR‐342‐5p. The expression of miR‐342‐5p was upregulated by transforming growth factor β, and inhibition of miR‐342‐5p attenuated the inhibitory effects of transforming growth factor β on lumen formation and sprouting by ECs. In addition, VEGF repressed miR‐342‐5p expression, and transfection of miR‐342‐5p repressed VEGFR2 and VEGFR3 expression and VEGF‐triggered Akt phosphorylation in ECs. miR‐342‐5p repressed angiogenesis in a laser‐induced choroidal neovascularization model in mice, highlighting its clinical potential.ConclusionsmiR‐342‐5p acts as a multifunctional angiogenic repressor mediating the effects and interaction among angiogenic pathways.

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“…Since the endothelial cells cover the entire surface of the bead, however, vessels may grow out in any direction; lumen-containing tubes must also be distinguished from migrating cells. The microbead assay has also been adapted to demonstrate angiogenic activity with HUVEC (Nakatsu et al, 2003), but HUVEC sprouting was only observed in the presence of a supporting layer of fibroblasts (Nakatsu et al, 2003). This is not surprising, given that HUVEC have a greater requirement for growth factors than do many other cell types, but presence of another cell type could complicate analysis of direct effects on endothelial cells.…”

Section: Assessment Of Sprouting Angiogenesis Using Endothelial Cell mentioning

confidence: 99%

In vitro assays of angiogenesis for assessment of angiogenic and anti-angiogenic agents

Goodwin¹

2007

Microvascular Research

259

208

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Blood vessels, either in insufficient numbers or in excess, contribute to the pathogenesis of many diseases. Agents that stimulate angiogenesis can improve blood flow in patients with ischemic diseases, whereas anti-angiogenic agents are used to treat disorders ranging from macular degeneration to cancer. In this review I describe in vitro assays that can be used to assess the activity of agents that affect angiogenesis. Means of quantifying endothelial cell matrix degradation, migration, proliferation, apoptosis and morphogenesis are discussed, as are embryoid body, aortic ring and metatarsal assays of vessel outgrowth. Strengths and limitations of these techniques are also addressed.

show abstract

Angiogenic sprouting and capillary lumen formation modeled by human umbilical vein endothelial cells (HUVEC) in fibrin gels: the role of fibroblasts and Angiopoietin-1☆

Cited by 414 publications

References 39 publications

Bovine Lactoferricin Inhibits Basic Fibroblast Growth Factor- and Vascular Endothelial Growth Factor165-Induced Angiogenesis by Competing for Heparin-Like Binding Sites on Endothelial Cells

Bovine Lactoferricin Inhibits Basic Fibroblast Growth Factor- and Vascular Endothelial Growth Factor165-Induced Angiogenesis by Competing for Heparin-Like Binding Sites on Endothelial Cells

miR‐342‐5p Is a Notch Downstream Molecule and Regulates Multiple Angiogenic Pathways Including Notch, Vascular Endothelial Growth Factor and Transforming Growth Factor β Signaling

In vitro assays of angiogenesis for assessment of angiogenic and anti-angiogenic agents

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