Low-dose 6-bromoindirubin-3'-oxime induces partial dedifferentiation of endothelial cells to promote increased neovascularization.
Endothelial cell (EC) dedifferentiation in relation to neovascularization is a poorly understood process. In this report, we addressed the role of Wnt signaling in the mechanisms of neovascularization in adult tissues. Here, we show that a low-dose of 6-bromoindirubin-3'-oxime (BIO), a competitive inhibitor of glycogen synthase kinase-3β, induced the stabilization of β-catenin and its subsequent direct interaction with the transcription factor NANOG in the nucleus of ECs. This event induced loss of VE-cadherin from the adherens junctions, increased EC proliferation accompanied by asymmetric cell division (ACD), and formed cellular aggregates in hanging drop assays indicating the acquisition of a dedifferentiated state. In a chromatin immunoprecipitation assay, nuclear NANOG protein bound to the NANOG- and VEGFR2-promoters in ECs, and the addition of BIO activated the NANOG-promoter-luciferase reporter system in a cell-based assay. Consequently, NANOG-knockdown decreased BIO-induced NOTCH-1 expression, thereby decreasing cell proliferation, ACD, and neovascularization. In a Matrigel plug assay, BIO induced increased neovascularization, secondary to the presence of vascular endothelial growth factor (VEGF). Moreover, in a mouse model of hind limb ischemia, BIO augmented neovascularization that was coupled with increased expression of NOTCH-1 in ECs and increased smooth muscle α-actin(+) cell recruitment around the neovessels. Thus, these results demonstrate the ability of a low-dose of BIO to augment neovascularization secondary to VEGF, a process that was accompanied by a partial dedifferentiation of ECs via β-catenin and the NANOG signaling pathway.
Department of Pharmacology, University of Illinois at Chicago, Chicago, Illinois, USA.
Erin E
EE
Kohler
Jugajyoti
J
Baruah
Norifumi
N
Urao
Masuko
M
Ushio-Fukai
Tohru
T
Fukai
Ishita
I
Chatterjee
Kishore K
KK
Wary
D043925:Angiogenesis Inducing Agents / Q000378:metabolism; D000818:Animals; D002449:Cell Aggregation / Q000187:drug effects; D054337:Cell Dedifferentiation / Q000187:drug effects*; D002455:Cell Division / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D004195:Disease Models, Animal; D004305:Dose-Response Relationship, Drug; D042783:Endothelial Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D005326:Fetal Proteins / Q000235:genetics; D006614:Hindlimb / Q000098:blood supply / Q000473:pathology; D018398:Homeodomain Proteins / Q000378:metabolism; D061307:Human Umbilical Vein Endothelial Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D007211:Indoles / Q000008:administration & dosage / Q000494:pharmacology*; D007511:Ischemia / Q000473:pathology; D051379:Mice; D000071317:Nanog Homeobox Protein; D018919:Neovascularization, Physiologic / Q000187:drug effects*; D010091:Oximes / Q000008:administration & dosage / Q000494:pharmacology*; D010641:Phenotype; D039904:Pluripotent Stem Cells / Q000187:drug effects / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding / Q000187:drug effects; D055550:Protein Stability / Q000187:drug effects; D020825:T-Box Domain Proteins / Q000235:genetics; D042461:Vascular Endothelial Growth Factor A / Q000494:pharmacology; D051176:beta Catenin / Q000378:metabolism; D000098908:Brachyury Protein
D043925:Angiogenesis Inducing Agents; D005326:Fetal Proteins; D018398:Homeodomain Proteins; D007211:Indoles; D000071317:Nanog Homeobox Protein; D010091:Oximes; D020825:T-Box Domain Proteins; D042461:Vascular Endothelial Growth Factor A; D051176:beta Catenin; D000098908:Brachyury Protein; C483321:6-bromoindirubin-3'-oxime; C486780:NANOG protein, human
6-Bromoindirubin-3′-oxime; Dedifferentiation; Endothelial cells; Hind limb ischemia; Neovascularization