Glycome and transcriptome regulation of vasculogenesis.
BACKGROUND
Therapeutic vasculogenesis is an emerging concept that can potentially be harnessed for the management of ischemic pathologies. The present study elucidates the potential coregulation of vasculogenesis by the heparan sulfate glycosaminoglycan-rich cell-surface glycome and the transcriptome.


RESULTS
Differentiation of embryonic stem cells into endothelial cells in an in vitro embryoid body is paralleled by an amplification of heparan sulfate glycosaminoglycan sulfation, which correlates with the levels of the enzyme N-deacetylase/N-sulfotransferase 1 (NDST1). Small hairpin RNA-mediated knockdown of NDST1 or modification of heparan sulfate glycosaminoglycans in embryonic stem cells with heparinases or sodium chlorate inhibited differentiation of embryonic stem cells into endothelial cells. This was translated to an in vivo zebrafish embryo model, in which the genetic knockdown of NDST1 resulted in impaired vascularization characterized by a concentration-dependent decrease in intersegmental vessel lumen and a large tail-vessel configuration, which could be rescued by use of exogenous sulfated heparan sulfate glycosaminoglycans. To explore the cross talk between the glycome and the transcriptome during vasculogenesis, we identified by microarray and then validated wild-type and NDST1 knockdown-associated gene-expression patterns in zebrafish embryos. Temporal analysis at 3 developmental stages critical for vasculogenesis revealed a cascade of pathways that may mediate glycocalyx regulation of vasculogenesis. These pathways were intimately connected to cell signaling, cell survival, and cell fate determination. Specifically, we demonstrated that forkhead box O3A/5 proteins and insulin-like growth factor were key downstream signals in this process.


CONCLUSIONS
The present study for the first time implicates interplay between the glycome and the transcriptome during vasculogenesis, revealing the possibility of harnessing specific cellular glyco-microenvironments for therapeutic vascularization.
Department of Medicine (R.H., D.M.H., S.P., S.B., S.S.), Brigham and Women's Hospital, Boston, Mass; Department of Molecular Medicine and Pathology (C.P.), University of Auckland, Auckland, New Zealand; Department of Biological Engineering (D.E., T.K., R.S.), Massachusetts Institute of Technology, Cambridge, Mass; and Harvard-MIT Division of Health Sciences and Technology (R.H., S.P., S.B., D.E., T.K., R.S., S.S.), Cambridge, Mass.
Rania
R
Harfouche
Department of Medicine (R.H., D.M.H., S.P., S.B., S.S.), Brigham and Women's Hospital, Boston, Mass; Department of Molecular Medicine and Pathology (C.P.), University of Auckland, Auckland, New Zealand; Department of Biological Engineering (D.E., T.K., R.S.), Massachusetts Institute of Technology, Cambridge, Mass; and Harvard-MIT Division of Health Sciences and Technology (R.H., S.P., S.B., D.E., T.K., R.S., S.S.), Cambridge, Mass.
Dirk M
DM
Hentschel
Department of Medicine (R.H., D.M.H., S.P., S.B., S.S.), Brigham and Women's Hospital, Boston, Mass; Department of Molecular Medicine and Pathology (C.P.), University of Auckland, Auckland, New Zealand; Department of Biological Engineering (D.E., T.K., R.S.), Massachusetts Institute of Technology, Cambridge, Mass; and Harvard-MIT Division of Health Sciences and Technology (R.H., S.P., S.B., D.E., T.K., R.S., S.S.), Cambridge, Mass.
Stephanie
S
Piecewicz
Department of Medicine (R.H., D.M.H., S.P., S.B., S.S.), Brigham and Women's Hospital, Boston, Mass; Department of Molecular Medicine and Pathology (C.P.), University of Auckland, Auckland, New Zealand; Department of Biological Engineering (D.E., T.K., R.S.), Massachusetts Institute of Technology, Cambridge, Mass; and Harvard-MIT Division of Health Sciences and Technology (R.H., S.P., S.B., D.E., T.K., R.S., S.S.), Cambridge, Mass.
Sudipta
S
Basu
Department of Medicine (R.H., D.M.H., S.P., S.B., S.S.), Brigham and Women's Hospital, Boston, Mass; Department of Molecular Medicine and Pathology (C.P.), University of Auckland, Auckland, New Zealand; Department of Biological Engineering (D.E., T.K., R.S.), Massachusetts Institute of Technology, Cambridge, Mass; and Harvard-MIT Division of Health Sciences and Technology (R.H., S.P., S.B., D.E., T.K., R.S., S.S.), Cambridge, Mass.
Cristin
C
Print
Department of Medicine (R.H., D.M.H., S.P., S.B., S.S.), Brigham and Women's Hospital, Boston, Mass; Department of Molecular Medicine and Pathology (C.P.), University of Auckland, Auckland, New Zealand; Department of Biological Engineering (D.E., T.K., R.S.), Massachusetts Institute of Technology, Cambridge, Mass; and Harvard-MIT Division of Health Sciences and Technology (R.H., S.P., S.B., D.E., T.K., R.S., S.S.), Cambridge, Mass.
David
D
Eavarone
Department of Medicine (R.H., D.M.H., S.P., S.B., S.S.), Brigham and Women's Hospital, Boston, Mass; Department of Molecular Medicine and Pathology (C.P.), University of Auckland, Auckland, New Zealand; Department of Biological Engineering (D.E., T.K., R.S.), Massachusetts Institute of Technology, Cambridge, Mass; and Harvard-MIT Division of Health Sciences and Technology (R.H., S.P., S.B., D.E., T.K., R.S., S.S.), Cambridge, Mass.
Tanyel
T
Kiziltepe
Department of Medicine (R.H., D.M.H., S.P., S.B., S.S.), Brigham and Women's Hospital, Boston, Mass; Department of Molecular Medicine and Pathology (C.P.), University of Auckland, Auckland, New Zealand; Department of Biological Engineering (D.E., T.K., R.S.), Massachusetts Institute of Technology, Cambridge, Mass; and Harvard-MIT Division of Health Sciences and Technology (R.H., S.P., S.B., D.E., T.K., R.S., S.S.), Cambridge, Mass.
Ram
R
Sasisekharan
Department of Medicine (R.H., D.M.H., S.P., S.B., S.S.), Brigham and Women's Hospital, Boston, Mass; Department of Molecular Medicine and Pathology (C.P.), University of Auckland, Auckland, New Zealand; Department of Biological Engineering (D.E., T.K., R.S.), Massachusetts Institute of Technology, Cambridge, Mass; and Harvard-MIT Division of Health Sciences and Technology (R.H., S.P., S.B., D.E., T.K., R.S., S.S.), Cambridge, Mass.
Shiladitya
S
Sengupta
D000818:Animals; D002454:Cell Differentiation / Q000502:physiology; D002478:Cells, Cultured; D053595:Embryonic Stem Cells / Q000166:cytology / Q000502:physiology*; D042783:Endothelial Cells / Q000166:cytology / Q000502:physiology*; D000071316:Forkhead Box Protein O3; D051858:Forkhead Transcription Factors / Q000235:genetics / Q000378:metabolism; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental; D055785:Gene Knockdown Techniques; D019276:Glycocalyx / Q000502:physiology; D054794:Glycomics; D019812:Heparan Sulfate Proteoglycans / Q000235:genetics / Q000378:metabolism; D007335:Insulin-Like Growth Factor II / Q000235:genetics / Q000378:metabolism; D051379:Mice; D023421:Models, Animal; D018919:Neovascularization, Physiologic / Q000502:physiology*; D010641:Phenotype; D015398:Signal Transduction / Q000502:physiology; D015238:Sulfotransferases / Q000235:genetics / Q000378:metabolism; D015027:Zebrafish
D000071316:Forkhead Box Protein O3; D051858:Forkhead Transcription Factors; C474033:FoxO3 protein, mouse; D019812:Heparan Sulfate Proteoglycans; D007335:Insulin-Like Growth Factor II; D015238:Sulfotransferases; C021350:heparitin sulfotransferase