Shear stress increases expression of the arterial endothelial marker ephrinB2 in murine ES cells via the VEGF-Notch signaling pathways.
OBJECTIVE
Arterial-venous specification in the embryo has been assumed to depend on the influence of fluid mechanical forces, but its cellular and molecular mechanisms are still poorly understood. Our previous in vitro study revealed that fluid shear stress induces endothelial cell (EC) differentiation by murine embryonic stem (ES) cells. In the present study we investigated whether shear stress regulates the arterial-venous specification of ES-cell-derived ECs.


RESULTS
When murine ES cell-derived VEGFR2(+) cells were exposed to shear stress, expression of the arterial EC marker protein ephrinB2 increased dose-dependently. The ephrinB2 mRNA levels also increased in response to shear stress, whereas the mRNA levels of the venous EC marker EphB4 decreased. Notch cleavage and translocation of the Notch intracellular domain (NICD) into the nucleus occurred as early as 30 minutes after the start of shear stress and increased with time. Gamma-Secretase inhibitors (DAPT and L685 458) and the recombinant extracellular domain of the Notch ligand DLL4 abolished the shear stress-induced NICD translocation, and that, in turn, blocked the shear stress-induced upregulation of ephrinB2 expression. In addition, the VEGF receptor kinase inhibitor SU1498 was found to suppress both the shear-stress-induced Notch cleavage and up-regulation of ephrinB2 expression.


CONCLUSIONS
Exposure to shear stress induces an increase in expression of ephrinB2 in murine ES cells via VEGF-Notch signaling pathways.
Department of Biomedical Engineering, Graduate School of Medicine, University of Tokyo, Japan.
Tomomi
T
Masumura
Kimiko
K
Yamamoto
Nobutaka
N
Shimizu
Syotaro
S
Obi
Joji
J
Ando
D000818:Animals; D001483:Base Sequence; D015415:Biomarkers / Q000378:metabolism; D001696:Biomechanical Phenomena; D002454:Cell Differentiation; D002478:Cells, Cultured; D017931:DNA Primers / Q000235:genetics; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism*; D036388:Ephrin-B2 / Q000235:genetics* / Q000378:metabolism*; D015870:Gene Expression; D051379:Mice; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D036224:Receptor, EphB4 / Q000235:genetics / Q000378:metabolism; D051880:Receptors, Notch / Q000378:metabolism*; D015398:Signal Transduction; D013234:Stem Cells / Q000166:cytology / Q000378:metabolism*; D013314:Stress, Mechanical; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism*; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000378:metabolism
D015415:Biomarkers; D017931:DNA Primers; D036388:Ephrin-B2; D012333:RNA, Messenger; D051880:Receptors, Notch; D042461:Vascular Endothelial Growth Factor A; C467480:vascular endothelial growth factor A, mouse; C086681:Ephb4 protein, mouse; D036224:Receptor, EphB4; D040301:Vascular Endothelial Growth Factor Receptor-2