Impact of Hey2 and COUP-TFII on genes involved in arteriovenous differentiation in primary human arterial and venous endothelial cells

Abstract

Arteries and veins show marked differences in their anatomy, physiology and genetic expression pattern. In this study, we analyzed impact of overexpression or downregulation of arterial marker gene Hey2 and venous marker gene COUP-TFII in human venous and arterial endothelial cells on genes involved in arteriovenous differentiation. Lentiviral overexpression of venous marker gene COUP-TFII in arterial endothelial cells led to downregulation of NICD4, arterial marker gene Hey2 and EphrinB2. Downregulation of Hey2 could be mediated by direct binding of COUP-TFII to Hey2 promoter as shown by ChIP, EMSA and promoter analysis. Downregulation of Hey2 by shRNA causes downregulation of EphrinB2 expression. Overexpression of arterial marker Hey2 in venous endothelial cells did not change expression pattern of COUP-TFII. Downregulation of venous marker gene COUP-TFII in venous endothelial cells resulted in upregulation of VEGF-A, Dll4 and EphrinB2 expression. Our data support an important role of Hey2 and COUP-TFII in arteriovenous differentiation of human endothelial cells.

Human brain arteriovenous malformations express lymphatic‐associated genes

“…In rodents, both loss-of- and gain-of-function Notch signaling have been shown to generate vascular malformations,42,43 while normalization of Notch signaling leads to malformation regression 10. Overexpression of COUP-TFII in vivo also resulted in vascular malformations in a conditional, EC-specific mouse model16 and in vitro overexpression of COUP-TFII alone can downregulate HEY2, a downstream target of Notch signaling 44. Finally, EC-specific overexpression of PROX1 results in hemorrhage, edema, anemia, and embryonic lethality at E13.5, possibly due to alterations in tight junction proteins such as ZO-1 and Occludin 45.…”

Cigarette smoke extract counteracts atheroprotective effects of high laminar flow on endothelial function

“…Primary cultures of HUVEC were isolated using 0.5% collagenase II solution (Worthington Biochemical Corp., Lakewood NJ, USA) [32], [33], [34]. Isolated HUVEC were cultured on 2% gelatin-coated plates in Medium 199 (Thermo Fisher Scientific, Waltham, MA, USA), supplemented with 10% fetal calf serum (Biochrom, Berlin, Germany), 0.5% self-isolated retina calf eye growth supplement [35], 100,000 U/l penicillin (Thermo Fisher Scientific, Waltham, MA, USA), 100 mg/l streptomycin (Thermo Fisher Scientific, Waltham, MA, USA) and 250 mg/l Fungizone (Thermo Fisher Scientific, Waltham, MA, USA).…”

“…NQO1-ARE and mutated NQO1-ARE containing pGL4 luciferase reporter vectors were kindly provided by Anna–Liisa Levonen and Hanna Leinonen (University of Eastern Finland, Kuopio, Finland) [34], [44]. In brief, human microvascular (HMEC-1) endothelial cells were transfected using Fugene HD in 24 well plates (Fugene HD: DNA-Ratio =3:1).…”

Hey bHLH Transcription Factors

“…8.5). On the other hand, downregulation or overexpression of Hey2 in human arterial or venous endothelial cells, respectively, did not influence CoupTF-II expression (Korten et al, 2013). On the other hand, downregulation or overexpression of Hey2 in human arterial or venous endothelial cells, respectively, did not influence CoupTF-II expression (Korten et al, 2013).…”

“…It represses Hey2 in atrial cardiomyocytes (Wu et al, 2013). This appears to be facilitated by Prox1 forming heterodimers with CoupTF-II, which lack repressive capacity toward Hey1 and Hey2 (Korten et al, 2013;Yoo et al, 2012). In lymphatic endothelia cells, however, Hey1/2 and CoupTF-II are coexpressed.…”