scholarly journals The VEGF-regulated transcription factor HLX controls the expression of guidance cues and negatively regulates sprouting of endothelial cells

Blood ◽  
2011 ◽  
Vol 117 (9) ◽  
pp. 2735-2744 ◽  
Author(s):  
Julia Testori ◽  
Bernhard Schweighofer ◽  
Iris Helfrich ◽  
Caterina Sturtzel ◽  
Karoline Lipnik ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Transcription Factor ◽  
Endothelial Cell Migration ◽  
Gene Repertoire ◽  
Gene Encoding ◽  
Hypoxic Conditions ◽  
Homeobox Transcription Factor ◽  
Endothelial Growth Factor

Abstract The HLX gene encoding a diverged homeobox transcription factor has been found to be up-regulated by vascular endothelial growth factor-A (VEGF-A) in endothelial cells. We have now investigated the gene repertoire induced by HLX and its potential biologic function. HLX strongly increased the transcripts for several repulsive cell-guidance proteins including UNC5B, plexin-A1, and semaphorin-3G. In addition, genes for transcriptional repressors such as HES-1 were up-regulated. In line with these findings, adenoviral overexpression of HLX inhibited endothelial cell migration, sprouting, and vessel formation in vitro and in vivo, whereas proliferation was unaffected. This inhibition of sprouting was caused to a significant part by HLX-mediated up-regulation of UNC5B as shown by short hairpin RNA (shRNA)–mediated down-modulation of the respective mRNA. VEGF-A stimulation of endothelial cells induced elevated levels of HLX over longer time periods resulting in especially high up-regulation of UNC5B mRNA as well as an increase in cells displaying UNC5B at their surface. However, induction of HLX was strongly reduced and UNC5B up-regulation completely abrogated when cells were exposed to hypoxic conditions. These data suggest that HLX may function to balance attractive with repulsive vessel guidance by up-regulating UNC5B and to down-modulate sprouting under normoxic conditions.

Vascular endothelial growth factor (VEGF) in cartilage neovascularization and chondrocyte differentiation: auto-paracrine role during endochondral bone formation

2000 ◽  
Vol 113 (1) ◽  
pp. 59-69 ◽  
Author(s):  
M.F. Carlevaro ◽  
S. Cermelli ◽  
R. Cancedda ◽  
F. Descalzi Cancedda
Keyword(s):  
Endothelial Cells ◽  
Cell Migration ◽  
Endothelial Cell ◽  
Hypertrophic Chondrocytes ◽  
Endothelial Cell Migration ◽  
Vegf Receptor ◽  
Hypertrophic Cartilage ◽  
Endothelial Growth Factor

Vascular endothelial growth factor/vascular permeability factor (VEGF/VPF) induces endothelial cell migration and proliferation in culture and is strongly angiogenic in vivo. VEGF synthesis has been shown to occur in both normal and transformed cells. The receptors for the factor have been shown to be localized mainly in endothelial cells, however, the presence of VEGF synthesis and the VEGF receptor in cells other than endothelial cells has been demonstrated. Neoangiogenesis in cartilage growth plate plays a fundamental role in endochondral ossification. We have shown that, in an avian in vitro system for chondrocyte differentiation, VEGF was produced and localized in cell clusters totally resembling in vivo cartilage. The factor was synthesized by hypertrophic chondrocytes and was released into their conditioned medium, which is highly chemotactic for endothelial cells. Antibodies against VEGF inhibited endothelial cell migration induced by chondrocyte conditioned media. Similarly, endothelial cell migration was inhibited also by antibodies directed against the VEGF receptor 2/Flk1 (VEGFR2). In avian and mammalian embryo long bones, immediately before vascular invasion, VEGF was distinctly localized in growth plate hypertrophic chondrocytes. In contrast, VEGF was not observed in quiescent and proliferating chondrocytes earlier in development. VEGF receptor 2 colocalized with the factor both in hypertrophic cartilage in vivo and hypertrophic cartilage engineered in vitro, suggesting an autocrine loop in chondrocytes at the time of their maturation to hypertrophic cells and of cartilage erosion. Regardless of cell exposure to exogenous VEGF, VEGFR-2 phosphorylation was recognized in cultured hypertrophic chondrocytes, supporting the idea of an autocrine functional activation of signal transduction in this non-endothelial cell type as a consequence of the endogenous VEGF production. In summary we propose that VEGF is actively responsible for hypertrophic cartilage neovascularization through a paracrine release by chondrocytes, with invading endothelial cells as a target. Furthermore, VEGF receptor localization and signal transduction in chondrocytes strongly support the hypothesis of a VEGF autocrine activity also in morphogenesis and differentiation of a mesoderm derived cell.

Reduced TIMP-2 in hypoxia enhances angiogenesis

2011 ◽  
Vol 300 (3) ◽  
pp. C557-C566 ◽  
Author(s):  
Nitza Lahat ◽  
Haim Bitterman ◽  
Miri Engelmayer-Goren ◽  
Doron Rosenzweig ◽  
Lea Weiss-Cerem ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Transcription Factor ◽  
Cell Migration ◽  
Endothelial Cell ◽  
Protein Secretion ◽  
Endothelial Cell Migration ◽  
Human Endothelial Cells ◽  
Monocyte Migration

Hypoxia, which characterizes ischemia, trauma, inflammation, and solid tumors, recruits monocytes, immobilizes them, and alters their function, leading to an anti-inflammatory and proangiogenic phenotype. Monocyte extravasation from the circulation and their migration in tissues are partially mediated by the balance between matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs). The mechanisms evoked by hypoxia that regulate monocyte migration and activation are not entirely clear. Specifically, the effect of hypoxia on TIMPs in these cells has hardly been investigated. We show that hypoxia reduces TIMP-2 secretion from human primary monocytes and from the monocyte-like cell lines U937 and THP-1 by three- to fourfold ( P < 0.01), by inhibiting TIMP-2 transcription through mechanisms that involve the transcription factor SP-1. Hypoxia also lowers TIMP-2 protein secretion from human endothelial cells (by 2-fold, P < 0.05). TIMP-2 levels do not influence the reduced migration of THP-1 cells in hypoxia; however, low TIMP-2 levels enhance endothelial cell migration/proliferation, their ability to form tubelike structures in vitro, and the appearance of mature blood vessels in a Matrigel plug assay in vivo. Thus we conclude that reduced TIMP-2 levels secreted from both hypoxic monocytes and endothelial cells are proangiogenic.

scholarly journals Nogo-B receptor is essential for angiogenesis in zebrafish via Akt pathway

Blood ◽  
2010 ◽  
Vol 116 (24) ◽  
pp. 5423-5433 ◽  
Author(s):  
Baofeng Zhao ◽  
Changzoon Chun ◽  
Zhong Liu ◽  
Mark A. Horswill ◽  
Kallal Pramanik ◽  
...  
Keyword(s):  
Vascular Endothelial Growth Factor ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Growth Factor ◽  
Umbilical Vein ◽  
Endothelial Cell Migration ◽  
Vascular Endothelial ◽  
Endothelial Growth Factor

Abstract Our previous work has shown that axon guidance gene family Nogo-B and its receptor (NgBR) are essential for chemotaxis and morphogenesis of endothelial cells in vitro. To investigate NogoB-NgBR function in vivo, we cloned the zebrafish ortholog of both genes and studied loss of function in vivo using morpholino antisense technology. Zebrafish ortholog of Nogo-B is expressed in somite while expression of zebrafish NgBR is localized in intersomitic vessel (ISV) and axial dorsal aorta during embryonic development. NgBR or Nogo-B knockdown embryos show defects in ISV sprouting in the zebrafish trunk. Mechanistically, we found that NgBR knockdown not only abolished its ligand Nogo-B–stimulated endothelial cell migration but also reduced the vascular endothelial growth factor (VEGF)–stimulated phosphorylation of Akt and vascular endothelial growth factor–induced chemotaxis and morphogenesis of human umbilical vein endothelial cells. Further, constitutively activated Akt (myristoylated [myr]Akt) or human NgBR can rescue the NgBR knockdown umbilical vein endothelial cell migration defects in vitro or NgBR morpholino-caused ISV defects in vivo. These data place Akt at the downstream of NgBR in both Nogo-B– and VEGF-coordinated sprouting of ISVs. In summary, this study identifies the in vivo functional role for Nogo-B and its receptor (NgBR) in angiogenesis in zebrafish.

Interaction between vascular endothelial cells and vascular intimal spindle-shaped cells in vitro

1983 ◽  
Vol 60 (1) ◽  
pp. 89-102
Author(s):  
D de Bono ◽  
C. Green
Keyword(s):  
Extracellular Matrix ◽  
Endothelial Cells ◽  
Vascular Endothelial Cells ◽  
Three Dimensional ◽  
Vascular Endothelial ◽  
Pure Cultures ◽  
Species Specific ◽  
Endothelial Growth Factor

The interactions between human or bovine vascular endothelial cells and fibroblast-like vascular intimal spindle-shaped cells have been studied in vitro, using species-specific antibodies to identify the different components in mixed cultures. Pure cultures of endothelial cells grow as uniform, nonoverlapping monolayers, but this growth pattern is lost after the addition of spindle cells, probably because the extracellular matrix secreted by the latter causes the endothelial cells to modify the way they are attached to the substrate. The result is a network of tubular aggregates of endothelial cells in a three-dimensional ‘polylayer’ of spindle-shaped cells. On the other hand, endothelial cells added to growth-inhibited cultures of spindle-shaped cells will grow in sheets over the surface of the culture. Human endothelial cells grown in contact with spindle-shaped cells have a reduced requirement for a brain-derived endothelial growth factor. The interactions of endothelial cells and other connective tissue cells in vitro may be relevant to the mechanisms of endothelial growth and blood vessel formation in vivo, and emphasize the potential importance of extracellular matrix in controlling endothelial cell behaviour.

scholarly journals Cystine/glutamate antiporter xCT (SLC7A11) facilitates oncogenic RAS transformation by preserving intracellular redox balance

2019 ◽  
Vol 116 (19) ◽  
pp. 9433-9442 ◽  
Author(s):  
Jonathan K. M. Lim ◽  
Alberto Delaidelli ◽  
Sean W. Minaker ◽  
Hai-Feng Zhang ◽  
Milena Colovic ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Transcription Factor ◽  
Redox Balance ◽  
Gene Encoding ◽  
Oncogenic Ras ◽  
Opposing Effects ◽  
Whole Transcriptome ◽  
Intracellular Redox

The RAS family of proto-oncogenes are among the most commonly mutated genes in human cancers and predict poor clinical outcome. Several mechanisms underlying oncogenic RAS transformation are well documented, including constitutive signaling through the RAF-MEK-ERK proproliferative pathway as well as the PI3K-AKT prosurvival pathway. Notably, control of redox balance has also been proposed to contribute to RAS transformation. However, how homeostasis between reactive oxygen species (ROS) and antioxidants, which have opposing effects in the cell, ultimately influence RAS-mediated transformation and tumor progression is still a matter of debate and the mechanisms involved have not been fully elucidated. Here, we show that oncogenic KRAS protects fibroblasts from oxidative stress by enhancing intracellular GSH levels. Using a whole transcriptome approach, we discovered that this is attributable to transcriptional up-regulation of xCT, the gene encoding the cystine/glutamate antiporter. This is in line with the function of xCT, which mediates the uptake of cystine, a precursor for GSH biosynthesis. Moreover, our results reveal that the ETS-1 transcription factor downstream of the RAS-RAF-MEK-ERK signaling cascade directly transactivates the xCT promoter in synergy with the ATF4 endoplasmic reticulum stress-associated transcription factor. Strikingly, xCT was found to be essential for oncogenic KRAS-mediated transformation in vitro and in vivo by mitigating oxidative stress, as knockdown of xCT strongly impaired growth of tumor xenografts established from KRAS-transformed cells. Overall, this study uncovers a mechanism by which oncogenic RAS preserves intracellular redox balance and identifies an unexpected role for xCT in supporting RAS-induced transformation and tumorigenicity.

Activating transcription factor 2 increases transactivation and protein stability of hypoxia-inducible factor 1α in hepatocytes

2009 ◽  
Vol 424 (2) ◽  
pp. 285-296 ◽  
Author(s):  
Jeong Hae Choi ◽  
Hyun Kook Cho ◽  
Yung Hyun Choi ◽  
JaeHun Cheong
Keyword(s):  
Transcription Factor ◽  
Protein Stability ◽  
Gene Transcription ◽  
Hypoxia Inducible Factor ◽  
Protein Stabilization ◽  
Hypoxic Conditions ◽  
Tumour Suppressor Protein ◽  
Activating Transcription Factor

HIF-1 (hypoxia inducible factor 1) performs a crucial role in mediating the response to hypoxia. However, other transcription factors are also capable of regulating hypoxia-induced target-gene transcription. In a previous report, we demonstrated that the transcription factor ATF-2 (activating transcription factor 2) regulates hypoxia-induced gene transcription, along with HIF-1α. In the present study, we show that the protein stability of ATF-2 is induced by hypoxia and the hypoxia-mimic CoCl2 (cobalt chloride), and that ATF-2 induction enhances HIF-1α protein stability via direct protein interaction. The knockdown of ATF-2 using small interfering RNA and translation-inhibition experiments demonstrated that ATF-2 plays a key role in the maintenance of the expression level and transcriptional activity of HIF-1α. Furthermore, we determined that ATF-2 interacts directly with HIF-1α both in vivo and in vitro and competes with the tumour suppressor protein p53 for HIF-1α binding. Collectively, these results show that protein stabilization of ATF-2 under hypoxic conditions is required for the induction of the protein stability and transactivation activity of HIF-1α for efficient hypoxia-associated gene expression.

CYLD regulates angiogenesis by mediating vascular endothelial cell migration

Blood ◽  
2010 ◽  
Vol 115 (20) ◽  
pp. 4130-4137 ◽  
Author(s):  
Jinmin Gao ◽  
Lei Sun ◽  
Lihong Huo ◽  
Min Liu ◽  
Dengwen Li ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Cell Migration ◽  
Endothelial Cell ◽  
Vascular Endothelial Cells ◽  
Vascular Endothelial Cell ◽  
Tube Formation ◽  
Endothelial Cell Migration ◽  
Vascular Endothelial

Cylindromatosis (CYLD) is a deubiquitinase that was initially identified as a tumor suppressor and has recently been implicated in diverse normal physiologic processes. In this study, we have investigated the involvement of CYLD in angiogenesis, the formation of new blood vessels from preexisting ones. We find that knockdown of CYLD expression significantly impairs angiogenesis in vitro in both matrigel-based tube formation assay and collagen-based 3-dimensional capillary sprouting assay. Disruption of CYLD also remarkably inhibits angiogenic response in vivo, as evidenced by diminished blood vessel growth into the angioreactors implanted in mice. Mechanistic studies show that CYLD regulates angiogenesis by mediating the spreading and migration of vascular endothelial cells. Silencing of CYLD dramatically decreases microtubule dynamics in endothelial cells and inhibits endothelial cell migration by blocking the polarization process. Furthermore, we identify Rac1 activation as an important factor contributing to the action of CYLD in regulating endothelial cell migration and angiogenesis. Our findings thus uncover a previously unrecognized role for CYLD in the angiogenic process and provide a novel mechanism for Rac1 activation during endothelial cell migration and angiogenesis.

β2-Glycoprotein I as a ‘Cofactor’ for anti-phospholipid reactivity with endothelial cells

Lupus ◽  
1998 ◽  
Vol 7 (2_suppl) ◽  
pp. 44-47 ◽  
Author(s):  
PL Meroni ◽  
N Del Papa ◽  
E Raschi ◽  
P Panzeri ◽  
MO Borghi ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Binding Site ◽  
Synthetic Peptide ◽  
Cell Activation ◽  
Heparan Sulphate ◽  
Membrane Structures ◽  
Glycoprotein I ◽  
Endothelial Growth Factor

β2-glycoprotein I (β2GPI) is a cofactor for anti-phospholipid (aPL) binding to cardiolipin (CL)-coated plates. β2GPI is also able to bind to endothelial cell (EC) membranes as supported by in-vivo as well as by in-vitro studies. The PL-binding site in the fifth domain of the molecule is involved in the adhesion to endothelium. Actually, specific mutations in this molecular portion abolish endothelium binding and a synthetic peptide spanning the sequence Glu274 –Cys288 of the CL-binding site displays comparable adhesion to EC monolayers. Heparan sulphate appears to be one of the anionic EC membrane structures with which cationic β2GPI interacts, as supported by studies with heparitinase-treated EC. β2GPI binding to EC might be related to its activity as endothelial growth factor or as a lipid-carrying glycoprotein. Adhesion of β2GPI to endothelial membranes offers suitable epitopes for circulating aPL that, once bound, can induce cell activation

scholarly journals BMP9 regulates endoglin-dependent chemokine responses in endothelial cells

Blood ◽  
2012 ◽  
Vol 120 (20) ◽  
pp. 4263-4273 ◽  
Author(s):  
Kira Young ◽  
Barbara Conley ◽  
Diana Romero ◽  
Eric Tweedie ◽  
Christine O'Neill ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Target Genes ◽  
Endothelial Cell Migration ◽  
Quantitative Mass Spectrometry ◽  
Chemokine Cxcl12 ◽  
Vessel Maturation ◽  
Rna Knockdown ◽  
Deficient Mice

Abstract BMP9 signaling has been implicated in hereditary hemorrhagic telangiectasia (HHT) and vascular remodeling, acting via the HHT target genes, endoglin and ALK1. This study sought to identify endothelial BMP9-regulated proteins that could affect the HHT phenotype. Gene ontology analysis of cDNA microarray data obtained after BMP9 treatment of primary human endothelial cells indicated regulation of chemokine, adhesion, and inflammation pathways. These responses included the up-regulation of the chemokine CXCL12/SDF1 and down-regulation of its receptor CXCR4. Quantitative mass spectrometry identified additional secreted proteins, including the chemokine CXCL10/IP10. RNA knockdown of endoglin and ALK1 impaired SDF1/CXCR4 regulation by BMP9. Because of the association of SDF1 with ischemia, we analyzed its expression under hypoxia in response to BMP9 in vitro, and during the response to hindlimb ischemia, in endoglin-deficient mice. BMP9 and hypoxia were additive inducers of SDF1 expression. Moreover, the data suggest that endoglin deficiency impaired SDF1 expression in endothelial cells in vivo. Our data implicate BMP9 in regulation of the SDF1/CXCR4 chemokine axis in endothelial cells and point to a role for BMP9 signaling via endoglin in a switch from an SDF1-responsive autocrine phenotype to an SDF1 nonresponsive paracrine state that represses endothelial cell migration and may promote vessel maturation.

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