scholarly journals Characterization of Heparin Affin Regulatory Peptide Signaling in Human Endothelial Cells

2005 ◽  
Vol 280 (23) ◽  
pp. 22454-22461 ◽  
Author(s):  
Apostolos Polykratis ◽  
Panagiotis Katsoris ◽  
José Courty ◽  
Evangelia Papadimitriou
Keyword(s):  
Endothelial Cells ◽  
Umbilical Vein ◽  
Tyrosine Phosphatase ◽  
Neuronal Cell ◽  
Cell Types ◽  
Tube Formation ◽  
Regulatory Peptide ◽  
Intracellular Signals ◽  
Umbilical Vein Endothelial Cells ◽  
Interfering Rna

Heparin affin regulatory peptide (HARP) is an 18-kDa secreted growth factor that has a high affinity for heparin and a potent role on tumor growth and angiogenesis. We have previously reported that HARP is mitogenic for different types of endothelial cells and also affects cell migration and differentiation (12). In this study we examined the signaling pathways involved in the migration and tube formation on matrigel of human umbilical vein endothelial cells (HUVEC) induced by HARP. We report for the first time that receptor-type protein-tyrosine phosphatase β/ζ (RPTPβ/ζ), which is a receptor for HARP in neuronal cell types, is also expressed in HUVEC. We also document that HARP signaling through RPTPβ/ζ leads to activation of Src kinase, focal adhesion kinase, phosphatidylinositol 3-kinase, and Erk1/2. Sodium orthovanadate, chondroitin sulfate-C, PP1, wortmannin, LY294002, and U0126 inhibit HARP-mediated signaling and HUVEC migration and tube formation. In addition, RPTPβ/ζ suppression using small interfering RNA technology interrupts intracellular signals and HUVEC migration and tube formation induced by HARP. These results establish the role of RPTPβ/ζ as a receptor of HARP in HUVEC and elucidate the HARP signaling pathway in endothelial cells.

Abstract 5440: Klf2 Inhibits Hif-1α Expression and Function in the Endothelium

Circulation ◽  
2008 ◽  
Vol 118 (suppl_18) ◽  
Author(s):  
Daiji Kawanami ◽  
Zhiyong Lin ◽  
Ganapati Mahabaleshwar ◽  
G. B Atkins ◽  
Anne Hamik ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Umbilical Vein ◽  
Hypoxia Inducible Factor ◽  
Proteasome Inhibitors ◽  
Cell Types ◽  
Tube Formation ◽  
Protein Levels ◽  
Umbilical Vein Endothelial Cells ◽  
And Function ◽  
Mediated Reduction

Introduction: Hypoxia-inducible factor 1 (HIF-1) is a central regulator of the hypoxic response in many cell types. In endothelial cells, HIF-1 induces the expression of key pro-angiogenic factors such as vascular endothelial growth factor (VEGF) to induce new blood vessel formation. Recent studies have identified Kruppel-like Factor 2 (KLF2) as a potent inhibitor of angiogenesis. However, the role of KLF2 in regulating the expression and function of HIF-1 has not been evaluated. Hypothesis: KLF2 inhibits HIF-1α expression and function. Methods and Results: Adenoviral overexpression of KLF2 in human umbilical vein endothelial cells (HUVECs) inhibited hypoxia-induced expression of HIF-1α, VEGF secretion, and matrigel tube formation. Conversely, siRNA-mediated knockdown of KLF2 in HUVECs increased HIF-1α expression and VEGF secretion. Consistent with this observation, KLF2−/− mouse embryonic fibroblasts (MEFs) showed accelerated HIF-1α accumulation in response to hypoxia and a marked induction of VEGF secretion (90.4±7.4 pg/ml in KLF2+/+ versus 234.6±6.1 pg/ml in KLF2−/−, p<0.01). From a mechanistic standpoint, KLF2-mediated reduction of HIF-1α level was attenuated by three distinct proteasome inhibitors (MG132, ALLN, or lactacystin), demonstrating that KLF2 mediated reduction of HIF-1α protein was proteasome-dependent. In addition, KLF2 maintained its ability to inhibit HIF-1α protein levels in VHL−/− cells, thermo-sensitive E1 deficient cells, and p53−/− cells. Finally, co-immunoprecipitation experiments revealed that KLF2 overexpression disrupted the interaction between HIF-1α and its chaperone Hsp90, suggesting that KLF2 promotes degradation of HIF-1α by affecting its folding and maturation. Conclusions: These observations identify KLF2 as a novel inhibitor of HIF-1α expression and function through a VHL/p53 independent but proteasome dependent pathway. As such, KLF2 may be a target for modulating the angiogenic response in disease states.

INTERACTION OF S PROTEIN/VITRONECTIN WITH CULTURED ENDOTHELIAL CELLS: PROMOTION OF ATTACHMENT AND SPECIFIC BINDING

1987 ◽  
Author(s):  
K T Preissner ◽  
E Anders ◽  
G Müller-Berghaus
Keyword(s):  
Endothelial Cells ◽  
Cell Attachment ◽  
Specific Binding ◽  
Umbilical Vein ◽  
Attachment Site ◽  
Cell Types ◽  
S Protein ◽  
Specific Association ◽  
Umbilical Vein Endothelial Cells ◽  
Adhesive Proteins

The interaction of the complement inhibitor S protein, which is identical to the serum spreading factor, vitronectin, with cultured human endothelial cells of macro- and microvas- cular origin was investigated. Purified S protein, coated for 2 h on polystyrene petri dishes, induced concentration- and time-dependent attachment and spreading of human umbilical vein endothelial cells (HUVEC) as well as human omental tissqe microvasular endothelial cells (HOTMEC) at 37°C. With 3 × 105 cells/ml (final concentration) more than 50% of the cells attached within 2 h incubation at 0.3 - 3 μg/ml S protein. The effect of S protein was specific, since only monospecific antibodies against S protein prevented attachment of cells, while antibodies against fibronectin, fibrinogen or von Wille-brand factor were uneffective. The pentapeptide Gly-Arg-Gly-Asp-Ser, which contains the cell-attachment site of these adhesive proteins including S protein, inhibited the activity of S protein to promote attachment of endothelial cells in a concentration-dependent fashion; at 200 μM peptide, less than 10% of the cells became attached. Direct binding of S protein to HUVEC and HOTMEC was studied with cells in suspension at a concentration of 1 × 106 cells/ml in the presence of 1% (w/v) human serum albumin and 1 mM CaCl2 and was maximal after 120 min. Both cell types bound S protein in a concentration-dependent fashion with an estimated dissociation constant KD=0.2pM. More than 80% of bound radiolabelled S protein was displaced by unlabelled S protein, whereas binding was reduced to about 50% by the addition in excess of either fibronectin, fibrinogen, von Willebrand factor or the pentapeptide. These findings provide evidence for the specific association of S protein with endothelial cells, ultimately leading to attachment and spreading of cells. Although the promotion of attachment was highly specific for S protein, other adhesive proteins than S protein, also known to associate with endothelial cells, may in part compete with direct S protein binding.

scholarly journals Hydrogel-Coated Textile Scaffolds as Three-Dimensional Growth Support for Human Umbilical Vein Endothelial Cells (HUVECs): Possibilities as Coculture System in Liver Tissue Engineering

2002 ◽  
Vol 11 (4) ◽  
pp. 369-377 ◽  
Author(s):  
Makarand V. Risbud ◽  
Erdal Karamuk ◽  
René Moser ◽  
Joerg Mayer
Keyword(s):  
Endothelial Cells ◽  
Umbilical Vein ◽  
Three Dimensional ◽  
Mesh Size ◽  
Cell Types ◽  
Cell Number ◽  
Human Umbilical Vein ◽  
Hydrogel Matrix ◽  
Umbilical Vein Endothelial Cells

Three-dimensional (3-D) scaffolds offer an exciting possibility to develop cocultures of various cell types. Here we report chitosan–collagen hydrogel-coated fabric scaffolds with defined mesh size and fiber diameter for 3-D culture of human umbilical vein endothelial cells (HUVECs). These scaffolds did not require pre-coating with fibronectin and they supported proper HUVEC attachment and growth. Scaffolds preserved endothelial cell-specific cobblestone morphology and cells were growing in compartments defined by the textile mesh. HUVECs on the scaffold maintained the property of contact inhibition and did not exhibit overgrowth until the end of in vitro culture (day 6). MTT assay showed that cells had preserved mitochondrial functionality. It was also noted that cell number on the chitosan-coated scaffold was lower than that of collagen-coated scaffolds. Calcein AM and ethidium homodimer (EtD-1) dual staining demonstrated presence of viable and metabolically active cells, indicating growth supportive properties of the scaffolds. Actin labeling revealed absence of actin stress fibers and uniform distribution of F-actin in the cells, indicating their proper attachment to the scaffold matrix. Confocal microscopic studies showed that HUVECs growing on the scaffold had preserved functionality as seen by expression of von Willebrand (vW) factor. Observations also revealed that functional HUVECs were growing at various depths in the hydrogel matrix, thus demonstrating the potential of these scaffolds to support 3-D growth of cells. We foresee the application of this scaffold system in the design of liver bioreactors wherein hepatocytes could be cocultured in parallel with endothelial cells to enhance and preserve liver-specific functions.

Abstract 103: MiR-4261 Controls Endothelial Cells Angiogenesis

2015 ◽  
Vol 117 (suppl_1) ◽  
Author(s):  
Qiulian Zhou ◽  
Dongchao Lv ◽  
Qi Sun ◽  
Ping Chen ◽  
Yihua Bei ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Molecular Mechanisms ◽  
Heart Diseases ◽  
Umbilical Vein ◽  
Tissue Perfusion ◽  
Tube Formation ◽  
Artery Disease ◽  
Incorporation Assay ◽  
Umbilical Vein Endothelial Cells

Myocardial infarction (MI) is among major causes of morbidity and mortality associated with coronary artery disease. Angiogenesis improves tissue perfusion and cardiac repair after MI. Therefore, angiogenesis is considered to be a novel therapeutic way for ischemic heart diseases. MicroRNAs (miRNAs, miRs) have been reported to play important roles in regulating post-ischemic neovascularization. The current study aims at investigating the role of miR-4261 in angiogenesis. We found that miR-4261 mimics increased, while miR-4261 inhibitors decreased the proliferation of human umbilical vein endothelial cells (HUVEC) using EdU incorporation assay (17.25%±1.31% vs 30.91%±0.92% in nc-mimics vs mir-4261-mimics, 17.91%±1.36% vs 8.51%±0.82% in nc-inhibitor vs mir-4261-inhibitor, respectively) and CCK-8 assays (0.84±0.04 vs 1.38±0.04 in nc-mimics vs mir-4261-mimics, 0.80±0.02 vs 0.72±0.01 in nc-inhibitor vs mir-4261-inhibitor, respectively). The wound healing assay showed that miR-4261 mimic transfection resulted in a significant increase in the migration of HUVEC compared to that of the negative controls while miR-4261 inhibition had the opposite effects. Tube formation assays showed that HUVEC transfected with miR-4261 mimics increased the number of tubes formed (57.25±2.56 vs 81.5±2.53 in nc-mimics vs mir-4261-mimics, respectively), while miR-4261 inhibitor-transfected cells had the opposite effect (56.55±0.45 vs 41.38±0.52 in nc-inhibitor vs mir-4261-inhibitor, respectively). These results indicate that miR-4261 play an important role in regulating angiogenesis. However, it remains unknown which target gene mediated the effects of miR-4261. Thus, it will be of great interest to further investigate the molecular mechanisms of miR-4261 in the proliferation, migration, and tube formation of HUVEC in vitro. MiR-4261 could be a potential therapeutic target to enhance angiogenesis.

Abstract 101: MiR-4260 Promotes the Proliferation, Migration, and Tube Formation of Human Umbilical Vein Endothelial Cells

2015 ◽  
Vol 117 (suppl_1) ◽  
Author(s):  
Qi Sun ◽  
Dongcao Lv ◽  
Qiulian Zhou ◽  
Yihua Bei ◽  
Junjie Xiao
Keyword(s):  
Endothelial Cells ◽  
Target Genes ◽  
Cell Function ◽  
Umbilical Vein ◽  
Tube Formation ◽  
Endothelial Cell Function ◽  
Human Umbilical Vein ◽  
And Migration ◽  
Umbilical Vein Endothelial Cells

MicroRNAs (miRNAs, miRs), endogenous small non-coding RNA, have been shown to act as essential regulators in angiogenesis which plays important roles in improving blood flow and cardiac function following myocardial infarction. The current study investigated the potential of miR-4260 in endothelial cell function and angiogenesis using human umbilical vein endothelial cells (HUVEC). Our data demonstrated that overexpression of miR-4260 was associated with increased proliferation and migration of HUVEC using EdU incorporation assay (17.25%±1.31 vs 25.78%±1.24 in nc-mimics vs miR-4260 mimics, respectively) and wound healing assay, respectively. While downregulation of miR-4260 inhibited the proliferation (17.90%±1.37 vs 10.66%±1.41 in nc-inhibitor vs miR-4260 inhibitor, respectively) and migration of HUVEC. Furthermore, we found that miR-4260 mimics increased (129.75±3.68 vs 147±3.13 in nc-mimics vs miR-4260 mimics, respectively), while miR-4260 inhibitor decreased the tube formation of HUVECs in vitro (123.25±2.17 vs 92±4.45 in nc-inhibitor vs miR-4260 inhibitor expression, respectively). Our data indicate that miR-4260 contributes to the proliferation, migration and tube formation of endothelial cells, and might be essential regulators for angiogenesis. Further study is needed to investigate the underlying mechanism that mediates the role of miR-4260 in angiogenesis by identifying its putative downstream target genes.

scholarly journals Suppression of Angiogenesis by Targeting Cyclin-Dependent Kinase 7 in Human Umbilical Vein Endothelial Cells and Renal Cell Carcinoma: An In Vitro and In Vivo Study

Cells ◽  
2019 ◽  
Vol 8 (11) ◽  
pp. 1469 ◽  
Author(s):  
Chung-Sheng Shi ◽  
Kuan-Lin Kuo ◽  
Mei-Sin Chen ◽  
Po-Ming Chow ◽  
Shing-Hwa Liu ◽  
...  
Keyword(s):  
Renal Cell Carcinoma ◽  
Endothelial Cells ◽  
Cell Carcinoma ◽  
Renal Cell ◽  
Umbilical Vein ◽  
Tube Formation ◽  
Antitumor Effects ◽  
Angiogenic Activity ◽  
Human Umbilical Vein ◽  
Umbilical Vein Endothelial Cells

Cancer cells rely on aberrant transcription for growth and survival. Cyclin-dependent kinases (CDKs) play critical roles in regulating gene transcription by modulating the activity of RNA polymerase II (RNAPII). THZ1, a selective covalent inhibitor of CDK7, has antitumor effects in several human cancers. In this study, we investigated the role and therapeutic potential of CDK7 in regulating the angiogenic activity of endothelial cells and human renal cell carcinoma (RCC). Our results revealed that vascular endothelial growth factor (VEGF), a critical activator of angiogenesis, upregulated the expression of CDK7 and RNAPII, and the phosphorylation of RNAPII at serine 5 and 7 in human umbilical vein endothelial cells (HUVECs), indicating the transcriptional activity of CDK7 may be involved in VEGF-activated angiogenic activity of endothelium. Furthermore, through suppressing CDK7 activity, THZ1 suppressed VEGF-activated proliferation and migration, as well as enhanced apoptosis of HUVECs. Moreover, THZ1 inhibited VEGF-activated capillary tube formation and CDK7 knockdown consistently diminished tube formation in HUVECs. Additionally, THZ1 reduced VEGF expression in human RCC cells (786-O and Caki-2), and THZ1 treatment inhibited tumor growth, vascularity, and angiogenic marker (CD31) expression in RCC xenografts. Our results demonstrated that CDK7-mediated transcription was involved in the angiogenic activity of endothelium and human RCC. THZ1 suppressed VEGF-mediated VEGFR2 downstream activation of angiogenesis, providing a new perspective for antitumor therapy in RCC patients.

Degradation of soluble VEGF receptor-1 by MMP-7 allows VEGF access to endothelial cells

Blood ◽  
2009 ◽  
Vol 113 (10) ◽  
pp. 2363-2369 ◽  
Author(s):  
Ta-Kashi Ito ◽  
Genichiro Ishii ◽  
Seiji Saito ◽  
Keiichi Yano ◽  
Ayuko Hoshino ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Umbilical Vein ◽  
Critical Role ◽  
Inhibitory Effect ◽  
Tube Formation ◽  
Membrane Receptors ◽  
Vegf Receptor ◽  
Migration Assay ◽  
Vegf Inhibitor ◽  
Umbilical Vein Endothelial Cells

AbstractVascular endothelial growth factor (VEGF) signaling in endothelial cells serves a critical role in physiologic and pathologic angiogenesis. Endothelial cells secrete soluble VEGF receptor-1 (sVEGFR-1/sFlt-1), an endogenous VEGF inhibitor that sequesters VEGF and blocks its access to VEGF receptors. This raises the question of how VEGF passes through this endogenous VEGF trap to reach its membrane receptors on endothelial cells, a step required for VEGF-driven angiogenesis. Here, we show that matrix metalloproteinase-7 (MMP-7) degrades human sVEGFR-1, which increases VEGF bioavailability around the endothelial cells. Using a tube formation assay, migration assay, and coimmunoprecipitation assay with human umbilical vein endothelial cells (HUVECs), we show that the degradation of sVEGFR-1 by MMP-7 liberates the VEGF165 isoform from sVEGFR-1. The presence of MMP-7 abrogates the inhibitory effect of sVEGFR-1 on VEGF-induced phosphorylation of VEGF receptor-2 on HUVECs. These data suggest that VEGF escapes the sequestration by endothelial sVEGFR-1 and promotes angiogenesis in the presence of MMP-7.

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