scholarly journals Endothelial Cell Tube Formation Assay for the In Vitro Study of Angiogenesis

10.3791/51312 ◽  
2014 ◽  
Author(s):  
Katie L. DeCicco-Skinner ◽  
Gervaise H. Henry ◽  
Christophe Cataisson ◽  
Tracy Tabib ◽  
J. Curtis Gwilliam ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Tube Formation ◽  
Tube Formation Assay

DARC Regulates Angiogenesis by Mediating Vascular Endothelial Cell Migration

2020 ◽  
Author(s):  
Xiaolin Wang ◽  
Yongqian Bian ◽  
Yuejun Li ◽  
Jing Li ◽  
Congying Zhao ◽  
...  
Keyword(s):  
Cell Migration ◽  
Endothelial Cell ◽  
Vascular Endothelial Cell ◽  
Tube Formation ◽  
Endothelial Cell Migration ◽  
Immunofluorescent Staining ◽  
Control Group ◽  
Rhoa Activation ◽  
And Control

Abstract Background: DARC (The Duffy antigen receptor for chemokines) is a kind of glycosylated membrane protein that binds to members of the CXC chemokine family associated with angiogenesis and has recently been reported to be implicated in diverse normal physiologic processes. This study aimed to investigate the involvement of DARC in angiogenesis, which is known to generate new capillary blood vessels from preexisting ones. Methods: HDMECs (Human dermal microvascular endothelial cells) were divided into two groups (DARC overexpression group, and control group). We used Brdu staining to detect cell proliferation, and wound healing assay to detect cell migration. Then tube formation assay were observed. Also, western blot and immunofluorescent staining were used to estimate the relationship between DARC and RhoA (Ras homolog gene family, member A). Results: HDMECs proliferation, migration, and tube formation were inhibited significantly when DARC was overexpressed intracellular. DARC impaired microfilament dynamics and intercellular connection in migrating cells, and RhoA activation underlay the effect of DARC on endothelial cell. Furthermore, DARC inhibited the formation of new capillaries in vitro. Conclusion: Our findings revealed the role of DARC in the angiogenic process and provided a novel mechanism for RhoA activation during endothelial cell migration and angiogenesis.

scholarly journals Intrauterine growth restriction decreases pulmonary alveolar and vessel growth and causes pulmonary artery endothelial cell dysfunction in vitro in fetal sheep

2011 ◽  
Vol 301 (6) ◽  
pp. L860-L871 ◽  
Author(s):  
Paul J. Rozance ◽  
Gregory J. Seedorf ◽  
Alicia Brown ◽  
Gates Roe ◽  
Meghan C. O'Meara ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Cell Growth ◽  
Pulmonary Artery ◽  
Intrauterine Growth Restriction ◽  
Tube Formation ◽  
No Production ◽  
Vascular Growth ◽  
Intrauterine Growth ◽  
Pulmonary Artery Endothelial Cell

Intrauterine growth restriction (IUGR) increases the risk for bronchopulmonary dysplasia (BPD). Abnormal lung structure has been noted in animal models of IUGR, but whether IUGR adversely impacts fetal pulmonary vascular development and pulmonary artery endothelial cell (PAEC) function is unknown. We hypothesized that IUGR would decrease fetal pulmonary alveolarization, vascular growth, and in vitro PAEC function. Studies were performed in an established model of severe placental insufficiency and IUGR induced by exposing pregnant sheep to elevated temperatures. Alveolarization, quantified by radial alveolar counts, was decreased 20% ( P < 0.005) in IUGR fetuses. Pulmonary vessel density was decreased 44% ( P < 0.01) in IUGR fetuses. In vitro, insulin increased control PAEC migration, tube formation, and nitric oxide (NO) production. This response was absent in IUGR PAECs. VEGFA stimulated tube formation, and NO production also was absent. In control PAECs, insulin increased cell growth by 68% ( P < 0.0001). Cell growth was reduced in IUGR PAECs by 29% at baseline ( P < 0.01), and the response to insulin was attenuated ( P < 0.005). Despite increased basal and insulin-stimulated Akt phosphorylation in IUGR PAECs, endothelial NO synthase (eNOS) protein expression as well as basal and insulin-stimulated eNOS phosphorylation were decreased in IUGR PAECs. Both VEGFA and VEGFR2 also were decreased in IUGR PAECs. We conclude that fetuses with IUGR are characterized by decreased alveolar and vascular growth and PAEC dysfunction in vitro. This may contribute to the increased risk for adverse respiratory outcomes and BPD in infants with IUGR.

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.

scholarly journals Roundabout4 Suppresses Glioma-Induced Endothelial Cell Proliferation, Migration and Tube Formation in Vitro by Inhibiting VEGR2-Mediated PI3K/AKT and FAK Signaling Pathways

2015 ◽  
Vol 35 (5) ◽  
pp. 1689-1705 ◽  
Author(s):  
Heng Cai ◽  
Yixue Xue ◽  
Zhen Li ◽  
Yi Hu ◽  
Zhenhua Wang ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Signaling Pathways ◽  
Conditioned Medium ◽  
Tube Formation ◽  
Endothelial Cell Proliferation ◽  
Akt Inhibitor ◽  
Over Expression

Background and Aims: Endothelial cell (EC) proliferation, migration, and tube formation are the critical steps for tumor angiogenesis, which is involved in the formation of new tumor blood vessels. Roundabout4 (Robo4), a new member of Robo proteins family, is specifically expressed in endothelial cells. This study aimed to investigate the effects of Robo4 on glioma-induced endothelial cell proliferation, migration and tube formation in vitro. Methods and Results: We found that Robo4 was endogenously expressed in Human Brain Microvascular Endothelial Cells (HBMECs), while Robo4 was significantly down-regulated in endothelial cells cultured in glioma conditioned medium. Robo4 over-expression remarkably suppressed glioma-induced endothelial cell proliferation, migration and tube formation in vitro. In addition, Robo4 influenced the glioma-induced angiogenesis via binding to its ligand Slit2. Further studies demonstrated that the knockdown of Robo4 up-regulated the phosphorylation of VEGFR2, PI3K, AKT and FAK in EC cultured in glioma conditioned medium. VEGFR2 inhibitor SU-1498, AKT inhibitor LY294002 and FAK inhibitor 14 (FAK inhibitor) blocked the Robo4 knockdown-mediated alteration in glioma angiogenesis in vitro. Conclusion: Our results proved that Robo4 suppressed glioma-induced endothelial cell proliferation, migration and tube formation in vitro by inhibiting VEGR2-mediated activation of PI3K/AKT and FAK signaling pathways.

scholarly journals Vascular endothelium protective property of curcumin

2021 ◽  
Vol 30 (2) ◽  
pp. 121-126
Author(s):  
Serdar Bayrak ◽  
Gizem Çalıbaşı-Koçal ◽  
Feriha Toksöz ◽  
Tuğra Gençpınar ◽  
Nazlı Mert Özüpek ◽  
...  
Keyword(s):  
Wound Healing ◽  
Cell Line ◽  
Umbilical Vein ◽  
Tube Formation ◽  
Control Group ◽  
Protective Agent ◽  
Vascular Protection ◽  
Tube Formation Assay ◽  
Huvec Cell

Objectives: In the present study, we aimed to examine the efficacy of curcumin on endothelial vascular protection on human umbilical vein endothelial cells (HUVEC) cells in the in vitro setting and to determine effective doses. Materials and methods: Cytotoxic effect and wound healing activity of curcumin on the HUVEC cell line were evaluated and angiogenesis was studied by tube formation assay. The cytotoxic activity of 0.5 to 8 μM curcumin on HUVEC cell line was tested using the WST-1 cell proliferation assay. In wound healing determination, the scratched cells were incubated with the half maximal effective concentration (EC50) dose of curcumin and wounds were monitored by the JuLITM Br live cell movie analyzer. Wound gaps were measured using the ImageJ software. To determine the angiogenesis, tube formation assay was performed and the results were analyzed. Results: The 1.77 μM of curcumin increased the cell viability by 150% after 48 h of treatment. According to the wound healing results, after 48 h of incubation, the control group and 1.77 μM curcumin exhibited 47.1% and 43.8% closure, respectively. The mean maximum and minimum tube lengths were found to be 21,858±3,945 and 12,438±3,817 pixel for curcumin, respectively with low fetal bovine serum (FBS) and curcumin with high FBS, respectively. Conclusion: Our results show that curcumin is a promising endothelial protective agent for HUVEC cell line.

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