scholarly journals Silymarin Ameliorates Diabetes-Induced Proangiogenic Response in Brain Endothelial Cells through a GSK-3β Inhibition-Induced Reduction of VEGF Release

2017 ◽  
Vol 2017 ◽  
pp. 1-9 ◽  
Author(s):  
Ahmed Alhusban ◽  
Enaam Alkhazaleh ◽  
Tamam El-Elimat
Keyword(s):  
Endothelial Cells ◽  
Glycogen Synthase ◽  
Antioxidant Properties ◽  
Tube Formation ◽  
Dependent Manner ◽  
Brain Endothelial Cells ◽  
Angiogenic Potential ◽  
Gsk 3Β ◽  
Vegf Release

Diabetes mellitus (DM) is a major risk factor for cardiovascular disease. Additionally, it was found to induce a dysfunctional angiogenic response in the brain that was attributed to oxidative stress. Milk thistle seed extract (silymarin) has potent antioxidant properties, though its potential use in ameliorating diabetes-induced aberrant brain angiogenesis is unknown. Glycogen synthase kinase-3β is a regulator of angiogenesis that is upregulated by diabetes. Its involvement in diabetes-induced angiogenesis is unknown. To evaluate the potential of silymarin to ameliorate diabetes-induced aberrant angiogenesis, human brain endothelial cells (HBEC-5i) were treated with 50 μg/mL advanced glycation end (AGE) products in the presence or absence of silymarin (50, 100 μM). The angiogenic potential of HBEC-5i was evaluated in terms of migration and in vitro tube formation capacities. The involvement of GSK-3β was also evaluated. AGE significantly increased the migration and tube formation rates of HBEC-5i by about onefold (p=0.0001). Silymarin reduced AGE-induced migration in a dose-dependent manner where 50 μM reduced migration by about 50%, whereas the 100 μM completely inhibited AGE-induced migration. Similarly, silymarin 50 μg/mL blunted AGE-induced tube formation (p=0.001). This effect was mediated through a GSK-3β-dependent inhibition of VEGF release. In conclusion, silymarin inhibits AGE-induced aberrant angiogenesis in a GSK-3β-mediated inhibition of VEGF release.

Accutin, a New Disintegrin, Inhibits Angiogenesis In Vitro and In Vivo by Acting as Integrin vβ3 Antagonist and Inducing Apoptosis

Blood ◽  
1998 ◽  
Vol 92 (9) ◽  
pp. 3268-3276 ◽  
Author(s):  
Chia Hsin Yeh ◽  
Hui-Chin Peng ◽  
Tur-Fu Huang
Keyword(s):  
Endothelial Cells ◽  
Umbilical Vein ◽  
Fluorescein Isothiocyanate ◽  
Adenosine Diphosphate ◽  
Tube Formation ◽  
Platelet Glycoprotein ◽  
Dependent Manner ◽  
Antiangiogenic Effect

Abstract Endothelial integrins play an essential role in angiogenesis and cell survival. Accutin, a new member of disintegrin family derived from venom of Agkistrodon acutus, potently inhibited human platelet aggregation caused by various agonists (eg, thrombin, collagen, and, adenosine diphosphate [ADP]) through the blockade of fibrinogen binding to platelet glycoprotein IIb/IIIa (ie, integrin IIbβ3). In this report, we describe that accutin specifically inhibited the binding of monoclonal antibody (MoAb) 7E3, which recognizes integrin vβ3, to human umbilical vein endothelial cells (HUVECs), but not those of other anti-integrin MoAbs such as 2β1, 3β1, and 5β1. Moreover, accutin, but not the control peptide GRGES, dose-dependently inhibited the 7E3 interaction with HUVECs. Both 7E3 and GRGDS, but not GRGES or Integrelin, significantly blocked fluorescein isothiocyanate-conjugated accutin binding to HUVEC. In functional studies, accutin exhibited inhibitory effects on HUVEC adhesion to immobilized fibrinogen, fibronectin and vitronectin, and the capillary-like tube formation on Matrigel in a dose- and RGD-dependent manner. In addition, it exhibited an effective antiangiogenic effect in vivo when assayed by using the 10-day-old embryo chick CAM model. Furthermore, it potently induced HUVEC apoptotic DNA fragmentation as examined by electrophoretic and flow cytometric assays. In conclusion, accutin inhibits angiogenesis in vivo and in vitro by blocking integrin vβ3 of endothelial cells and by inducing apoptosis. The antiangiogenic activity of disintegrins might be explored as the target of developing the potential antimetastatic agents. © 1998 by The American Society of Hematology.

scholarly journals P11.30 Special VEGF90kDa form in exosome from GBM cells activates TAZ in endothelial cells to promote angiogenesis and contributes bevacizumab resistance

2019 ◽  
Vol 21 (Supplement_3) ◽  
pp. iii49-iii49
Author(s):  
Z Wang ◽  
Y Yuan ◽  
C Xu ◽  
Y Liu
Keyword(s):  
Endothelial Cells ◽  
Western Blotting ◽  
Umbilical Vein ◽  
Tube Formation ◽  
Cellular Migration ◽  
Vegf Expression ◽  
Angiogenic Potential ◽  
Huvec Cells

Abstract BACKGROUND Glioblastoma (GBM) has obvious blood vessels proliferation, which is one of the important histological diagnostic criteria. Bevacizumab, VEGF targeted inhibitor, is found inefficient in 2/3 cases after the clinical trial. Here, we found a specific 90kDa form of VEGF (VEGF90K) in exosomes of GBM could activate TAZ expression in endothelial cells to promote angiogenesis, which might also contribute to Bevacizumab treatment resistance. MATERIAL AND METHODS Exosomes were isolated from U87, LN229 GBM cell culture medium and characterized using transmission electron microscopy, nanoparticle analysis, and western blotting. VEGF expression in GBM and TAZ expression in human umbilical vein endothelial cells (HUVEC) are inhibited by shRNA. In vitro migration, proliferation, and tube formation assays were used to assess the angiogenic potential of HUVEC cells. Western blotting was used to analysis TAZ expression in HUVEC. Bevacizumab or exosome inhibitor GW4869 were used separately or together to evaluation their blocking effects on angiogenic functions of GBM cells. RESULTS we found a unique 90kDa form of VEGF (VEGF90K) from exosome of GBM cells could activate TAZ expression in HUVEC cells which correlated with higher levels of cellular migration, proliferation, and tube formation. Knockdown VEGF expression in GBM exosomes or TAZ expression in HUVEC cells could decrease the angiogenic function of GBM cells. GW4869 combine Bevacizumab had significantly inhibited the angiogenic ability of GBM cells in vivo and in vitro. CONCLUSION A specific exosome-associated VEGF from GBM can promote angiogenesis by activating TAZ expression in endothelial cells. Targeted inhibition of exosome secretion can significantly inhibit tumor angiogenesis and restore bevacizumab effect. Our study highlights the unique properties of VEGF in exosome which might help to explore new treatment strategies of GBM.

scholarly journals MicroRNA-126 enhances the biological function of endothelial progenitor cells under oxidative stress via PI3K/Akt/GSK-3β and ERK1/2 signaling pathways

2020 ◽  
Author(s):  
Qinqin Wu ◽  
Benling Qi ◽  
Xiaoyu Duan ◽  
Xiaoyan Ming ◽  
Fengqin Yan ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Biological Function ◽  
Glycogen Synthase ◽  
Early Stage ◽  
Tube Formation ◽  
Dependent Manner ◽  
Endothelial Progenitor ◽  
H2o2 Treatment ◽  
Reverse Transcription Pcr ◽  
Gsk 3Β

Endothelial progenitor cell (EPC) transplantation is a safe and effective method to treat acute myocardial infarction (AMI). However, oxidative stress leads to the death of a large number of EPCs in the early stage of transplantation, severely weakening the therapeutic effect. Previous studies demonstrated that microRNAs (miRNAs) regulate the biological function of EPCs. The aim of the current study was to investigate the effect of miRNA on the biological function of EPCs under oxidative stress. Quantitative reverse transcription PCR was performed to detect the expression of miR-126, miR-508-5p, miR-150, and miR-16 in EPCs from rats, among which miR-126 showed a relatively higher expression. Treatment with H2O2 decreased miR-126 expression in EPCs in a dose-dependent manner. EPCs were further transfected with miR-126 mimics or inhibitors, followed by H2O2 treatment. Overexpression of miR-126 enhanced the proliferation, migration, and tube formation of H2O2-treated EPCs. MiR-126 overexpression also inhibited reactive oxygen species and malondialdehyde levels and enhanced superoxide dismutase levels, as well as increased angiopoietin (Ang)1 expression and decreased Ang2 expression in H2O2-treated EPCs. Moreover, miR-126 participated in the regulation of phosphatidylinositol 3-kinase (PI3K)/ protein kinase B (Akt)/ glycogen synthase kinase-3β (GSK-3β) and extracellular signal-regulated kinase (ERK)1/2 signaling in EPCs, where both pathways were activated after miR-126 overexpression in H2O2-treated EPCs. Overall, we showed that miR-126 promoted the biological function of EPCs under H2O2-induced oxidative stress by activating the PI3K/Akt/GSK-3β and ERK1/2 signaling pathway, which may serve as a new therapeutic approach to treat AMI.

scholarly journals NDRG1 Targets TAF15 to Promote Endothelial Dysfunction and Hypoxia-Induced Pulmonary Hypertension

2021 ◽  
Author(s):  
Chengwei Li ◽  
Liang Dong ◽  
Ning Zhu ◽  
Xiujuan Zhang ◽  
Ruzetuoheti Yiminniyaze ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Pulmonary Hypertension ◽  
Endothelial Dysfunction ◽  
Vascular Remodeling ◽  
Tube Formation ◽  
Cell Counting ◽  
Dependent Manner ◽  
Rat Models ◽  
Chain Reactions

Abstract Background: The mechanism underlying vascular remodeling of hypoxia-induced pulmonary hypertension (HPH) is not fully elucidated. We hypothesized that hypoxia promotes expression of N-myc downstream regulated gene-1 (NDRG1) in human pulmonary arterial endothelial cells (HPAECs), which in turn leads to endothelial dysfunction and contributing to HPH. Methods: Lung samples were obtained from qualified patients and HPH rat models. Quantitative polymerase chain reactions, western blotting and immunohistochemistry were used to measure the expression of NDRG1. EdU incorporation assays, cell counting kit-8 (CCK-8) assays, transwell migration assays, and matrigel assays were conducted to detect the role of NDRG1 in HPACE function in vitro. HPH models were established in SD rats and were treated with plasmids expressing short hairpin RNAs (shRNAs) to silence NDRG1. The candidate binding partner(s) of NDRG1 was screened and validated via co-immunoprecipitation and immunofluorescence staining. Results: NDRG1 is up-regulated by hypoxia in a time-dependent manner in HPAECs. Expression of NDRG1 was increased in lung tissues of HPH patient and rat model. In vitro, silencing NDRG1 attenuated proliferation, migration and tube formation of HPAECs under hypoxia, while NDRG1 over-expression promoted these behaviors of HPAECs in normoxia. NDRG1 knock-down alleviated vascular remodeling and right ventricular hypertrophy in rat models of HPH. NDRG1 can directly interact with TATA-box binding protein associated factor 15 (TAF15) and promote its nuclear localization. Bioinformatics study found that Notch1 signaling was downstream of TAF15 in endothelial cells. TAF15 can promote HPAECs dysfunction via binding to Notch1 promoter region and subsequently increasing Notch1 expression. Conclusions: Taken together, hypoxia-induced up-regulation of NDRG1 contributes to endothelial dysfunction and HPH development through TAF15 upregulation of Notch1, suggesting the applicability of targeting NDRG1 in clinical treatment of HPH.

Abstract 195: Simvastatin Rescues Impaired Angiogenesis Potential of Cerebrovascular Endothelial Cells by AGEs : The Role of Autophagy

Stroke ◽  
2014 ◽  
Vol 45 (suppl_1) ◽  
Author(s):  
Xuwei Hou ◽  
Zhaohui Hu ◽  
Ningfu Wang
Keyword(s):  
Endothelial Cells ◽  
Tube Formation ◽  
Western Blot Assay ◽  
Angiogenic Potential ◽  
Cellular Autophagy ◽  
Glycation End Products ◽  
Angiogenic Effect ◽  
Cerebrovascular Endothelial Cells ◽  
High Level

Background: Statin therapy is beneficial for ischemic stroke (IS) partly due to its pro-angiogenic effect. In diabetes patients, the accumulations of advanced glycation end products (AGEs) impair the angiogenic potential of endothelial cells (ECs), comprising collateral circulation in ischemic area. Autophagy affects endothelial function. The effect of statins on EC autophagy remains largely unknown. This study was to: 1.investigate the effect of Statin on the angiogenesis ability of ECs impaired by AGEs; 2. explore the autophagy of ECs in this process. Methods: primary cerebromicrovascular endothelial cells (CMVECs) were isolated from Wistar rats and cultured in medium. CMVECs were treated with AGEs. Subsequently, Simvastatin was added to cells. The angiogenic potential was evaluated by tube formation in vitro. Two autophagy marker, Beclin1 and LC3 protein expression were detected by western blot assay. The autophagosome quantitative analysis was evaluated by GFP-LC3-II assay. Results: AGEs (25mg/L, 50mg/L and 100mg/L) reduced the number of tube like structures in a dose dependant manner, accompanied by dramatically enhanced cellular autophagy level indicated by the increased GFP-LC3-II puncta number/cell and increased Beclin1 and LC3 expressions. Autophagy inhibitor 3-MA reversed the AGEs impaired angiogenic ability of CMVES. Simvastatin administration (10μmol/L) markedly increased the angiogenic potential of CMVES evaluated by tube formation ability, together with decreased Beclin1 and LC3, as well as decreased GFP-LC3-II puncta number/cell. AGEs significantly inhibits the PI3K/Akt/mTOR, while Simvastatin re-activate this pathway. More interestingly, the addition of mTOR inhibitor, rapamycin, which enhance the cellular autophagy, abolished the rescue effect of Simvastatin on the angiogenic potential of CMVECs impaired by AGEs. Conclusion: This study provides evidence that AGEs impair the angiogenesis ability via triggering high level of autophagy of CMVECs. Simvastatin rescue the angiogenic ability of CMVECs by surprising autophagy level via activation of PI3K/Akt/mTOR pathway.

scholarly journals The cancer angiogenesis co-culture assay: In vitro quantification of the angiogenic potential of tumoroids

PLoS ONE ◽  
2021 ◽  
Vol 16 (7) ◽  
pp. e0253258
Author(s):  
Sarah Line Bring Truelsen ◽  
Nabi Mousavi ◽  
Haoche Wei ◽  
Lucy Harvey ◽  
Rikke Stausholm ◽  
...  
Keyword(s):  
Predictive Biomarkers ◽  
Tube Formation ◽  
Angiogenic Factor ◽  
Dependent Manner ◽  
Vascular Endothelial ◽  
Colorectal Tumour ◽  
Angiogenic Potential ◽  
Endothelial Growth Factor ◽  
Dose Dependent

The treatment response to anti-angiogenic agents varies among cancer patients and predictive biomarkers are needed to identify patients with resistant cancer or guide the choice of anti-angiogenic treatment. We present “the Cancer Angiogenesis Co-Culture (CACC) assay”, an in vitro Functional Precision Medicine assay which enables the study of tumouroid induced angiogenesis. This assay can quantify the ability of a patient-derived tumouroid to induce vascularization by measuring the induction of tube formation in a co-culture of vascular cells and tumoroids established from the primary colorectal tumour or a metastasis. Furthermore, the assay can quantify the sensitivity of patient-derived tumoroids to anti-angiogenic therapies. We observed that tube formation increased in a dose-dependent manner upon treatment with the pro-angiogenic factor vascular endothelial growth factor A (VEGF-A). When investigating the angiogenic potential of tumoroids from 12 patients we found that 9 tumoroid cultures induced a significant increase in tube formation compared to controls without tumoroids. In these 9 angiogenic tumoroid cultures the tube formation could be abolished by treatment with one or more of the investigated anti-angiogenic agents. The 3 non-angiogenic tumoroid cultures secreted VEGF-A but we observed no correlation between the amount of tube formation and tumoroid-secreted VEGF-A. Our data suggests that the CACC assay recapitulates the complexity of tumour angiogenesis, and when clinically verified, could prove a valuable tool to quantify sensitivity towards different anti-angiogenic agents.

scholarly journals ERK1/2 and HIF1αAre Involved in Antiangiogenic Effect of Polyphenols-Enriched Fraction from Chilean Propolis

2015 ◽  
Vol 2015 ◽  
pp. 1-11 ◽  
Author(s):  
Alejandro Cuevas ◽  
Nicolás Saavedra ◽  
Martina Rudnicki ◽  
Dulcineia S. P. Abdalla ◽  
Luis A. Salazar
Keyword(s):  
Endothelial Cells ◽  
Western Blot ◽  
Mrna Expression ◽  
Ethanolic Extract ◽  
Tube Formation ◽  
Dependent Manner ◽  
Antiangiogenic Effect ◽  
Dose Dependent

Propolis has been shown to modulate the angiogenesis in bothin vitroandin vivomodels. Thus, we aimed to evaluate the antiangiogenic properties of an ethanolic extract of Chilean propolis (EEP) and Pinocembrin (Pn). Migration, formation of capillary-like structures of endothelial cells, and sprouting from rat aortic rings were used to assess the antiangiogenic properties of EEP or Pn. In addition, microRNAs and VEGFA mRNA expression were studied by qPCR. ERK1/2 phosphorylation and HIF1αstabilization were assessed by western blot. EEP or Pn attenuated the migration, the capillary-like tube formation, and the sprouting in thein vitroassays. In addition, the activation of HIF1αand ERK1/2 and the VEGFA mRNA expression was significantly inhibited in a dose-dependent manner. In summary, these results suggest that HIF1αand ERK1/2 phosphorylation could be involved in the antiangiogenic effect of Chilean propolis, but more studies are needed to corroborate these findings.

Accutin, a New Disintegrin, Inhibits Angiogenesis In Vitro and In Vivo by Acting as Integrin vβ3 Antagonist and Inducing Apoptosis

Blood ◽  
1998 ◽  
Vol 92 (9) ◽  
pp. 3268-3276 ◽  
Author(s):  
Chia Hsin Yeh ◽  
Hui-Chin Peng ◽  
Tur-Fu Huang
Keyword(s):  
Endothelial Cells ◽  
Umbilical Vein ◽  
Fluorescein Isothiocyanate ◽  
Adenosine Diphosphate ◽  
Tube Formation ◽  
Platelet Glycoprotein ◽  
Dependent Manner ◽  
Antiangiogenic Effect

Endothelial integrins play an essential role in angiogenesis and cell survival. Accutin, a new member of disintegrin family derived from venom of Agkistrodon acutus, potently inhibited human platelet aggregation caused by various agonists (eg, thrombin, collagen, and, adenosine diphosphate [ADP]) through the blockade of fibrinogen binding to platelet glycoprotein IIb/IIIa (ie, integrin IIbβ3). In this report, we describe that accutin specifically inhibited the binding of monoclonal antibody (MoAb) 7E3, which recognizes integrin vβ3, to human umbilical vein endothelial cells (HUVECs), but not those of other anti-integrin MoAbs such as 2β1, 3β1, and 5β1. Moreover, accutin, but not the control peptide GRGES, dose-dependently inhibited the 7E3 interaction with HUVECs. Both 7E3 and GRGDS, but not GRGES or Integrelin, significantly blocked fluorescein isothiocyanate-conjugated accutin binding to HUVEC. In functional studies, accutin exhibited inhibitory effects on HUVEC adhesion to immobilized fibrinogen, fibronectin and vitronectin, and the capillary-like tube formation on Matrigel in a dose- and RGD-dependent manner. In addition, it exhibited an effective antiangiogenic effect in vivo when assayed by using the 10-day-old embryo chick CAM model. Furthermore, it potently induced HUVEC apoptotic DNA fragmentation as examined by electrophoretic and flow cytometric assays. In conclusion, accutin inhibits angiogenesis in vivo and in vitro by blocking integrin vβ3 of endothelial cells and by inducing apoptosis. The antiangiogenic activity of disintegrins might be explored as the target of developing the potential antimetastatic agents.© 1998 by The American Society of Hematology.

Effect of DNA damage response by quinazolinone analogue HMJ-38 on human umbilical vein endothelial cells

2013 ◽  
Vol 33 (6) ◽  
pp. 590-601 ◽  
Author(s):  
J-H Chiang ◽  
J-S Yang ◽  
C-C Lu ◽  
M-J Hour ◽  
K-C Liu ◽  
...  
Keyword(s):  
Dna Damage ◽  
Endothelial Cells ◽  
Dna Damage Response ◽  
Glycogen Synthase ◽  
Umbilical Vein ◽  
Human Umbilical Vein ◽  
Damage Response ◽  
Gsk 3Β ◽  
Umbilical Vein Endothelial Cells

The present study aims to explore the mechanism of quinazolinone analogue HMJ-38-induced DNA damage in endothelial cells in vitro. We attempt to evaluate the antiangiogenetic response utilizing human umbilical vein endothelial cells (HUVECs). Herein, the results demonstrated that HMJ-38 incubation triggered DNA damage behavior and showed a longer DNA migration in HUVECs based on the comet assay and the analysis of DNA agarose gel electrophoresis to contact DNA smears. We further gained to determine a marker of DNA double strand breaks, phosphorylated histone H2A.X (Ser139) (γH2A.X), in HMJ-38-treated HUVECs by flow cytometry and Western blotting assay. We consider that HMJ-38 has caused an increase in γH2A.X, and DNA damage seemed to mediate through DNA-dependent serine/threonine protein kinase (DNA-PK) binding to Ku70/Ku80 as well as advanced activated p-Akt (Ser473) and stimulated phosphorylated glycogen synthase kinase-3β (p-GSK-3β) conditions in HUVECs. Importantly, the effect of above DNA damage response was prevented by N-acetyl-l-cysteine (a reactive oxygen species scavenger), and NU7026 (a DNA-PK inhibitor) could attenuate DNA-PK catalytic subunit and phosphorylation of H2A.X on Ser139 expression in comparison with HMJ-38 alone treated HUVECs. Therefore, HMJ-38-provoked DNA damage stress in HUVECs probably led to the activation of γH2A.X/DNA-PK/GSK-3β signaling. In summary, our novel finding provides more information addressing the pharmacological approach of newly synthesized HMJ-38 for further development and therapeutic application in antiangiogenetic effect of cancer chemotherapy.

MG53 inhibits angiogenesis through regulating focal adhesion kinase signaling

2021 ◽  
Author(s):  
Jinling Dong ◽  
Haiyan Zhou ◽  
Yongjie Li ◽  
Rong Li ◽  
Ni Chen ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Focal Adhesion Kinase ◽  
Signaling Pathways ◽  
Focal Adhesion ◽  
Cell Function ◽  
Striated Muscle ◽  
Tube Formation ◽  
Dependent Manner ◽  
Endothelial Cell Function

Abstract Mitsugumin 53 (MG53), which is expressed predominantly in striated muscle, has been demonstrated to be a myokine/cardiokine secreted from striated muscle under specific conditions. The important roles of MG53 in non-striated muscle tissues have also been examined in multiple disease models. However, no previous study has implicated MG53 in the control of endothelial cell function. In order to explore the effects of MG53 on endothelial cells, human umbilical vein endothelial cells (HUVECs) were stimulated with recombinant human MG53 (rhMG53). Then rhMG53 uptake, focal adhesion kinase (FAK)/Src/Akt/ERK1/2 signaling pathway activation, cell migration and tube formation were determined in vitro. The efficacy of rhMG53 in regulating angiogenesis was also detected in postnatal mouse retinas. The results demonstrated that rhMG53 directly entered into endothelial cells in a cholesterol-dependent manner. The uptake rhMG53 directly bound to FAK in endothelial cells, which resulted in significant decrease of FAK phosphorylation at Y397. Accompanied by the dephosphorylation of FAK, rhMG53 uncoupled FAK-Src interaction and reduced the phosphorylation of Src at Y416. Consequently, the activation of FAK/Src downstream signaling pathways, such as Akt and ERK1/2, was also significantly inhibited by rhMG53. Furthermore, rhMG53 remarkably decreased HUVEC migration and tube formation in vitro and postnatal mouse retinal angiogenesis in vivo. Taken together, these data indicate that rhMG53 inhibits angiogenesis through regulating FAK/Src/Akt/ERK1/2 signaling pathways. This may provide a novel molecular mechanism for the impaired angiogenesis in ischemic diseases.

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