AGE/RAGE-Induced EMP Release via the NOX-Derived ROS Pathway

Objective. Diabetes is associated with accelerated formation of advanced glycation end products (AGEs) that are extensively found in circulating endothelial microparticles (EMPs). This study aimed to investigate whether AGEs have a direct effect on EMP formation and the possible underlying mechanism. Methods. In vitro, cultured human umbilical vein endothelial cells (HUVECs) were incubated with AGEs (200 and 400 μg/ml) for 24 hours with or without pretreatment with anti-RAGE antibody, NOX inhibitor, or ROS scavenger. The number of CD31-positive EMPs was assessed by flow cytometry. Results. The number of EMPs was significantly increased in HUVECs stimulated by AGEs in a dose-dependent manner. In addition, receptors for AGEs (RAGE), NAD(P)H oxidase (NOX), and reactive oxygen species (ROS) were increased by AGEs as compared to the control group. These changes could be reversed when HUVECs were pretreated with anti-RAGE antibody. Moreover, inhibition of NOX as well as antioxidant treatment reduced the release of EMPs induced by AGEs. Conclusion. Our study suggested that AGEs increased EMP generation, which was mediated by RAGE signaling through NOX-derived ROS.

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Ethyl Acetate Fraction of Teucrium Polium Extract Abolishes Human Umbilical Vein Endothelial Cells (HUVEC) Tubulogenesis in Collagen Bed through Suppression of Cell Proliferation/VEGF Secretion

Iranian Journal of Allergy Asthma and Immunology ◽

10.18502/ijaai.v18i3.1121 ◽

2019 ◽

Cited By ~ 1

Author(s):

Vahide Askari ◽

Somayeh Shamlou ◽

Ali Mostafaie ◽

Sara Khaleqi

Keyword(s):

Endothelial Cells ◽

Ethyl Acetate ◽

Umbilical Vein ◽

Ethyl Acetate Fraction ◽

Dependent Manner ◽

Angiogenic Effect ◽

Teucrium Polium ◽

Umbilical Vein Endothelial Cells ◽

Dose Dependent

Angiogenesis has essential role in growth and metastasis of tumors. Development of therapies aimed to suppress angiogenesis using medicinal plants is one of the effective approaches for prevention/treatment of cancer. The current study was performed to investigate in vitro anti-angiogenic effect of Teucrium Polium (TP) extract and its fractions. The aerial part of Teucrium Polium was powdered and extracted with 50% ethanol. The extract was fractionated in to aqueous (AQ), n-butanol (BU), ethyl acetate (EA) and n-hexane (HE) fractions. Anti-angiogenic effect of TP was examined on human umbilical vein endothelial cells (HUVECs) in three-dimensional collagen matrix. The endothelial cells form capillary-like branches that can be visualized using phase contrast microscope and the number of tube-like structures can be quantified as a measure of in vitro angiogenesis. Furthermore, anti-proliferative and vascular endothelial growth factor(VEGF )suppressive effect of TP as important factors in the process of angiogenesis were assessed using3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT)and quantitative ELISA, respectively. Based on our findings, among the TP fractions, EA fraction showed the highest inhibitory activity on angiogenesis. This fraction with IC50: 68 µg/mL, inhibited angiogenesis at 60 µg/mL. The crude extract and other fractions of TP inhibited angiogenesis in a dose-dependent manner at doses higher concentrations than EA fraction, significantly.TP extract and EA fraction were able to inhibit proliferation of HUVEC and inhibited VEGF secretion in a dose dependent manner. The ethyl acetate fraction at 60 µg/ml inhibited VEGF secretion perfectly. Our data indicated that ethyl acetate fraction of Teucrium Polium could be a potential candidate for the prevention of angiogenesis in cancer and other related disorders. However, this suggestion needs more quantitative and in vivo investigations for confirmation.

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PPAR-αAgonist Fenofibrate Upregulates Tetrahydrobiopterin Level through Increasing the Expression of Guanosine 5′-Triphosphate Cyclohydrolase-I in Human Umbilical Vein Endothelial Cells

PPAR Research ◽

10.1155/2011/523520 ◽

2011 ◽

Vol 2011 ◽

pp. 1-8 ◽

Cited By ~ 12

Author(s):

Jinbo Liu ◽

Changlin Lu ◽

Fuwang Li ◽

Haining Wang ◽

Liyun He ◽

...

Keyword(s):

Endothelial Cells ◽

Umbilical Vein ◽

Control Group ◽

Peroxisome Proliferator ◽

Dependent Manner ◽

Concentration Dependent Manner ◽

Peroxisome Proliferator Activated Receptor ◽

Human Umbilical Vein ◽

Umbilical Vein Endothelial Cells

Tetrahydrobiopterin (BH4) is an essential cofactor for endothelial nitric oxide (NO) synthase. Guanosine 5′-triphosphate cyclohydrolase-I (GTPCH-I) is a key limiting enzyme for BH4 synthesis. In the present in vitro study, we investigated whether peroxisome proliferator-activated receptorα(PPAR-α) agonist fenofibrate could recouple eNOS by reversing low-expression of intracellular BH4 in endothelial cells and discussed the potential mechanisms. After human umbilical vein endothelial cells (HUVECs) were treated with lipopolysaccharide (LPS) for 24 hours, the levels of cellular eNOS, BH4 and cell supernatant NO were significantly reduced compared to control group. And the fluorescence intensity of intracellular ROS was significantly increased. But pretreated with fenofibrate (10 umol/L) for 2 hours before cells were induced by LPS, the levels of eNOS, NO, and BH4 were significantly raised compared to LPS treatment alone. ROS production was markedly reduced in fenofibrate group than LPS group. In addition, our results showed that the level of intracellular GTPCH-I detected by western blot was increased in a concentration-dependent manner after being treated with fenofibrate. These results suggested that fenofibrate might help protect endothelial function and against atherosclerosis by increasing level of BH4 and decreasing production of ROS through upregulating the level of intracellular GTPCH-I.

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Shear-dependent inhibition of granulocyte adhesion to cultured endothelium by dextran sulfate

Blood ◽

10.1182/blood.v73.5.1324.bloodjournal7351324 ◽

1989 ◽

Vol 73 (5) ◽

pp. 1324-1330

Author(s):

K Ley ◽

E Lundgren ◽

E Berger ◽

KE Arfors

Keyword(s):

Shear Stress ◽

Dextran Sulfate ◽

Leukocyte Adhesion ◽

Umbilical Vein ◽

Dependent Manner ◽

Dependent Inhibition ◽

Umbilical Vein Endothelial Cells ◽

Dose Dependent ◽

Adhesion Process

Adhesion of polymorphonuclear granulocytes (PMNs) in microvessels occurs in the presence of shear forces exerted by the blood flow. To model this in vitro, phorbol myristate acetate (PMA)-activated PMN were exposed to shear stress on cultured human umbilical vein endothelial cells (HUVECs) and on plastic dishes coated with bovine serum albumin (BSA). PMN adhesion to HUVECs averaged 36% of the total PMNs added and was reduced to 21% by shear stress of approximately 1.5 dynes.cm-2. On BSA, adhesion was reduced from 59% to 35%. Dextran sulfate (molecular weight 500,000) inhibited PMN adhesion in a dose-dependent manner when shear stress was applied. At a concentration of 1 mg.ml-1, inhibition was 72% on HUVECs and 76% on BSA. Half-maximal inhibition was reached at approximately 1 microgram.mL-1 dextran sulfate, corresponding to 2 nmol/L. Without shear stress, dextran sulfate had no effect on HUVECs and only a moderate effect on BSA. The murine monoclonal antibody (MoAb) 60.3, recognizing an epitope on the leukocyte adhesion glycoprotein CD18, inhibited PMN adhesion equally well with and without shear. A low dose of MoAb 60.3 enhanced the effect of dextran sulfate without shear stress. Flow cytometry (FACS) did not show inhibition of MoAb 60.3 binding to PMNs by dextran sulfate. These results indicate that a dextran sulfate-inhibitable adhesion process is important for PMN adhesion in the presence of shear stress.

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Shear-dependent inhibition of granulocyte adhesion to cultured endothelium by dextran sulfate

Blood ◽

10.1182/blood.v73.5.1324.1324 ◽

1989 ◽

Vol 73 (5) ◽

pp. 1324-1330 ◽

Cited By ~ 78

Author(s):

K Ley ◽

E Lundgren ◽

E Berger ◽

KE Arfors

Keyword(s):

Shear Stress ◽

Dextran Sulfate ◽

Leukocyte Adhesion ◽

Umbilical Vein ◽

Dependent Manner ◽

Dependent Inhibition ◽

Umbilical Vein Endothelial Cells ◽

Dose Dependent ◽

Adhesion Process

Abstract Adhesion of polymorphonuclear granulocytes (PMNs) in microvessels occurs in the presence of shear forces exerted by the blood flow. To model this in vitro, phorbol myristate acetate (PMA)-activated PMN were exposed to shear stress on cultured human umbilical vein endothelial cells (HUVECs) and on plastic dishes coated with bovine serum albumin (BSA). PMN adhesion to HUVECs averaged 36% of the total PMNs added and was reduced to 21% by shear stress of approximately 1.5 dynes.cm-2. On BSA, adhesion was reduced from 59% to 35%. Dextran sulfate (molecular weight 500,000) inhibited PMN adhesion in a dose-dependent manner when shear stress was applied. At a concentration of 1 mg.ml-1, inhibition was 72% on HUVECs and 76% on BSA. Half-maximal inhibition was reached at approximately 1 microgram.mL-1 dextran sulfate, corresponding to 2 nmol/L. Without shear stress, dextran sulfate had no effect on HUVECs and only a moderate effect on BSA. The murine monoclonal antibody (MoAb) 60.3, recognizing an epitope on the leukocyte adhesion glycoprotein CD18, inhibited PMN adhesion equally well with and without shear. A low dose of MoAb 60.3 enhanced the effect of dextran sulfate without shear stress. Flow cytometry (FACS) did not show inhibition of MoAb 60.3 binding to PMNs by dextran sulfate. These results indicate that a dextran sulfate-inhibitable adhesion process is important for PMN adhesion in the presence of shear stress.

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Effect of Synchronous Versus Sequential Regimens on the Pharmacokinetics and Biodistribution of Regorafenib with Irradiation

Pharmaceutics ◽

10.3390/pharmaceutics13030386 ◽

2021 ◽

Vol 13 (3) ◽

pp. 386

Author(s):

Tung-Hu Tsai ◽

Yu-Jen Chen ◽

Li-Ying Wang ◽

Chen-Hsi Hsieh

Keyword(s):

Stereotactic Body Radiation Therapy ◽

In Vivo Studies ◽

Control Group ◽

High Dose ◽

Dependent Manner ◽

Hep G2 ◽

Time Curve ◽

Dose Dependent

This study was performed to evaluate the interaction between conventional or high-dose radiotherapy (RT) and the pharmacokinetics (PK) of regorafenib in concurrent or sequential regimens for the treatment of hepatocellular carcinoma. Concurrent and sequential in vitro and in vivo studies of irradiation and regorafenib were designed. The interactions of RT and regorafenib in vitro were examined in the human hepatoma Huh-7, HA22T and Hep G2 cell lines. The RT–PK phenomenon and biodistribution of regorafenib under RT were confirmed in a free-moving rat model. Regorafenib inhibited the viability of Huh-7 cells in a dose-dependent manner. Apoptosis in Huh-7 cells was enhanced by RT followed by regorafenib treatment. In the concurrent regimen, RT decreased the area under the concentration versus time curve (AUC)regorafenib by 74% (p = 0.001) in the RT2 Gy × 3 fraction (f’x) group and by 69% (p = 0.001) in the RT9 Gy × 3 f’x group. The AUCregorafenib was increased by 182.8% (p = 0.011) in the sequential RT2Gy × 1 f’x group and by 213.2% (p = 0.016) in the sequential RT9Gy × 1 f’x group. Both concurrent regimens, RT2Gy × 3 f’x and RT9Gy × 3 f’x, clearly decreased the biodistribution of regorafenib in the heart, liver, lung, spleen and kidneys, compared to the control (regorafenib × 3 d) group. The concurrent regimens, both RT2Gy × 3 f’x and RT9Gy × 3 f’x, significantly decreased the biodistribution of regorafenib, compared with the control group. The PK of regorafenib can be modulated both by off-target irradiation and stereotactic body radiation therapy (SBRT).

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Dihydromyricetin Attenuates TNF-α-Induced Endothelial Dysfunction through miR-21-Mediated DDAH1/ADMA/NO Signal Pathway

BioMed Research International ◽

10.1155/2018/1047810 ◽

2018 ◽

Vol 2018 ◽

pp. 1-12 ◽

Cited By ~ 1

Author(s):

Dafeng Yang ◽

Shenglan Tan ◽

Zhousheng Yang ◽

Pei Jiang ◽

Caie Qin ◽

...

Keyword(s):

Nitric Oxide ◽

Endothelial Dysfunction ◽

Umbilical Vein ◽

Underlying Mechanism ◽

Tube Formation ◽

Signal Pathway ◽

Dependent Manner ◽

Necrosis Factor Alpha ◽

Atherosclerotic Process ◽

Umbilical Vein Endothelial Cells

Accumulating studies demonstrate that dihydromyricetin (DMY), a compound extracted from Chinese traditional herb, Ampelopsis grossedentata, attenuates atherosclerotic process by improvement of endothelial dysfunction. However, the underlying mechanism remains poorly understood. Thus, the aim of this study is to investigate the potential mechanism behind the attenuating effects of DMY on tumor necrosis factor alpha- (TNF-α-) induced endothelial dysfunction. In response to TNF-α, microRNA-21 (miR-21) expression was significantly increased in human umbilical vein endothelial cells (HUVECs), in line with impaired endothelial dysfunction as evidenced by decreased tube formation and migration, endothelial nitric oxide synthase (eNOS) (ser1177) phosphorylation, dimethylarginine dimethylaminohydrolases 1 (DDAH1) expression and metabolic activity, and nitric oxide (NO) concentration as well as increased asymmetric dimethylarginine (ADMA) levels. In contrast, DMY or blockade of miR-21 expression ameliorated endothelial dysfunction in HUVECs treated with TNF-α through downregulation of miR-21 expression, whereas these effects were abolished by overexpression of miR-21. In addition, using a nonspecific NOS inhibitor, L-NAME, also abrogated the attenuating effects of DMY on endothelial dysfunction. Taken together, these data demonstrated that miR-21-mediated DDAH1/ADMA/NO signal pathway plays an important role in TNF-α-induced endothelial dysfunction, and DMY attenuated endothelial dysfunction induced by TNF-α in a miR-21-dependent manner.

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Endothelial AMP-Activated Kinase α1 Phosphorylates eNOS on Thr495 and Decreases Endothelial NO Formation

International Journal of Molecular Sciences ◽

10.3390/ijms19092753 ◽

2018 ◽

Vol 19 (9) ◽

pp. 2753 ◽

Cited By ~ 6

Author(s):

Nina Zippel ◽

Annemarieke Loot ◽

Heike Stingl ◽

Voahanginirina Randriamboavonjy ◽

Ingrid Fleming ◽

...

Keyword(s):

Endothelial Cells ◽

Vascular Reactivity ◽

Umbilical Vein ◽

No Production ◽

Dependent Manner ◽

Calmodulin Binding ◽

Umbilical Vein Endothelial Cells ◽

Endothelial Nitric Oxide ◽

Specific Deletion

AMP-activated protein kinase (AMPK) is frequently reported to phosphorylate Ser1177 of the endothelial nitric-oxide synthase (eNOS), and therefore, is linked with a relaxing effect. However, previous studies failed to consistently demonstrate a major role for AMPK on eNOS-dependent relaxation. As AMPK also phosphorylates eNOS on the inhibitory Thr495 site, this study aimed to determine the role of AMPKα1 and α2 subunits in the regulation of NO-mediated vascular relaxation. Vascular reactivity to phenylephrine and acetylcholine was assessed in aortic and carotid artery segments from mice with global (AMPKα−/−) or endothelial-specific deletion (AMPKαΔEC) of the AMPKα subunits. In control and AMPKα1-depleted human umbilical vein endothelial cells, eNOS phosphorylation on Ser1177 and Thr495 was assessed after AMPK activation with thiopental or ionomycin. Global deletion of the AMPKα1 or α2 subunit in mice did not affect vascular reactivity. The endothelial-specific deletion of the AMPKα1 subunit attenuated phenylephrine-mediated contraction in an eNOS- and endothelium-dependent manner. In in vitro studies, activation of AMPK did not alter the phosphorylation of eNOS on Ser1177, but increased its phosphorylation on Thr495. Depletion of AMPKα1 in cultured human endothelial cells decreased Thr495 phosphorylation without affecting Ser1177 phosphorylation. The results of this study indicate that AMPKα1 targets the inhibitory phosphorylation Thr495 site in the calmodulin-binding domain of eNOS to attenuate basal NO production and phenylephrine-induced vasoconstriction.

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Endothelial nitric oxide production during in vitro simulation of external limb compression

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00288.2001 ◽

2002 ◽

Vol 282 (6) ◽

pp. H2066-H2075 ◽

Cited By ~ 23

Author(s):

Guohao Dai ◽

Olga Tsukurov ◽

Michael Chen ◽

Jonathan P. Gertler ◽

Roger D. Kamm

Keyword(s):

Nitric Oxide ◽

Mrna Expression ◽

Umbilical Vein ◽

Control Group ◽

No Production ◽

Vein Thrombosis ◽

Enos Mrna ◽

Umbilical Vein Endothelial Cells ◽

Endothelial Nitric Oxide

External pneumatic compression (EPC) is effective in preventing deep vein thrombosis (DVT) and is thought to alter endothelial thromboresistant properties. We investigated the effect of EPC on changes in nitric oxide (NO), a critical mediator in the regulation of vasomotor and platelet function. An in vitro cell culture system was developed to simulate flow and vessel collapse conditions under EPC. Human umbilical vein endothelial cells were cultured and subjected to tube compression (C), pulsatile flow (F), or a combination of the two (FC). NO production and endothelial nitric oxide synthase (eNOS) mRNA expression were measured. The data demonstrate that in the F and FC groups, there is a rapid release of NO followed by a sustained increase. NO production levels in the F and FC groups were almost identical, whereas the C group produced the same low amount of NO as the control group. Conditions F and FC also upregulate eNOS mRNA expression by a factor of 2.08 ± 0.25 and 2.11 ± 0.21, respectively, at 6 h. Experiments with different modes of EPC show that NO production and eNOS mRNA expression respond to different time cycles of compression. These results implicate enhanced NO release as a potentially important factor in the prevention of DVT.

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Aspergillus fumigatus inhibits angiogenesis through the production of gliotoxin and other secondary metabolites

Blood ◽

10.1182/blood-2009-07-231209 ◽

2009 ◽

Vol 114 (26) ◽

pp. 5393-5399 ◽

Cited By ~ 78

Author(s):

Ronen Ben-Ami ◽

Russell E. Lewis ◽

Konstantinos Leventakos ◽

Dimitrios P. Kontoyiannis

Keyword(s):

Invasive Aspergillosis ◽

Aspergillus Fumigatus ◽

Capillary Tube ◽

Umbilical Vein ◽

Tube Formation ◽

Dependent Manner ◽

Antiangiogenic Effect ◽

Umbilical Vein Endothelial Cells

AbstractIn susceptible hosts, angioinvasion by Aspergillus fumigatus triggers thrombosis, hypoxia, and proinflammatory cytokine release, all of which are stimuli for angiogenesis. We sought to determine whether A fumigatus directly modulates angiogenesis. A fumigatus culture filtrates profoundly inhibited the differentiation, migration, and capillary tube formation of human umbilical vein endothelial cells in vitro. To measure angiogenesis at the site of infection, we devised an in vivo Matrigel assay in cyclophosphamide-treated BALB/c mice with cutaneous invasive aspergillosis. Angiogenesis was significantly suppressed in Matrigel plugs implanted in A fumigatus–infected mice compared with plugs from uninfected control mice. The antiangiogenic effect of A fumigatus was completely abolished by deletion of the global regulator of secondary metabolism, laeA, and to a lesser extent by deletion of gliP, which controls gliotoxin production. Moreover, pure gliotoxin potently inhibited angiogenesis in vitro in a dose-dependent manner. Finally, overexpression of multiple angiogenesis mediator–encoding genes was observed in the lungs of cortisone-treated mice during early invasive aspergillosis, whereas gene expression returned rapidly to baseline levels in cyclophosphamide/cortisone-treated mice. Taken together, these results indicate that suppression of angiogenesis by A fumigatus both in vitro and in a neutropenic mouse model is mediated through secondary metabolite production.

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Protein-free phospholipid emulsion treatment improved cardiopulmonary function and survival in porcine sepsis

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.00285.2002 ◽

2003 ◽

Vol 284 (2) ◽

pp. R550-R557 ◽

Cited By ~ 22

Author(s):

Roy D. Goldfarb ◽

Thomas S. Parker ◽

Daniel M. Levine ◽

Dana Glock ◽

Imran Akhter ◽

...

Keyword(s):

Density Lipoprotein ◽

Fibrin Clot ◽

Control Group ◽

High Dose ◽

Dependent Manner ◽

Serum Lipoproteins ◽

Treatment Groups ◽

Tnf Α ◽

Dose Dependent

Lipoprotein phospholipid (PL) plays a major role in neutralization of endotoxin. This study tested the hypothesis that prophylactic administration of a PL-enriched emulsion (PRE), which augments PL content of serum lipoproteins and neutralizes endotoxin in vitro, would preserve cardiovascular function and improve survival in porcine septic peritonitis. A control group was compared with low-, mid-, and high-dose treatment groups that received PRE by primed continuous infusion for 48 h. A fibrin clot containing live Escherichia coli 0111.B4 was implanted intraperitoneally 30 min after the priming dose. Survival increased in a dose-dependent manner and was correlated with serum PL. Infused PL was associated with high-density lipoprotein in the low-dose group and all serum lipoproteins at higher doses. Treatment significantly lowered serum endotoxin and tumor necrosis factor (TNF)-α, preserved cardiac output and ejection fraction, and attenuated increases in systemic and pulmonary vascular resistances. This study demonstrated that augmentation of lipoprotein PL via administration of PRE improved survival and offered a novel therapeutic approach to sepsis.

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