Activation of pro-(matrix metalloproteinase-2) (pro-MMP-2) by thrombin is membrane-type-MMP-dependent in human umbilical vein endothelial cells and generates a distinct 63 kDa active species

2001 ◽  
Vol 357 (1) ◽  
pp. 107-115 ◽  
Author(s):  
Marc A. LAFLEUR ◽  
Morley D. HOLLENBERG ◽  
Susan J. ATKINSON ◽  
Vera KNÄUPER ◽  
Gillian MURPHY ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Matrix Metalloproteinase ◽  
Umbilical Vein ◽  
Coagulation Cascade ◽  
Matrix Metalloproteinase 2 ◽  
Intermediate Form ◽  
Dependent Manner ◽  
Human Umbilical Vein ◽  
Membrane Type ◽  
Umbilical Vein Endothelial Cells

Thrombin, a critical enzyme in the coagulation cascade, has also been associated with angiogenesis and activation of the zymogen form of matrix metalloproteinase-2 (MMP-2 or gelatinase-A). We show that thrombin activated pro-MMP-2 in a dose- and time-dependent manner in cultured human umbilical-vein endothelial cells (HUVECs) to generate a catalytically active 63kDa protein that accumulated as the predominant form in the conditioned medium. This 63kDa thrombin-activated MMP-2 is distinct from the 62kDa species found following concanavalin A or PMA stimulated pro-MMP-2 activation. Hirudin and leupeptin blocked thrombin-induced pro-MMP-2 activation, demonstrating that the proteolytic activity of thrombin is essential. However, activation was also dependent upon membrane-type-MMP (MT-MMP) action, since it was blocked by EDTA, o-phenanthroline, hydroxamate metalloproteinase inhibitors, tissue inhibitor of metalloproteinase-2 (TIMP-2) and TIMP-4, but not TIMP-1. Thrombin inefficiently cleaved recombinant 72kDa pro-MMP-2, but efficiently cleaved the 64kDa MT-MMP-processed intermediate form in the presence of cells. Thrombin also rapidly (within 1h) increased cellular MT-MMP activity, and at longer time points (>6h) it increased expression of MT1-MMP mRNA and protein. Thus signalling via proteinase-activated receptors (PARs) may play a role in thrombin-induced MMP-2 activation, though this does not appear to involve PAR1, PAR2, or PAR4 in HUVECs. These results indicate that in HUVECs the activation of pro-MMP-2 by thrombin involves increased MT-MMP activity and preferential cleavage of the MT-MMP-processed 64kDa MMP-2 form in the presence of cells. The integration of these proteinase systems in the vascular endothelium may be important during thrombogenesis and tissue remodelling associated with neovascularization.

scholarly journals Homocysteine Induces Apoptosis of Human Umbilical Vein Endothelial Cells via Mitochondrial Dysfunction and Endoplasmic Reticulum Stress

2017 ◽  
Vol 2017 ◽  
pp. 1-13 ◽  
Author(s):  
Zhimin Zhang ◽  
Congying Wei ◽  
Yanfen Zhou ◽  
Tao Yan ◽  
Zhengqiang Wang ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Er Stress ◽  
Mitochondrial Dysfunction ◽  
Umbilical Vein ◽  
Dependent Manner ◽  
Human Umbilical Vein ◽  
Homologous Protein ◽  
Intracellular Ros ◽  
Underlying Mechanisms ◽  
Umbilical Vein Endothelial Cells

Homocysteine- (Hcy-) induced endothelial cell apoptosis has been suggested as a cause of Hcy-dependent vascular injury, while the proposed molecular pathways underlying this process are unclear. In this study, we investigated the adverse effects of Hcy on human umbilical vein endothelial cells (HUVEC) and the underlying mechanisms. Our results demonstrated that moderate-dose Hcy treatment induced HUVEC apoptosis in a time-dependent manner. Furthermore, prolonged Hcy treatment increased the expression of NOX4 and the production of intracellular ROS but decreased the ratio of Bcl-2/Bax and mitochondrial membrane potential (MMP), resulting in the leakage of cytochrome c and activation of caspase-3. Prolonged Hcy treatment also upregulated glucose-regulated protein 78 (GRP78), activated protein kinase RNA-like ER kinase (PERK), and induced the expression of C/EBP homologous protein (CHOP) and the phosphorylation of NF-κb. The inhibition of NOX4 decreased the production of ROS and alleviated the Hcy-induced HUVEC apoptosis and ER stress. Blocking the PERK pathway partly alleviated Hcy-induced HUVEC apoptosis and the activation of NF-κb. Taken together, our results suggest that Hcy-induced mitochondrial dysfunction crucially modulated apoptosis and contributed to the activation of ER stress in HUVEC. The excessive activation of the PERK pathway partly contributed to Hcy-induced HUVEC apoptosis and the phosphorylation of NF-κb.

Factor Xa inhibits HMGB1-induced septic responses in human umbilical vein endothelial cells and in mice

2014 ◽  
Vol 112 (10) ◽  
pp. 757-769 ◽  
Author(s):  
Wonhwa Lee ◽  
Sae-Kwang Ku ◽  
Jong-Sup Bae
Keyword(s):  
Endothelial Cells ◽  
Nuclear Dna ◽  
Inflammatory Responses ◽  
Umbilical Vein ◽  
Cecal Ligation ◽  
Post Treatment ◽  
Coagulation Cascade ◽  
Factor X ◽  
Human Umbilical Vein ◽  
Umbilical Vein Endothelial Cells

SummaryNuclear DNA-binding protein high mobility group box 1 (HMGB1) acts as a late mediator of severe vascular inflammatory conditions, such as sepsis. Activated factor X (FXa) is an important player in the coagulation cascade responsible for thrombin generation, and it influences cell signalling in various cell types by activating protease-activated receptors (PARs). However, the effect of FXa on HMGB1-induced inflammatory response has not been studied. First, we addressed this issue by monitoring the effects of post-treatment with FXa on lipopolysaccharide (LPS)- and cecal ligation and puncture (CLP)-mediated release of HMGB1 and HMGB1-mediated regulation of pro-inflammatory responses in human umbilical vein endothelial cells (HUVECs) and septic mice. Post-treatment with FXa was found to suppress LPS-mediated release of HMGB1 and HMGB1-mediated cytoskeletal rearrangements. FXa also inhibited HMGB1-mediated hyperpermeability and leukocyte migration in septic mice. In addition, FXa inhibited the production of tumour necrosis factor-α and interleukin (IL)-1β. FXa also facilitated the downregulation of CLP-induced release of HMGB1, production of IL-6, and mortality. Collectively, these results suggest that FXa may be regarded as a candidate therapeutic agent for treating vascular inflammatory diseases by inhibiting the HMGB1 signalling pathway.

Angiotensin II AT1 receptors and Na+/K+ ATPase in human umbilical vein endothelial cells

1997 ◽  
Vol 155 (3) ◽  
pp. 587-593 ◽  
Author(s):  
A Muscella ◽  
S Marsigliante ◽  
MA Carluccio ◽  
GP Vinson ◽  
C Storelli
Keyword(s):  
Endothelial Cells ◽  
Atpase Activity ◽  
Umbilical Vein ◽  
Maximal Response ◽  
Receptor Subtype ◽  
Dependent Manner ◽  
Ang Ii ◽  
Human Umbilical Vein ◽  
Umbilical Vein Endothelial Cells ◽  
Significant Increment

Cultured human umbilical vein endothelial cells (HUVECs) at passage 4 specifically bound 70 +/- 12 fmol [3,5-3H]Tyr4-Ile5-angiotensin (Ang) II/mg protein, with a Kd of 0.9 +/- 0.36 nM. Binding was eliminated in cells preincubated with a monoclonal antibody (6313/G2) raised against the subtype AT1 of the Ang II receptor. Freshly seeded HUVECs were positive for 6313/G2 antibody by immunocytochemistry, and such immunoreactivity was still retained at passage 4. Incubation of HUVECs for 20 min with different concentrations of Ang II provoked a significant increment in Na+/K+ ATPase activity compared with controls, in a dose- and time-dependent manner. Maximal response was obtained with 1000 pM Ang II after 20 min stimulation and resulted in a 2.2-fold increment in Na+/K+ ATPase activity. This stimulation was abolished when cells were incubated with 1000 pM Ang II in the presence of 1 microM of the specific AT1 subtype inhibitor, DuP753. Moreover, preincubation of HUVECs with 6313/G2 or with 1 mM dithiothreitol also inhibited the stimulatory effect of Ang II. These results suggest that the AT1 receptor subtype mediates the Na+/K+ ATPase response to Ang II in these cells.

A New Isoxazolic Compound Acts as α7 Nicotinic Receptor Agonist in Human Umbilical Vein Endothelial Cells

2014 ◽  
Vol 69 (7-8) ◽  
pp. 291-299 ◽  
Author(s):  
Magdalena P. Cortés ◽  
Rocío Alvarez ◽  
Evelyn Sepúlveda ◽  
Felipe Jiménez-Aspee ◽  
Luis Astudillo ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Umbilical Vein ◽  
Therapeutic Angiogenesis ◽  
Dependent Manner ◽  
Human Umbilical Vein ◽  
Α7 Nachr ◽  
Significant Difference ◽  
Umbilical Vein Endothelial Cells

Recent evidence suggests that the α7 nicotinic acetylcholine receptors (α7 nAChRs) participate in the development of angiogenesis and could be a new endothelial target for revascularization in therapeutic angiogenesis. It has been shown that in human umbilical vein endothelial cells (HUVECs) α7 nAChR agonists increase the intracellular calcium concentration ([Ca2+]i), thus inducing proliferation and vessel formation which are important stages of angiogenesis. In the present study we evaluated the effect of new isoxazole compounds on the cytosolic Ca2+ signal in HUVECs using the fluorescent Ca2+ indicator Fluo-3AM and probing the involvement of α7 nAChR by means of pharmacological tools. HUVECs expressed mainly α7 nAChR, since there was no significant difference in the increase in [Ca2+]i induced by nicotine, a non-selective nicotinic agonist, in relation to choline, a selective α7 nAChR agonist. The increase in [Ca2+]i induced by 1 mM choline was inhibited significantly (p = 0.014) in cells which had been pre-incubated for 15 min with methyllycaconitine (MLA), a selective α7 nAChR antagonist. The studied compounds 1, 2, and 3 induced an increase in [Ca2+]i in a dose-dependent manner. Compound 1 at 10 mM induced a greater increase in [Ca2+]i than compounds 2 and 3. The increase in [Ca2+]i induced by compound 1 was significantly inhibited by MLA (p = 0.013) and completely inhibited by mecamylamine, a non-selective nAChR antagonist, indicating that the isoxazolic compound 1 acts as an α7 nAChR agonist.

The protective effect of daidzein on high glucose-induced oxidative stress in human umbilical vein endothelial cells

2016 ◽  
Vol 71 (1-2) ◽  
pp. 21-28 ◽  
Author(s):  
Mi Hwa Park ◽  
Jae-Won Ju ◽  
Mihyang Kim ◽  
Ji-Sook Han
Keyword(s):  
Oxidative Stress ◽  
Nitric Oxide ◽  
Endothelial Cells ◽  
Protective Effect ◽  
High Glucose ◽  
Umbilical Vein ◽  
Vascular Complications ◽  
Dependent Manner ◽  
Human Umbilical Vein ◽  
Umbilical Vein Endothelial Cells

AbstractEndothelial cell dysfunction is considered a major cause of vascular complications in diabetes. In the present study, we investigated the protective effect of daidzein, a natural isoflavonoid, against high-glucose–induced oxidative damage in human umbilical vein endothelial cells (HUVECs). Treatment with a high concentration of glucose (30 mM) induced oxidative stress in the endothelial cells, against which daidzein protected the cells as demonstrated by significantly increased cell viability. In addition, lipid peroxidation, intracellular reactive oxygen species (ROS) generation, and indirect nitric oxide levels induced by the high glucose treatment were significantly reduced in the presence of daidzein (0.02–0.1 mM) in a dose-dependent manner. High glucose levels induced the overexpression of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and NF-κB proteins in HUVECs, which was suppressed by treatment with 0.04 mM daidzein. These findings indicate the potential of daidzein to reduce high glucose-induced oxidative stress.

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