Abstract 183: Carbon Monoxide Induces Macrophage VEGF Production and Promotes Angiogenic Behavior in Endothelial Cells

2014 ◽  
Vol 34 (suppl_1) ◽  
Author(s):  
Andrew E Leake ◽  
Guiying Hong ◽  
Christopher B Washington ◽  
Ankur J Shukla ◽  
Ulka Sachdev ◽  
...  
Keyword(s):  
Carbon Monoxide ◽  
Umbilical Vein ◽  
Fold Increase ◽  
Peritoneal Macrophages ◽  
Phenotypic Change ◽  
Enos Expression ◽  
Future Studies ◽  
Angiogenic Effect

Introduction: Carbon monoxide (CO) has potent anti-inflammatory and pro-healing properties. We have previously shown that CO enhances endothelial cell (EC) angiogenic behavior in vitro. More striking, however, is that conditioned medium (CM) from macrophages isolated from CO treated rats can promote angiogenesis indirectly. We sought to examine the mechanism of this indirect angiogenic effect of CO. Methods: Peritoneal macrophages were collected from rats after inhaled CO treatment (250 PPM for 1 hr) or air treatment. Macrophages were cultured overnight and CM was collected to treat human umbilical vein ECs (HUVECs). VEGF and eNOS expression was assessed by Western blot. Results: CM from macrophages isolated from CO treated rats (CO) showed a 3-fold increase in VEGF vs Air CM (Ratio CO/Air=2.96, Range 1.6-4.2) (Figure). VEGF levels were similar between the macrophages from CO or air treated rats (CO/Air=0.89). HUVECs treated with CO or Air CM also showed no difference in VEGF levels (CO/Air=1.07). However, eNOS expression was significantly increased in HUVECs treated with CO CM vs. Air CM (CO/Air=26.4, Range 20-32; p=0.004). HUVECs cultured in a CO chamber or a standard incubator exhibited no change in eNOS expression (CO/Air, 1.38 p=0.11). Conclusion: In vivo CO promotes macrophage secretion of VEGF. These cells can promote angiogenic behavior in ECs through this VEGF and other secreted products. These macrophage products upregulate eNOS but not VEGF expression in HUVECs. The effect of the CO CM on eNOS expression in HUVECs could not be reproduced by direct CO treatment of the cells. Future studies will further characterize the phenotypic change of the macrophages induced by inhaled CO that favors angiogenesis and healing.

A novel vasculo-angiogenic effect of cilostazol mediated by cross-talk between multiple signalling pathways including the ERK/p38 MAPK signalling transduction cascade

2012 ◽  
Vol 123 (3) ◽  
pp. 147-159 ◽  
Author(s):  
Ting-Hsing Chao ◽  
Shih-Ya Tseng ◽  
Yi-Heng Li ◽  
Ping-Yen Liu ◽  
Chung-Lung Cho ◽  
...  
Keyword(s):  
Protein Kinase ◽  
P38 Mapk ◽  
Umbilical Vein ◽  
Tube Formation ◽  
Mitogen Activated Protein Kinase ◽  
Signalling Pathways ◽  
No Production ◽  
Angiogenic Effect

Cilostazol is an anti-platelet agent with vasodilatory activity that acts by increasing intracellular concentrations of cAMP. Recent reports have suggested that cilostazol may promote angiogenesis. In the present study, we have investigated the effect of cilostazol in promoting angiogenesis and vasculogenesis in a hindlimb ischaemia model and have also examined its potential mechanism of action in vitro and in vivo. We found that cilostazol treatment significantly increased colony formation by human early EPCs (endothelial progenitor cells) through a mechanism involving the activation of cAMP/PKA (protein kinase A), PI3K (phosphoinositide 3-kinase)/Akt/eNOS (endothelial NO synthase) and ERK (extracellular-signal-regulated kinase)/p38 MAPK (mitogen-activated protein kinase) signalling pathways. Cilostazol also enhanced proliferation, chemotaxis, NO production and vascular tube formation in HUVECs (human umbilical vein endothelial cells) through activation of multiple signalling pathways downstream of PI3K/Akt/eNOS. Cilostazol up-regulated VEGF (vascular endothelial growth factor)-A165 expression and secretion of VEGF-A in HUVECs through activation of the PI3K/Akt/eNOS pathway. In a mouse hindlimb ischaemia model, recovery of blood flow ratio (ipsilateral/contralateral) 14 days after surgery was significantly improved in cilostazol-treated mice (10 mg/kg of body weight) compared with vehicle-treated controls (0.63±0.07 and 0.43±0.05 respectively, P<0.05). Circulating CD34+ cells were also increased in cilostazol-treated mice (3614±670 compared with 2151±608 cells/ml, P<0.05). Expression of VEGF and phosphorylation of PI3K/Akt/eNOS and ERK/p38 MAPK in ischaemic muscles were significantly enhanced by cilostazol. Our data suggest that cilostazol produces a vasculo-angiogenic effect by up-regulating a broad signalling network that includes the ERK/p38 MAPK, VEGF-A165, PI3K/Akt/eNOS and cAMP/PKA pathways.

Abstract 11: Thioredoxin-Interacting Protein Is Required for VEGF-Induced Angiogenesis Through Regulation of VEGFR2 Internalization

2012 ◽  
Vol 32 (suppl_1) ◽  
Author(s):  
Shin-Young Park ◽  
Chen Yan ◽  
Bradford C Berk
Keyword(s):  
Umbilical Vein ◽  
Fold Increase ◽  
Tube Formation ◽  
In Vivo Studies ◽  
Vegf Receptor ◽  
Cell Fractionation ◽  
Interacting Protein ◽  
Thioredoxin Interacting Protein

Introduction— Thioredoxin-interacting protein (TXNIP) is an arrestin-like scaffold protein. We have shown previously that it is necessary for the transactivation of the vascular endothelial growth factor (VEGF) receptor 2 (VEGFR2) as well as promoting the migration and survival of endothelial cells (ECs). However, its roles in VEGF-induced angiogenesis and in vivo studies of TXNIP function have not been elucidated. Hypothesis— TXNIP regulates VEGF-mediated angiogenesis through modulation of angiogenic signaling pathways in ECs. Methods and Results— To determine the functions of TXNIP in ECs, we generated endothelial-specific TXNIP knockout (EC-TXNIP KO) mice (TXNIPflox/flox: Tie2-Cre/+). These mice displayed impaired capillary growth of the retinal vasculature compared to control mice. Furthermore, aortic rings from EC-TXNIP KO mice exhibited fewer and shorter vascular sprouts than those in control mice. To investigate the role of TXNIP in the regulation of VEGF-induced angiogenesis, we determined the subcellular localization of TXNIP in human umbilical vein EC (HUVEC). Immunofluorescence and cell fractionation studies revealed that upon VEGF stimulation (10ng/ml). TXNIP translocated from cytoplasm to the plasma membrane. There was a 9 fold increase of membrane associated TXNIP with a peak at 15 minutes compared to non-VEGF treatment cells. We hypothesized that membrane associated TXNIP may modulate VEGFR2 internalization and thereby affect VEGF-induced signaling and angiogenesis. To investigate this, we performed in vitro cell surface biotinylation assays in HUVEC. VEGFR2 internalization was decreased by 65% in TXNIP siRNA knockdown cells compared to control siRNA treated cells following VEGF stimulation. Consistent with this result, VEGF-induced phosphorylation of VEGFR2, PLCγ and ERK1/2 was decreased by knockdown of TXNIP. Significantly, TXNIP knockdown inhibited VEGF-induced proliferation and tube formation in vitro. Conclusion— Our results suggest that TXNIP can modulate VEGF-induced angiogenesis and signaling by regulation of VEGFR2 internalization.

scholarly journals Macrophage-mediated phagocytosis of apoptotic cholangiocytes contributes to reversal of experimental biliary fibrosis

2010 ◽  
Vol 298 (3) ◽  
pp. G323-G334 ◽  
Author(s):  
Yury Popov ◽  
Deanna Y. Sverdlov ◽  
K. Ramakrishnan Bhaskar ◽  
Anisha K. Sharma ◽  
Gunda Millonig ◽  
...  
Keyword(s):  
Fold Increase ◽  
Peritoneal Macrophages ◽  
Cytokeratin 19 ◽  
Functional Link ◽  
Effector Cells ◽  
Duct Ligation ◽  
Complete Reversal ◽  
Massive Apoptosis

Studies have suggested the reversibility of liver fibrosis, but the mechanisms of fibrosis reversal are poorly understood. We investigated the possible functional link between apoptosis, macrophages, and matrix turnover in rat liver during reversal of fibrosis secondary to bile duct ligation (BDL). Biliary fibrosis was induced by BDL for 4 wk. After Roux-en-Y (RY)-bilio-jejunal-anastomosis, resolution of fibrosis was monitored for up to 12 wk by hepatic collagen content, matrix metalloproteinase (MMP) expression and activities, and fibrosis-related gene expression. MMP expression and activities were studied in macrophages after engulfment of apoptotic cholangiocytes in vitro. Hepatic collagen decreased to near normal at 12 wk after RY-anastomosis. During reversal, profibrogenic mRNA declined, whereas expression of several profibrolytic MMPs increased. Fibrotic septa showed fragmentation at week 4 and disappeared at week 12. Peak histological remodeling at week 4 was characterized by massive apoptosis of cytokeratin 19+ cholangiocytes, >90% in colocalization with CD68+ macrophages, and a 2- to 7.5-fold increase in matrix-degrading activities. In vitro, phagocytosis of apoptotic cholangiocytes induced matrix-degrading activities and MMP-3, -8, and -9 in rat peritoneal macrophages. We concluded that reconstruction of bile flow after BDL leads to an orchestrated fibrolytic program that results in near complete reversal of advanced fibrosis. The peak of connective tissue remodeling and fibrolytic activity is associated with massive apoptosis of cholangiocytes and their phagocytic clearance by macrophages in vivo. Macrophages upregulate MMPs and become fibrolytic effector cells upon apoptotic cholangiocyte engulfment in vitro, suggesting that phagocytosis-associated MMP induction in macrophages significantly contributes to biliary fibrosis reversal.

Aggretin, a snake venom–derived endothelial integrin α2β1 agonist, induces angiogenesis via expression of vascular endothelial growth factor

Blood ◽  
2004 ◽  
Vol 103 (6) ◽  
pp. 2105-2113 ◽  
Author(s):  
Ching-Hu Chung ◽  
Wen-Bin Wu ◽  
Tur-Fu Huang
Keyword(s):  
Vascular Endothelial Growth Factor ◽  
Endothelial Cell ◽  
Growth Factor ◽  
Umbilical Vein ◽  
Endothelial Cell Proliferation ◽  
Vascular Endothelial ◽  
Angiogenic Effect ◽  
Endothelial Growth Factor

Abstract Aggretin, a collagen-like α2β1 agonist purified from Calloselasma rhodostoma venom, was shown to increase human umbilical vein endothelial cell (HUVEC) proliferation and HUVEC migration toward immobilized aggretin was also increased. These effects were blocked by A2-IIE10, an antibody raised against integrin α2. Aggretin bound to HUVECs in a dose-dependent and saturable manner, which was specifically inhibited by A2-IIE10, as examined by flow cytometry. Aggretin elicited significant angiogenic effects in both in vivo and in vitro angiogenesis assays, and incubation of HUVECs with aggretin activated phosphatidylinositol 3-kinase (PI3K), Akt, and extracellular-regulated kinase 1/2 (ERK1/2); these effects were blocked by A2-IIE10 or vascular endothelial growth factor (VEGF) monoclonal antibody (mAb). The angiogenic effect induced by aggretin may be via the production of VEGF because the VEGF level was elevated and VEGF mAb pretreatment inhibited Akt/ERK1/2 activation as well as the in vivo angiogenesis induced by aggretin. The VEGF production induced by aggretin can be blocked by A2-IIE10 mAb pretreatment. In conclusion, aggretin induces endothelial cell proliferation, migration, and angiogenesis by interacting with integrin α2β1, leading to activation of PI3K, Akt, and ERK1/2 pathways, and the increased expression of VEGF may be responsible for its angiogenic activity.

scholarly journals Icariin improves eNOS/NO pathway to prohibit the atherogenesis of apolipoprotein E-null mice

2017 ◽  
Vol 95 (6) ◽  
pp. 625-633 ◽  
Author(s):  
Hong-Bo Xiao ◽  
Guo-Guang Sui ◽  
Xiang-Yang Lu
Keyword(s):  
Nitric Oxide ◽  
Apolipoprotein E ◽  
Body Mass ◽  
Umbilical Vein ◽  
Ros Production ◽  
Enos Expression ◽  
Production Body ◽  
Umbilical Vein Endothelial Cells

Impaired endothelial nitric oxide synthase (eNOS)/nitric oxide (NO) pathway induces atherogenesis. The present study examined whether icariin improves the eNOS/NO pathway to prohibit the atherogenesis of apolipoprotein E-null (ApoE−/−) mice. In vitro, primary human umbilical vein endothelial cells (HUVECs) were randomly divided into 7 groups: control; vehicle; icariin 10; lyphosphatidylcholine (LPC) group; LPC + icariin 1; LPC + icariin 3; and LPC + icariin 10. In vivo, 80 mice were separated randomly into 4 groups (n = 20): control, ApoE−/−, ApoE−/− + icariin 10, and ApoE−/− + icariin 30. ApoE−/− mice had significantly more atherosclerosis in the aortic root together with increased aortic ROS production, body mass, plasma triglyceride (TG) and total cholesterol (TC) concentration, decreased aortic eNOS expression, and plasma NO concentration. LPC (10 μg/mL) treatment induced a big decline in NO level in the conditioned medium and eNOS expression, and an increase in intracellular reactive oxygen species (ROS) production in HUVECs. Icariin treatment decreased atherogenesis, ROS production, body mass, plasma TG concentration, and plasma TC concentration, and increased NO concentration and eNOS expression. These findings suggested icariin could improve eNOS/NO-pathway to prohibit the atherogenesis of apolipoprotein E-null mice by restraining oxidative stress.

The effects of tumour necrosis factor on host—parasite relations in murine Mesocestoides corti (Cestoda) infection

Parasitology ◽  
1992 ◽  
Vol 105 (3) ◽  
pp. 453-459 ◽  
Author(s):  
P. Jenkins ◽  
S. Spiers ◽  
J. B. Dixon ◽  
S. D. Carter ◽  
S. May
Keyword(s):  
Tumour Necrosis Factor ◽  
Post Infection ◽  
Tumour Necrosis ◽  
Fold Increase ◽  
Peritoneal Macrophages ◽  
Hepatic Necrosis ◽  
Peritoneal Cells ◽  
Necrosis Factor

SUMMARYThe regulatory role of tumour necrosis factor (TNF) was investigated in murine infection with tetrathyridia of Mesocestoides corti. Recombinant TNFα reduced macrophage larvicidal activity in vitro. M. corti primed mice for TNF release in response to bacterial lipopolysaccharide (LPS) in vivo. TNF activity was amplified 100-fold at 14 days post-infection (p.i.), with a further rise at day 28 p.i. Maximal inflammatory reaction was observed histologically in the liver at the height of TNF activity. Hepatic necrosis was located within inflammatory foci, but not within the vicinity of the parasite itself, suggesting that TNF may contribute to the pathogenesis of infection. Peritoneal cells from infected mice, when stimulated with tetrathyridia in vitro, showed a 4-fold increase in TNFα activity at day 14 p.i. However, when peritoneal cells were stimulated with LPS in vitro, a marked increase in TNFα secretion was observed at 2 months post-infection followed by a slow decline. It is suggested that impaired macrophage effector function, previously attributed to endogenous endotoxin, which gains access to peritoneal macrophages through an inability of the liver to detoxify endotoxin, may be mediated through TNFα.

scholarly journals Zileuton, a 5-Lipoxygenase Inhibitor, Exerts Anti-Angiogenic Effect by Inducing Apoptosis of HUVEC via BK Channel Activation

Cells ◽  
2019 ◽  
Vol 8 (10) ◽  
pp. 1182 ◽  
Author(s):  
Lim ◽  
Park ◽  
Um ◽  
Lee ◽  
Kwak
Keyword(s):  
Umbilical Vein ◽  
Arachidonic Acid Metabolism ◽  
Bk Channel ◽  
Endothelial Cell Proliferation ◽  
Lipoxygenase Inhibitor ◽  
Apoptotic Pathway ◽  
Angiogenic Effect ◽  
Umbilical Vein Endothelial Cells

The arachidonic acid metabolism through 5-lipoxygenase (5-LO) pathways is involved in modulating both tumorigenesis and angiogenesis. Although anti-carcinogenic activities of certain 5-LO inhibitors have been reported, the role of zileuton, a well known 5-LO inhibitor, on the endothelial cell proliferation and angiogenesis has not been fully elucidated. Here, we report that zileuton has an anti-angiogenic effect, and the underlying mechanisms involved activation of the large-conductance Ca2+-activated K+ (BK) channel. Our results show that zileuton significantly prevented vascular endothelial growth factor (VEGF)-induced proliferation of human umbilical vein endothelial cells (HUVECs) in vitro, as well as in vivo. However, such anti-angiogenic effect of zileuton was abolished by iberiotoxin (IBTX), a BK channel blocker, suggesting zileuton-induced activation of BK channel was critical for the observed anti-angiogenic effect of zileuton. Furthermore, the anti-angiogenic effect of zileuton was, at least, due to the activation of pro-apoptotic signaling cascades which was also abolished by IBTX. Additionally, zileuton suppressed the expression of VCAM-1, ICAM-1, ETS related gene (Erg) and the production of nitric oxide (NO). Taken together, our results show that zileuton prevents angiogenesis by activating the BK channel dependent-apoptotic pathway, thus highlighting its therapeutic capacity in angiogenesis-related diseases, such as cancer.

Abstract 236: Carbon Monoxide Mediated Changes in Macrophages are Regulated by Vagal Activation

2017 ◽  
Vol 37 (suppl_1) ◽  
Author(s):  
Karim Salem ◽  
Guiying Hong ◽  
Andrew Leake ◽  
Ankur Aggarwal ◽  
Edith Tzeng
Keyword(s):  
Carbon Monoxide ◽  
High Mobility ◽  
Fold Increase ◽  
Arterial Injury ◽  
Peritoneal Macrophages ◽  
Significant Inhibition ◽  
Sprague Dawley ◽  
Mild Reduction ◽  
Anti Inflammatory

Introduction: Carbon monoxide (CO) has potent vasoproective effects in vivo through changes in macrophage (MΦ) function. Because of these marked changes, we examined the vagal cholinergic anti-inflammatory pathway in regulating the actions of inhaled CO. We previously showed that vagotomy inhibited the ability of inhaled CO to prevent intimal hyperplasia following arterial injury in rats. We now hypothesize that the anti-inflammatory effects of inhaled CO can be blocked through pharmacologic vagal inhibition. Methods: Sprague-Dawley rats (3 rats/group) were treated with inhaled CO (250 parts per million) for 1 hr or remained in room air. Some were injected with atropine (0.05 mg/kg SQ) 1 hr prior to and just prior to inhaled CO treatment. Peritoneal macrophages (MΦ) were collected 1 hr after CO treatment or comparable time period in control rats and were cultured overnight under standard conditions. Media from these cells were collected for Western blot analysis for high mobility box group 1 (HMGB1) and VEGF. Results: MΦ isolated from CO treated rats released increased levels of HMGB1 and VEGF into the media compared to MΦs from air treated rats (1.7 and 4 fold increase, respectively; Fig.). These molecules mediate the proendothelial actions of inhaled CO. MΦs isolated from atropine treated rats showed a mild reduction of HMGB1 and VEGF secretion. In contrast, MΦ from atropine and CO treated rats showed a significant inhibition of HMGB1 release (P<0.01 vs. all other treatment groups) and a trend toward significance in the reduction in VEGF release (Fig.). Conclusions: Inhaled CO activates vagal signaling to mediate changes in MΦ behavior. Inhibition of vagal signaling with atropine blocked the changes in MΦ induced by inhaled CO, confirming the importance of the vagus nerve in mediating the vasoprotective actions of CO. These findings suggest the potential ability to reproduce the therapeutic actions of CO through pharmacologic vagal stimulation.

Ultrastructural observations on the in vitro cytoadherence of Plasmodium falciparum infected red blood cells

1990 ◽  
Vol 48 (3) ◽  
pp. 914-915
Author(s):  
D.J.P. Ferguson ◽  
A.R. Berendt ◽  
J. Tansey ◽  
K. Marsh ◽  
C.I. Newbold
Keyword(s):  
Plasmodium Falciparum ◽  
Red Blood Cells ◽  
Blood Cells ◽  
Umbilical Vein ◽  
Cell Types ◽  
Amelanotic Melanoma ◽  
Cytoplasmic Process ◽  
Umbilical Vein Endothelial Cells

In human malaria, the most serious clinical manifestation is cerebral malaria (CM) due to infection with Plasmodium falciparum. The pathology of CM is thought to relate to the fact that red blood cells containing mature forms of the parasite (PRBC) cytoadhere or sequester to post capillary venules of various tissues including the brain. This in vivo phenomenon has been studied in vitro by examining the cytoadherence of PRBCs to various cell types and purified proteins. To date, three Ijiost receptor molecules have been identified; CD36, ICAM-1 and thrombospondin. The specific changes in the PRBC membrane which mediate cytoadherence are less well understood, but they include the sub-membranous deposition of electron-dense material resulting in surface deformations called knobs. Knobs were thought to be essential for cytoadherence, lput recent work has shown that certain knob-negative (K-) lines can cytoadhere. In the present study, we have used electron microscopy to re-examine the interactions between K+ PRBCs and both C32 amelanotic melanoma cells and human umbilical vein endothelial cells (HUVEC).We confirm previous data demonstrating that C32 cells possess numerous microvilli which adhere to the PRBC, mainly via the knobs (Fig. 1). In contrast, the HUVEC were relatively smooth and the PRBCs appeared partially flattened onto the cell surface (Fig. 2). Furthermore, many of the PRBCs exhibited an invagination of the limiting membrane in the attachment zone, often containing a cytoplasmic process from the endothelial cell (Fig. 2).

Desmopressin (DDAVP) Enhances Platelet Adhesion to the Extracellular Matrix of Cultured Human Endothelial Cells through Increased Expression of Tissue Factor

1997 ◽  
Vol 77 (05) ◽  
pp. 0975-0980 ◽  
Author(s):  
Angel Gálvez ◽  
Goretti Gómez-Ortiz ◽  
Maribel Díaz-Ricart ◽  
Ginés Escolar ◽  
Rogelio González-Sarmiento ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Endothelial Cells ◽  
Tissue Factor ◽  
Platelet Adhesion ◽  
Umbilical Vein ◽  
Procoagulant Activity ◽  
Experimental Conditions ◽  
Chromogenic Assay

SummaryThe effect of desmopressin (DDAVP) on thrombogenicity, expression of tissue factor and procoagulant activity (PCA) of extracellular matrix (ECM) generated by human umbilical vein endothelial cells cultures (HUVEC), was studied under different experimental conditions. HUVEC were incubated with DDAVP (1, 5 and 30 ng/ml) and then detached from their ECM. The reactivity towards platelets of this ECM was tested in a perfusion system. Coverslips covered with DD A VP-treated ECMs were inserted in a parallel-plate chamber and exposed to normal blood anticoagulated with low molecular weight heparin (Fragmin®, 20 U/ml). Perfusions were run for 5 min at a shear rate of 800 s1. Deposition of platelets on ECMs was significantly increased with respect to control ECMs when DDAVP was used at 5 and 30 ng/ml (p <0.05 and p <0.01 respectively). The increase in platelet deposition was prevented by incubation of ECMs with an antibody against human tissue factor prior to perfusion. Immunofluorescence studies positively detected tissue factor antigen on DDAVP derived ECMs. A chromogenic assay performed under standardized conditions revealed a statistically significant increase in the procoagulant activity of the ECMs produced by ECs incubated with 30 ng/ml DDAVP (p <0.01 vs. control samples). Northern blot analysis revealed increased levels of tissue factor mRNA in extracts from ECs exposed to DDAVP. Our data indicate that DDAVP in vitro enhances platelet adhesion to the ECMs through increased expression of tissue factor. A similar increase in the expression of tissue factor might contribute to the in vivo hemostatic effect of DDAVP.

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