scholarly journals Impact of Na+ permeation on collective migration of pulmonary arterial endothelial cells

PLoS ONE ◽  
2021 ◽  
Vol 16 (4) ◽  
pp. e0250095
Author(s):  
Ningyong Xu ◽  
Linn Ayers ◽  
Viktoriya Pastukh ◽  
Mikhail Alexeyev ◽  
Troy Stevens ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Wound Closure ◽  
Combined Treatment ◽  
Intercellular Adhesion ◽  
Second Phase ◽  
Collective Migration ◽  
Channel Protein ◽  
Pharmacological Inhibition ◽  
Floating Particles

Collective migration of endothelial cells is important for wound healing and angiogenesis. During such migration, each constituent endothelial cell coordinates its magnitude and direction of migration with its neighbors while retaining intercellular adhesion. Ensuring coordination and cohesion involves a variety of intra- and inter-cellular signaling processes. However, the role of permeation of extracellular Na+ in collective cell migration remains unclear. Here, we examined the effect of Na+ permeation in collective migration of pulmonary artery endothelial cell (PAEC) monolayers triggered by either a scratch injury or a barrier removal over 24 hours. In the scratch assay, PAEC monolayers migrated in two approximately linear phases. In the first phase, wound closure started with fast speed which then rapidly reduced within 5 hours after scratching. In the second phase, wound closure maintained at slow and stable speed from 6 to 24 hours. In the absence of extracellular Na+, the wound closure distance was reduced by >50%. Fewer cells at the leading edge protruded prominent lamellipodia. Beside transient gaps, some sustained interendothelial gaps also formed and progressively increased in size over time, and some fused with adjacent gaps. In the absence of both Na+ and scratch injury, PAEC monolayer migrated even more slowly, and interendothelial gaps obviously increased in size towards the end. Pharmacological inhibition of the epithelial Na+ channel (ENaC) using amiloride reduced wound closure distance by 30%. Inhibition of both the ENaC and the Na+/Ca2+ exchanger (NCX) using benzamil further reduced wound closure distance in the second phase and caused accumulation of floating particles in the media. Surprisingly, pharmacological inhibition of the Ca2+ release-activated Ca2+ (CRAC) channel protein 1 (Orai1) using GSK-7975A, the transient receptor potential channel protein 1 and 4 (TRPC1/4) using Pico145, or both Orai1 and TRPC1/4 using combined GSK-7975A and Pico145 treatment did not affect wound closure distance dramatically. Nevertheless, the combined treatment appeared to cause accumulation of floating particles. Note that GSK-7975A also inhibits small inward Ca2+ currents via Orai2 and Orai3 channels, whereas Pico145 also blocks TRPC4, TRPC5, and TRPC1/5 channels. By contrast, gene silence of Orai1 by shRNAs led to a 25% reduction of wound closure in the first 6 hours but had no effect afterwards. However, in the absence of extracellular Na+ or cellular injury, Orai1 did not affect PAEC collective migration. Overall, the data reveal that Na+ permeation into cells contributes to PAEC monolayer collective migration by increasing lamellipodial formation, reducing accumulation of floating particles, and improving intercellular adhesion.

scholarly journals Induced pluripotent stem cell-derived endothelial cells promote angiogenesis and accelerate wound closure in a murine excisional wound healing model

2018 ◽  
Vol 38 (4) ◽  
Author(s):  
Zoë E. Clayton ◽  
Richard P. Tan ◽  
Maria M. Miravet ◽  
Katarina Lennartsson ◽  
John P. Cooke ◽  
...  
Keyword(s):  
Stem Cells ◽  
Wound Healing ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Adult Stem Cells ◽  
Wound Closure ◽  
Chronic Wounds ◽  
Control Treatment ◽  
Excisional Wound ◽  
Induced Pluripotent

Chronic wounds are a major complication in patients with cardiovascular diseases. Cell therapies have shown potential to stimulate wound healing, but clinical trials using adult stem cells have been tempered by limited numbers of cells and invasive procurement procedures. Induced pluripotent stem cells (iPSCs) have several advantages of other cell types, for example they can be generated in abundance from patients’ somatic cells (autologous) or those from a matched donor. iPSCs can be efficiently differentiated to functional endothelial cells (iPSC-ECs). Here, we used a murine excisional wound model to test the pro-angiogenic properties of iPSC-ECs in wound healing. Two full-thickness wounds were made on the dorsum of NOD-SCID mice and splinted. iPSC-ECs (5 × 105) were topically applied to one wound, with the other serving as a control. Treatment with iPSC-ECs significantly increased wound perfusion and accelerated wound closure. Expression of endothelial cell (EC) surface marker, platelet endothelial cell adhesion molecule (PECAM-1) (CD31), and pro-angiogenic EC receptor, Tie1, mRNA was up-regulated in iPSC-EC treated wounds at 7 days post-wounding. Histological analysis of wound sections showed increased capillary density in iPSC-EC wounds at days 7 and 14 post-wounding, and increased collagen content at day 14. Anti-GFP fluorescence confirmed presence of iPSC-ECs in the wounds. Bioluminescent imaging (BLI) showed progressive decline of iPSC-ECs over time, suggesting that iPSC-ECs are acting primarily through short-term paracrine effects. These results highlight the pro-regenerative effects of iPSC-ECs and demonstrate that they are a promising potential therapy for intractable wounds.

scholarly journals PKM2 regulates endothelial cell junction dynamics and angiogenesis via ATP production

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Jesús Gómez-Escudero ◽  
Cristina Clemente ◽  
Diego García-Weber ◽  
Rebeca Acín-Pérez ◽  
Jaime Millán ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Inflammatory Disease ◽  
Chronic Inflammatory Disease ◽  
Cell Junctions ◽  
Cell Junction ◽  
Collective Migration ◽  
Sprouting Angiogenesis ◽  
Cell Cell

Abstract Angiogenesis, the formation of new blood vessels from pre-existing ones, occurs in pathophysiological contexts such as wound healing, cancer, and chronic inflammatory disease. During sprouting angiogenesis, endothelial tip and stalk cells coordinately remodel their cell-cell junctions to allow collective migration and extension of the sprout while maintaining barrier integrity. All these processes require energy, and the predominant ATP generation route in endothelial cells is glycolysis. However, it remains unclear how ATP reaches the plasma membrane and intercellular junctions. In this study, we demonstrate that the glycolytic enzyme pyruvate kinase 2 (PKM2) is required for sprouting angiogenesis in vitro and in vivo through the regulation of endothelial cell-junction dynamics and collective migration. We show that PKM2-silencing decreases ATP required for proper VE-cadherin internalization/traffic at endothelial cell-cell junctions. Our study provides fresh insight into the role of ATP subcellular compartmentalization in endothelial cells during angiogenesis. Since manipulation of EC glycolysis constitutes a potential therapeutic intervention route, particularly in tumors and chronic inflammatory disease, these findings may help to refine the targeting of endothelial glycolytic activity in disease.

Shear stress enhances human endothelial cell wound closure in vitro

2000 ◽  
Vol 279 (1) ◽  
pp. H293-H302 ◽  
Author(s):  
Maria Luiza C. Albuquerque ◽  
Christopher M. Waters ◽  
Ushma Savla ◽  
H. William Schnaper ◽  
Annette S. Flozak
Keyword(s):  
Endothelial Cells ◽  
Shear Stress ◽  
Endothelial Cell ◽  
Wound Closure ◽  
Type I Collagen ◽  
Umbilical Vein ◽  
Type I ◽  
Laminar Shear Stress ◽  
Human Coronary Artery ◽  
Closure Rate

Repair of the endothelium occurs in the presence of continued blood flow, yet the mechanisms by which shear forces affect endothelial wound closure remain elusive. Therefore, we tested the hypothesis that shear stress enhances endothelial cell wound closure. Human umbilical vein endothelial cells (HUVEC) or human coronary artery endothelial cells (HCAEC) were cultured on type I collagen-coated coverslips. Cell monolayers were sheared for 18 h in a parallel-plate flow chamber at 12 dyn/cm2 to attain cellular alignment and then wounded by scraping with a metal spatula. Subsequently, the monolayers were exposed to a laminar shear stress of 3, 12, or 20 dyn/cm2 under shear-wound-shear (S-W-sH) or shear-wound-static (S-W-sT) conditions for 6 h. Wound closure was measured as a percentage of original wound width. Cell area, centroid-to-centroid distance, and cell velocity were also measured. HUVEC wounds in the S-W-sH group exposed to 3, 12, or 20 dyn/cm2 closed to 21, 39, or 50%, respectively, compared with only 59% in the S-W-sT cells. Similarly, HCAEC wounds closed to 29, 49, or 33% (S-W-sH) compared with 58% in the S-W-sT cells. Cell spreading and migration, but not proliferation, were the major mechanisms accounting for the increases in wound closure rate. These results suggest that physiological levels of shear stress enhance endothelial repair.

scholarly journals Lymphocyte function-associated antigen-1 (LFA-1) interaction with intercellular adhesion molecule-1 (ICAM-1) is one of at least three mechanisms for lymphocyte adhesion to cultured endothelial cells.

1988 ◽  
Vol 107 (1) ◽  
pp. 321-331 ◽  
Author(s):  
M L Dustin ◽  
T A Springer
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Adhesion Molecule ◽  
Umbilical Vein ◽  
Peripheral Blood Lymphocytes ◽  
Intercellular Adhesion ◽  
Intercellular Adhesion Molecule ◽  
Lymphocyte Function ◽  
Intercellular Adhesion Molecule 1 ◽  
Lymphocyte Adhesion

Intercellular adhesion molecule-1 (ICAM-1) on the surface of cultured umbilical vein and saphenous vein endothelial cells was upregulated between 2.5- and 40-fold by rIL-1, rTNF, LPS and rIFN gamma corresponding to up to 5 X 10(6) sites/cell. Endothelial cell ICAM-1 was a single band of 90 kD in SDS-PAGE. Purified endothelial cell ICAM-1 reconstituted into liposomes and bound to plastic was an excellent substrate for both JY B lymphoblastoid cell and T lymphoblast adhesion. Adhesion to endothelial cell ICAM-1 in planar membranes was blocked completely by monoclonal antibodies to lymphocyte function associated antigen-1 (LFA-1) or ICAM-1. Adhesion to artificial membranes was most sensitive to ICAM-1 density within the physiological range found on resting and stimulated endothelial cells. Adhesion of JY B lymphoblastoid cells, normal and genetically LFA-1 deficient T lymphoblasts and resting peripheral blood lymphocytes to endothelial cell monolayers was also assayed. In summary, LFA-1 dependent (60-90% of total adhesion) and LFA-1-independent basal adhesion was observed and the use of both adhesion pathways by different interacting cell pairs was increased by monokine or lipopolysaccharide stimulation of endothelial cells. The LFA-1-dependent adhesion could be further subdivided into an LFA-1/ICAM-1-dependent component which was increased by cytokines and a basal LFA-1-dependent, ICAM-1-independent component which did not appear to be affected by cytokines. We conclude that ICAM-1 is a regulated ligand for lymphocyte-endothelial cell adhesion, but at least two other major adhesion pathways exist.

scholarly journals Polymerisation of fibrin αC-domains promotes endothelial cell migration and proliferation

2014 ◽  
Vol 112 (12) ◽  
pp. 1244-1251 ◽  
Author(s):  
Sergiy Yakovlev ◽  
Irina Mikhailenko ◽  
Galina Tsurupa ◽  
Alexey Belkin ◽  
Leonid Medved
Keyword(s):  
Cell Proliferation ◽  
Wound Healing ◽  
Endothelial Cells ◽  
Cell Migration ◽  
Endothelial Cell ◽  
Wound Closure ◽  
Signalling Pathway ◽  
Endothelial Cell Migration ◽  
Specific Antagonist ◽  
Cell Migration And Proliferation

SummaryUpon conversion of fibrinogen into fibrin, fibrinogen αC-domains containing the RGD recognition motif form ordered αC polymers. Our previous study revealed that polymerisation of these domains promotes integrin-dependent adhesion and spreading of endothelial cells, as well as integrin-mediated activation of the FAK and ERK1/2 signalling pathways. The major goal of this study was to test the impact of αC-domain polymerisation on endothelial cell migration and proliferation during wound healing, and to clarify the mechanism underlying superior activity of αC polymers toward endothelial cells. In an in vitro wound healing assay, confluent endothelial cell monolayers on tissue culture plates coated with the αC monomer or αC polymers were wounded by scratching and wound closure was monitored by timelapse videomicroscopy. Although the plates were coated with equal amounts of αC species, as confirmed by ELISA, wound closure by the cells occurred much faster on αC polymers, indicating that αC-domain polymerisation promotes cell migration and proliferation. In agreement, endothelial cell proliferation was also more efficient on αC polymers, as revealed by cell proliferation assay. Wound closure on both types of substrates was equally inhibited by the integrin-blocking GRGDSP peptide and a specific antagonist of the ERK1/2 signalling pathway. In contrast, blocking the FAK signaling pathway by a specific antagonist decreased wound closure only on αC polymers. These results indicate that polymerisation of the αC-domains enhances integrin-dependent endothelial cell migration and proliferation mainly through the FAK signalling pathway. Furthermore, clustering of integrin-binding RGD motifs in αC polymers is the major mechanism triggering these events.

The Endothelial Cell and the Factor VIII Bypassing Activity of Prothrombin Complex Concentrate

1988 ◽  
Vol 60 (02) ◽  
pp. 226-229 ◽  
Author(s):  
Jerome M Teitel ◽  
Hong-Yu Ni ◽  
John J Freedman ◽  
M Bernadette Garvey
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Factor Viii ◽  
Prothrombin Complex Concentrate ◽  
Vascular Endothelial Cells ◽  
Vascular Endothelial ◽  
Monolayer Cultures ◽  
Coagulant Activity ◽  
Time Courses ◽  
Privileged Site

SummarySome classical hemophiliacs have a paradoxical hemostatic response to prothrombin complex concentrate (PCC). We hypothesized that vascular endothelial cells (EC) may contribute to this “factor VIII bypassing activity”. When PCC were incubated with suspensions or monolayer cultures of EC, they acquired the ability to partially bypass the defect of factor VIII deficient plasma. This factor VIII bypassing activity distributed with EC and not with the supernatant PCC, and was not a general property of intravascular cells. The effect of PCC was even more dramatic on fixed EC monolayers, which became procoagulant after incubation with PCC. The time courses of association and dissociation of the PCC-derived factor VIII bypassing activity of fixed and viable EC monolayers were both rapid. We conclude that EC may provide a privileged site for sequestration of constituents of PCC which express coagulant activity and which bypass the abnormality of factor VIII deficient plasma.

Immunoelectrophoretic Analysis of Membrane Glycoproteins in Cultured Human Endothelial Cells

1990 ◽  
Vol 63 (02) ◽  
pp. 303-311
Author(s):  
Tone Børsum
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Rabbit Antiserum ◽  
Platelet Glycoprotein ◽  
Membrane Glycoproteins ◽  
Human Endothelial Cells ◽  
Immunoelectrophoretic Analysis ◽  
Glycoprotein Complex ◽  
Crossed Immunoelectrophoresis ◽  
Triton X

SummaryHuman endothelial cells isolated from umbilical cordswere solubilized in Triton X-100 and examined by crossedimmunoelec-trophoresis using rabbit antiserum against endothelial cells. Endogenous labelling of the endothelialcell proteins with 14Cmannose followed by crossed immunoelectrophoresis and autoradiography revealed about 10 immunoprecipitates. Four of these endothelial cell glycoproteins were labelled by lactoperoxidase catalyzed iodination and thus were surface located. Three of the surface located glycoproteins showed reduced electrophoretic mobility after incubation of the endothelial cells with neuraminidase and were therefore sialoglycoproteins. Amphiphilicity of endothelial cell glycoproteins was studied by crossed hydrophobic interaction immunoelectrophoresis with phenyl-Sepharose in the intermediate gel. Amphiphilic proteins also show increasing electrophoretic migration velocity with decreasing concentration of Triton X-100 in the first dimension gels. Five of the endothelial cell glycoproteins were shown to be amphiphilic using these two techniques.Two monoclonal antibodies against the platelet glycoprotein complex Ilb-IIIa and glycoprotein IlIa, respectively, reacted with the same precipitate of endothelial cells. When a polyclonal antibody against the platelet glycoprotein complex Ilb-IIIa was incorporated into the intermediate gel the position of two endothelial cell precipitates were lowered. One of these was a sialoglycoprotein.

Alterations in Vascular Endothelial Cell-related Plasma Proteins In Thalassaemic Patients and their Correlation with Clinical Symptoms

1995 ◽  
Vol 74 (04) ◽  
pp. 1045-1049 ◽  
Author(s):  
P Butthep ◽  
A Bunyaratvej ◽  
Y Funahara ◽  
H Kitaguchi ◽  
S Fucharoen ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Immunosorbent Assay ◽  
Plasma Proteins ◽  
Clinical Symptoms ◽  
Vascular Endothelial Cells ◽  
Vascular Endothelial Cell ◽  
Enzyme Linked Immunosorbent Assay ◽  
Vascular Endothelial ◽  
Leg Ulcers

SummaryAn increased level of plasma thrombomodulin (TM) in α- and β- thalassaemia was demonstrated using an enzyme-linked immunosorbent assay (ELISA). Nonsplenectomized patients with β-thalassaemia/ haemoglobin E (BE) had higher levels of TM than splenectomized cases (BE-S). Patients with leg ulcers (BE-LU) were found to have the highest increase in TM level. Appearance of larger platelets in all types of thalassaemic blood was observed indicating an increase in the number of younger platelets. These data indicate that injury of vascular endothelial cells is present in thalassaemic patients.

An Electron Microscopic Study of Platelet Thrombus Formation in the Rabbit with Particular Regard to 5-Hydroxytryptamine Release

1967 ◽  
Vol 18 (03/04) ◽  
pp. 592-604 ◽  
Author(s):  
H. R Baumgartner ◽  
J. P Tranzer ◽  
A Studer
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Vessel Wall ◽  
Thrombus Formation ◽  
Endothelial Damage ◽  
Electron Microscopic ◽  
Rabbit Ear ◽  
Basement Lamina ◽  
Platelet Thrombus

SummaryElectron microscopic and histologic examination of rabbit ear vein segments 4 and 30 min after slight endothelial damage have yielded the following findings :1. Platelets do not adhere to damaged endothelial cells.2. If the vessel wall is denuded of the whole endothelial cell, platelets adhere to the intimai basement lamina as do endothelial cells.3. The distance between adherent platelets as well as endothelial cells and intimai basement lamina measures 10 to 20 mµ, whereas the distance between aggregated platelets is 30 to 60 mµ.4. 5-hydroxytryptamine (5-HT) is released from platelets during viscous metamorphosis at least in part as 5-HT organelles.It should be noted that the presence of collagen fibers is not necessary for platelet thrombus formation in vivo.

Endothelial Cell Protein S Synthesis Is Upregulated by the Complex of IL-6 and Soluble IL-6 Receptor

1997 ◽  
Vol 77 (05) ◽  
pp. 1014-1019 ◽  
Author(s):  
W Craig Hooper ◽  
Donald J Phillips ◽  
Bruce L Evatt
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Cell Line ◽  
Neutralizing Antibodies ◽  
Hepatoma Cell ◽  
Protein S ◽  
Oncostatin M ◽  
Cell Protein ◽  
Microvascular Endothelial Cell ◽  
Hepatoma Cell Line

SummaryWe have recently demonstrated that the proinflammatory cytokine, interleukin-6 (IL-6), could upregulate the production of protein S in the human hepatoma cell line, HepG-2, but not in endothelial cells. In this study, we have demonstrated that the combination of exogenous IL-6 and soluble IL-6 receptor (sIL-6R) could significantly upregulate protein S production in both primary human umbilical vein endothelial cells (HUVEC) and in the immortalized human microvascular endothelial cell line, HMEC-1. The IL-6/sIL-6R complex was also able to rapidly induce tyrosine phosphorylation of the IL-6 transducer, gpl30. Neutralizing antibodies directed against either IL-6 or gpl30 blocked protein S upregulation by the IL-6/sIL-6R complex. It was also observed that exogenous sIL-6R could also upregulate protein S by forming a complex with IL-6 constitutively produced by the endothelial cell. Two other cytokines which also utilize the gpl30 receptor, oncostatin M (OSM) and leukemia inhibitory factor (LIF), were also able to upregulate endothelial cell protein S. This study demonstrates a mechanism that allows endothelial cells to respond to IL-6 and also illustrates the potential importance of circulating soluble receptors in the regulation of the anticoagulation pathway.

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