scholarly journals ATP increases the migration of microglia across the brain endothelial cell monolayer

2016 ◽  
Vol 36 (2) ◽  
Author(s):  
Tomoji Maeda ◽  
Manato Inagaki ◽  
Yu Fujita ◽  
Takehiro Kimoto ◽  
Chiaki Tanabe-Fujimura ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Matrix Metalloproteinases ◽  
Real Time ◽  
Culture System ◽  
Cell Monolayer ◽  
Brain Endothelial Cell ◽  
Endothelial Cell Monolayer ◽  
Microglial Migration ◽  
The Brain

To elucidate the mechanism of microglial migration across the blood–brain barrier (BBB), we developed an in vitro co-culture system and analysed real-time BBB integrity during transmigration. We show that ATP promotes microglia transmigration via a mechanism involving microglial matrix metalloproteinases (MMPs).

scholarly journals Viable “Haemophilus somnus” Induces Myosin Light-Chain Kinase-Dependent Decrease in Brain Endothelial Cell Monolayer Resistance

2007 ◽  
Vol 75 (9) ◽  
pp. 4572-4581 ◽  
Author(s):  
E. Behling-Kelly ◽  
David McClenahan ◽  
K. S. Kim ◽  
C. J. Czuprynski
Keyword(s):  
Endothelial Cell ◽  
Light Chain ◽  
Myosin Light Chain Kinase ◽  
Myosin Light Chain ◽  
Cell Monolayer ◽  
Brain Endothelial Cell ◽  
Endothelial Cell Monolayer ◽  
Necrosis Factor Alpha ◽  
Haemophilus Somnus

ABSTRACT “Haemophilus somnus” causes thrombotic meningoencephalitis in cattle. Our laboratory has previously reported that H. somnus has the ability to adhere to, but not invade, bovine brain endothelial cells (BBEC) in vitro. The goal of this study was to determine if H. somnus alters brain endothelial cell monolayer integrity in vitro, in a manner that would be expected to contribute to inflammation of the central nervous system (CNS). Monolayer integrity was monitored by measuring transendothelial electrical resistance (TEER) and albumin flux. BBEC incubated with H. somnus underwent rapid cytoskeletal rearrangement, significant increases in albumin flux, and reductions in TEER. Decreased monolayer TEER was preceded by phosphorylation of the myosin regulatory light chain and was partially dependent on tumor necrosis factor alpha and myosin light-chain kinase but not interleukin-1β. Neither heat-killed H. somnus, formalin-fixed H. somnus, nor purified lipooligosaccharide altered monolayer integrity within a 2-h incubation period, whereas conditioned medium from H. somnus-treated BBEC caused a modest reduction in TEER. The data from this study support the hypothesis that viable H. somnus alters integrity of the blood-brain barrier by promoting contraction of BBEC and increasing paracellular permeability of the CNS vasculature.

Role of thrombin in endothelial cell monolayer repair in vitro

1994 ◽  
Vol 20 (4) ◽  
pp. 621-628 ◽  
Author(s):  
Paul J. DiMuzio ◽  
Kerri J. Pratt ◽  
Pauline K. Park ◽  
R.Anthony Carabasi
Keyword(s):  
Endothelial Cell ◽  
Cell Monolayer ◽  
Endothelial Cell Monolayer

Characterization of an In Vitro Model to Study the Permeability of Human Arterial Endothelial Cell Monolayers

1988 ◽  
Vol 60 (02) ◽  
pp. 240-246 ◽  
Author(s):  
Erna G Langeler ◽  
Victor W M van Hinsbergh
Keyword(s):  
Endothelial Cell ◽  
Calcium Ions ◽  
Electrical Resistance ◽  
Serum Proteins ◽  
Cell Monolayer ◽  
Extracellular Calcium ◽  
Endothelial Cell Monolayer ◽  
Passage Rate ◽  
Cell Monolayers

SummaryA model has been developed to study the transport of fluid and macromolecules through human arterial umbilical cord endothelial cell monolayers in vitro. Cells were cultured on fibronectin- coated polycarbonate filters and formed within a few days a tight monolayer, with an electrical resistance of 17 ± 4 Ohm · cm2. The cells were connected by close cell contacts with tight junctions. The passáge-rate of horse radish peroxidase (HRP) through these filters was 20-40 fold lower than through filters without an endothelial monolayer. The continuous presence of 10% human serum was needed to maintain the electrical resistance of the monolayer and its barrier function towards macromolecules. Chelation of extracellular calcium resulted in an increased permeability and a decreased electrical resistance of the monolayer. This process was reversible by re-addition of calcium ions to the cells. The permeation rate of dextrans of various molecular weights (9-480 kD) was inversely related to the molecular mass of the molecule. No difference was measured between the passage rate of dextran of 480 kD and dextran of 2,000 kD. Incubation of the endothelial cell monolayer with 2-deoxy-D-glucose resulted in a decreased permeability but it had no effect on electrical resistance. This suggests that the passage-process is energy- dependent.Fluid permeation through the endothelial cell monolayer on filters was measured in a perfusion chamber under 20 mmHg hydrostatic pressure. It was decreased by the presence of serum proteins and responded reversibly on the chelation and readdition of extracellular calcium ions.

scholarly journals Platelet aggregation in flowing blood at a site of injury to an endothelial cell monolayer: quantitation and real-time imaging with the TAB monoclonal antibody

Blood ◽  
1990 ◽  
Vol 75 (2) ◽  
pp. 390-398
Author(s):  
EF Grabowski
Keyword(s):  
Monoclonal Antibody ◽  
Endothelial Cell ◽  
Real Time ◽  
Platelet Adhesion ◽  
Cell Monolayer ◽  
Thromboxane B2 ◽  
Endothelial Cell Monolayer ◽  
Lysine Acetylsalicylate ◽  
Flowing Blood ◽  
A Site

Epifluorescence videomicroscopy permits real-time imaging of platelet adhesion/aggregation to a defined microinjury of a monolayer of endothelial cells exposed to flowing blood. The fluorescent label is the TAB murine monoclonal antibody directed against human platelet GP IIB, together with a fluorescein-conjugated goat F(ab')2 against murine immunoglobulin. The combination assures specificity for platelet membranes, yet leaves platelet function intact. TAB is first added to gently mixed, citrated human blood; the second antibody is added 1 hour after the first, mixing continuing for a second hour. Bovine aortic endothelial cell monolayers (ECMs), grown on rectangular cover glasses precoated with microfibrillar collagen, comprise one wall of a flow chamber mounted on a vertical microscope stage. A loop of 6–0 sterile suture is drawn across the ECM in order to create microinjuries of width 70 +/- 15 microns (mean +/- SD) oriented in a direction transverse to flow. Platelet adhesion/aggregation is virtually absent on intact and confluent regions of the monolayer. On micro-injury sites and at shear rates of 60 to 1,080 sec-1, however, computer-enhanced images obtained by means of videomicroscopy show arrival and adherence of single platelets resulting in the formation of platelet aggregates elongated in the flow direction. When the monolayers are pretreated with 1.0 mmol/L lysine acetylsalicylate, the mean aggregate thickness increases (2P less than .05) to 260 +/- 58% (mean +/- SE, N = 6) of control, aggregates are regularly shed downstream, and the surface area of the injury site covered by platelets is augmented (2P less than .05) from 14.8 +/- 3.9% to 49.2 +/- 4.7% (mean +/- SE, N = 6). Donor ingestion of aspirin, on the other hand, leads to an increase (2P less than .01) in percent surface coverage to 42.7 +/- 8.5 without a concomitant increase in mean aggregate thickness. In parallel with the above, outflow levels of serum thromboxane and prostacyclin are measured by radioimmunoassays (RIAs) for thromboxane B2 and 6-Keto-PGF1 alpha, respectively. Thromboxane B2 is increased (2P less than .01) by monolayer pretreatment with lysine acetylsalicylate from 5.08 +/- 1.47 to 9.35 +/- 2.42, but decreased (2P less than .05) after oral aspirin to 1.21 +/- 0.38 ng/mL (mean +/- SE, N = 6). Levels of 6-Keto-PGF1 alpha were reduced (2P less than .05) by monolayer pretreatment from 0.48 +/- 0.046 to 0.36 +/- 0.016 ng/mL. Platelet adhesion/aggregation at a site of injury to an endothelial cell monolayer, therefore, can be imaged in flowing blood in real time using a monoclonal antibody approach.(ABSTRACT TRUNCATED AT 400 WORDS)

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