Tissue Factor Coagulant Activity is Regulated by the Plasma Membrane Microenvironment

2018 ◽  
Vol 118 (06) ◽  
pp. 990-1000 ◽  
Author(s):  
Yuanjie Yu ◽  
Anita Böing ◽  
Chi Hau ◽  
Najat Hajji ◽  
Wolfram Ruf ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Plasma Membrane ◽  
Tissue Factor ◽  
Plasma Membranes ◽  
Low Density ◽  
Percoll Gradient ◽  
Protein Disulphide Isomerase ◽  
Coagulant Activity ◽  
Different Types ◽  
Detergent Resistant Membrane

Background Tissue factor (TF) can be present in a non-coagulant and coagulant form. Whether the coagulant activity is affected by the plasma membrane microenvironment is unexplored. Objective This article studies the presence and coagulant activity of human TF in plasma membrane micro-domains. Methods Plasma membranes were isolated from human MIA PaCa2 cells, MDA-MB-231 cells and human vascular smooth muscle cells by Percoll gradient ultracentrifugation after cell disruption. Plasma membranes were fractionated by OptiPrep gradient ultracentrifugation, and the presence of TF, flotillin, caveolin, clathrin, protein disulphide isomerase (PDI), TF pathway inhibitor (TFPI) and phosphatidylserine (PS) were determined. Results Plasma membranes contain two detergent-resistant membrane (DRM) compartments differing in density and biochemical composition. High-density DRMs (DRM-H) have a density (ρ) of 1.15 to 1.20 g/mL and contain clathrin, whereas low-density DRMs (DRM-L) have a density between 1.09 and 1.13 g/mL and do not contain clathrin. Both DRMs contain TF, flotillin and caveolin. PDI is detectable in DRM-H, TFPI is not detectable in either DMR-H or DRM-L and PS is detectable in DRM-L. The DRM-H-associated TF (> 95% of the TF antigen) lacks detectable coagulant activity, whereas the DRM-L-associated TF triggers coagulation. This coagulant activity is inhibited by lactadherin and thus PS-dependent, but seemed insensitive to 16F16, an inhibitor of PDI. Conclusion Non-coagulant and coagulant TF are present within different types of DRMs in the plasma membrane, and the composition of these DRMs may affect the TF coagulant activity.

Alternatively spliced human tissue factor (asHTF) is not pro-coagulant

2007 ◽  
Vol 97 (01) ◽  
pp. 11-14 ◽  
Author(s):  
Petra Censarek ◽  
Anett Bobbe ◽  
Maria Grandoch ◽  
Karsten Schrör ◽  
Artur-Aron Weber
Keyword(s):  
Smooth Muscle ◽  
Tissue Factor ◽  
Human Tissue ◽  
Thrombin Generation ◽  
Expression System ◽  
Hek293 Cells ◽  
Transfected Cells ◽  
Mammalian Expression ◽  
Coagulant Activity ◽  
Alternatively Spliced

SummaryIt has been proposed that alternatively-spliced human tissue factor (asHTF) is pro-coagulant. We have evaluated the function of asHTF in a mammalian expression system. Full-length human tissue factor (HTF) and asHTF were cloned from smooth muscle cells and over-expressed in HEK293 cells. As expected, a marked pro-coagulant activity (FX activation, thrombin generation) was observed on the surface, in lysates, and on microparticles from HTF transfected cells. In contrast, no pro-coagulant activity of as HTF was observed.

scholarly journals Partial purification and characterization of oestrogen receptors in subfractions of hepatocyte plasma membranes

1980 ◽  
Vol 191 (3) ◽  
pp. 743-760 ◽  
Author(s):  
Richard J. Pietras ◽  
Clara M. Szego
Keyword(s):  
Plasma Membrane ◽  
Binding Sites ◽  
Wheat Germ Agglutinin ◽  
Wheat Germ ◽  
Plasma Membranes ◽  
Specific Binding ◽  
Low Density ◽  
Marker Enzymes ◽  
Low Density Plasma ◽  
Density Plasma

To assess the subcellular distribution of oestrogen-binding components in their native state, plasma membrane and other cell fractions were prepared from hepatocytes in the absence of [3H]oestradiol-17β. Cells from livers of ovariectomized rats were disrupted, with submaximal homogenization in buffered isotonic sucrose with CaCl2 and proteinase inhibitor, and fractionated by using isotonic media. Fractions were characterized by determinations of enzyme activities, biochemical constituents and ligand binding. Specific binding of 2nm-[3H]oestradiol-17β to intact cells and their fractions was detemined after equilibration for 1.5h at 4°C. More than 92% of the radioactivity from representative preparations was verified as authentic oestradiol by thin-layer chromatography. Activities of plasma-membrane marker enzymes as well as binding sites for oestrogen and for wheat germ agglutinin were present principally in particulate fractions, rather than in 105000g-supernatant fractions. However, by using alternative homogenization procedures (i.e. hypotonic media), known to fragment and strip structural components, oestradiol-binding sites and activities of plasma-membrane marker enzymes were distributed predominantly into cytosol. By using the more conservative procedures, plasma membranes of low (ρ=1.13–1.16) and high (ρ=1.16–1.18) density were purified from crude nuclear fractions. A second low-density subfraction of plasma membrane was prepared from microsome-rich fractions. Activities of plasma-membrane marker enzymes were enriched to about 28 and four times that of the homogenate in plasma membranes of low and high density respectively. Binding sites for wheat germ agglutinin and oestradiol were concentrated in low-density plasma membranes to 46–63 times that of the homogenate. Specific binding of oestrogen in low-density plasma membranes purified from crude nuclei was saturable, with an apparent association constant of 3.5nm. At saturation, such oestradiol receptors corresponded to 526fmol/mg of membrane protein. A Hill plot showed a moderate degree of positive co-operativity in the interaction of hormone with plasma membranes. Specific binding of [3H]oestradiol-17β was reduced by a 200-fold molar excess of unlabelled oestradiol-17β, oestriol or diethylstilbestrol, but not by oestradiol-17α, cortisol, testosterone or progesterone. Binding was also blocked by prior exposure of membranes to trypsin or to 60°C, but remained essentially undiminished by extraction of membranes with either hypotonic or high-salt buffers. Extraction with 0.1% (v/v) Triton X-100 partially solubilized the oestrogen-binding component(s) of plasma membranes. Particle-free extracts were resolved on 5–20% (w/v) sucrose density gradients with either 0.01m- or 0.4m-KCl, and the fractions were analysed by adsorption to hydroxyapatite. In low-salt gradients macromolecule-bound oestrogen sedimented at predominantly 7.4S and binding was 1560 times that of the homogenate. Under high-salt conditions oestradiol-binding activity occurred at both 3.6S and 4.9S.

Electron microscopy and x-ray microanalysis of calcium-binding sites of the plasma membrane of Beroe giant smooth muscle fibre

1982 ◽  
Vol 55 (1) ◽  
pp. 353-364
Author(s):  
G. Nicaise ◽  
M.L. Hernandez-Nicaise ◽  
L. Malaval
Keyword(s):  
Smooth Muscle ◽  
Plasma Membrane ◽  
Calcium Binding ◽  
Plasma Membranes ◽  
Calcium Content ◽  
Muscle Fibres ◽  
Fixed Tissue ◽  
X Ray ◽  
Wavelength Dispersive Spectrometry ◽  
High Calcium

When they are fixed with glutaraldehyde in the presence of calcium ions, the plasma membranes of Beroe giant smooth muscle fibres display micropapillae filled with an electron-dense deposit. After freeze-fracturing of fixed tissue, the micropapillae are still present, therefore their shape and size are determined during or before glutaraldehyde fixation, and are not due to rearrangement during subsequent steps of tissue processing; intramembranous particles are seen at the periphery rather than at the top of micropapillae. In conventional stained sections, the surface of most micropapillae is surrounded by fine fibrils; when the fuzzy coat is separated from the muscle cell by a clear space, this fibrillar material becomes conspicuous and links the micropapillae to the coat. After calcium-free (EGTA) fixation, the plasma membrane is completely free of electron-dense sites but “empty' micropapillae can be seen. X-ray microanalysis of single electron-dense deposits by wavelength-dispersive spectrometry reveals a high calcium content. A weak osmiophily is suspected, but does not seem to interfere with this analysis of calcium. The highest peak-to-background ratios for calcium were obtained using the smallest aperture of the Wehnelt of the analytical microscope. In the Discussion, the micropapillae are compared to similar structures described by other authors in a variety of cell types.

scholarly journals A CHOLINERGIC COMPONENT IN THE INNERVATION OF THE LONGITUDINAL SMOOTH MUSCLE OF THE GUINEA PIG VAS DEFERENS

1969 ◽  
Vol 41 (2) ◽  
pp. 462-476 ◽  
Author(s):  
Peter M. Robinson
Keyword(s):  
Smooth Muscle ◽  
Plasma Membrane ◽  
Guinea Pig ◽  
Smooth Muscle Cells ◽  
Plasma Membranes ◽  
Vas Deferens ◽  
Muscle Cells ◽  
Muscle Membrane ◽  
Smooth Muscle Tissue ◽  
Longitudinal Smooth Muscle

Acetylcholinesterase (AChE) has been detected on the plasma membrane of about 25% of the axons in the longitudinal smooth muscle tissue of guinea pig vas deferens. These axons are presumably cholinergic. No enzyme was detected in the remaining 75% of axons. These axons are presumably adrenergic. The plasma membrane of the Schwann cells associated with the cholinergic axons also stained for AChE. Some axon bundles contained only cholinergic or adrenergic axons while others contained both types of axon. When a cholinergic axon approached within 1100 A of a smooth muscle cell, there was a patch of AChE activity on the muscle membrane adjacent to the axon. It is suggested that these approaches are the points of effective transmission from cholinergic axons to smooth muscle cells. Butyrylcholinesterase activity was detected on the plasma membranes of all axons and smooth muscle cells in this tissue.

scholarly journals ARCHITECTURE AND NERVE SUPPLY OF MAMMALIAN SMOOTH MUSCLE TISSUE

1957 ◽  
Vol 3 (6) ◽  
pp. 867-878 ◽  
Author(s):  
Rudolf Caesar ◽  
George A. Edwards ◽  
Helmut Ruska
Keyword(s):  
Endoplasmic Reticulum ◽  
Smooth Muscle ◽  
Plasma Membrane ◽  
Smooth Muscle Cells ◽  
Muscle Cell ◽  
Muscle Tissue ◽  
Plasma Membranes ◽  
Muscle Cells ◽  
Autonomic Nerves ◽  
Smooth Muscle Tissue

Smooth muscle tissue from mouse urinary bladder, uterus, and gall bladder has been studied by means of the electron microscope. The smooth muscle cells are distinctly and completely separated from each other by a cytolemma comparable to the sarcolemma of striated muscle. The tissue is thus cellular and not syncytial. With this evidence, supported by electron microscopy of other tissues, we question the existence of true syncytia in animal tissues. Individual cell membranes necessary for the electrophysiologic events exist in smooth muscle, and its nerve and conduction in a tissue such as uterus or bladder can occur at the cellular level as well as at the tissue area level. The smooth muscle cell contains myofilaments, nucleus, endoplasmic reticulum, mitochondria, Golgi complex, centrosome, and pinocytotic vesicles. These structures are described in some detail, and their probable interrelations and functions are discussed. The autonomic nerves innervating smooth muscle cells are composed of axons and lemnoblasts. The axon is suspended by the mesaxon formed by the infolded plasma membrane of the lemnoblast. The respective plasma membranes separate axon and lemnoblast from each other and from surrounding muscle cells. The axons of autonomic nerves never penetrate the plasma membrane of the muscle cell, but pass or intrude into muscle cell pockets, forming a contact between axonal plasma membrane and smooth muscle plasma membrane. The lemnoblast shows well developed endoplasmic reticulum with Palade granules, mitochondria, and a long, elliptical nucleus. The axon contains neurofilaments, mitochondria, and synaptic vesicles; the quantity of the latter two being significantly greater in the periphery of lemnoblasts and near axon-muscle contact regions. We regard the contact regions as the synapses between the autonomic nerves and the smooth muscle cells.

Differing Procoagulant Activities in the Extracted Lipids of High and Low Density Platelet Plasma Membranes

1972 ◽  
Vol 28 (02) ◽  
pp. 206-212 ◽  
Author(s):  
A. J Barber ◽  
D. C Triantaphyllopoulos ◽  
G. A Jamieson
Keyword(s):  
Plasma Membrane ◽  
Plasma Membranes ◽  
Lipid Fraction ◽  
Anticoagulant Effect ◽  
Low Density ◽  
Platelet Factor ◽  
Functional Differences ◽  
Lipid Fractions ◽  
High Concentrations ◽  
Generation Test

SummaryPlatelet factor 3 activity has been examined in the high (d,1.120) and low (d,1.090) density platelet membranes obtained by the glycerol-lysis technique, and in the lipid fractions extracted from them. The following observations were made : procoagulant activity, as determined by the thromboplastin generation test, was equal in the two types of membrane and was higher than in the intracellular fractions ; the lipid fraction extracted from the low density membranes had a higher activity than that from the high density membranes; at high concentrations this extracted phospholipid had a mild anticoagulant effect. These results demonstrate functional differences between the two types of platelet membrane and favor a plasma membrane origin for platelet factor 3.

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