SEARCH · PubMed Health

Results for “Extracellular vesicles”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 721 records · Page 40Linked to original sources

PubMed1975

Structure of cultured fibroblasts from dermatosparaxic calves.

Clonal cell lines from the dermis of a dermatosparaxic calf were grown in tissue culture. After fixation in a mixture of glutaraldehyde and osmium, they were prepared for electron microscopy. Most cells contained a well-developed Golgi region, lysosomes, mitochondria, and dilated cisternae of rough endoplasmic reticulum. They also contained numerous, large bundles of intracellular fialments, many lipid droplets and extensive arrays of vesicles. Cultures accumulated substantial amounts of extracellular fibrillar material. The fibrils were loosely packed and indistinctly cross-banded. Bundles of intracellular filaments were commonly parallel in adjacent cells and also parallel to extracellular fibrils. These cytoplasmic features may result from the inability of the secreted collagen to form normal fibrils.

Animals↗

PubMed1989

Ultrastructural demonstration of increased sulfated proteoglycans and calcium associated with chondrocyte cytoplasmic processes and matrix vesicles in rat growth plate cartilage.

A monoclonal antibody (3-B-3) to chondroitin 6-sulfated proteoglycan was used with immunoperoxidase electron microscopy to study the relationship of chondrocyte cytoplasmic processes and matrix vesicles in rat epiphyseal growth plate cartilage. Immunoperoxidase staining of the chondrocyte plasmalemma was found at all levels in the growth plate and was most prominent in the hypertrophic zone. The plasmalemma and matrix of the cytoplasmic process often demonstrated stronger reactivity than the remainder of the cell surface. Matrix vesicles showed weak to strong surface or internal reactivity. The majority of them stained very similarly to the cytoplasmic process. X-ray microanalysis of specimens processed by rapid freezing and freeze substitution confirmed that both sulfur and calcium were localized within or in close association with both the cytoplasmic process and the matrix vesicle, suggesting a chemical combination of calcium with sulfated proteoglycans at both sites. These results indicate that there is a selective increase in the concentration of membrane-associated sulfated proteoglycan and calcium in the cell process, from which matrix vesicles may be released into the extracellular matrix.

Animals↗

PubMed2005

Calcium increases endocytotic vesicle size and accelerates membrane fission in insulin-secreting INS-1 cells.

In many cells, endocytotic membrane retrieval is accelerated by Ca2+. The effect of Ca2+ on single endocytotic vesicles and fission pore kinetics was examined by measuring capacitance and conductance changes in small membrane patches of insulin-secreting INS-1 cells. In intact cells, elevation of Ca2+ by glucose stimulation induced a 1.8-fold increase in membrane internalisation. This surprisingly resulted from an increased unitary capacitance of endocytotic vesicles whereas the frequency of endocytosis was unaltered. This effect of glucose was prevented by inhibition of L- or R-type Ca2+ channels. Extracellular (pipette) Ca2+ was found to regulate endocytotic vesicle capacitance in a bimodal manner. Vesicle capacitance was increased at intermediate Ca2+ (2.6 mM), but not at high Ca2+ (10 mM). Similar results were obtained upon direct application of 100 nM and 0.5 mM Ca2+ to the intracellular surface of inside-out excised membrane patches, and in these experiments the increase in vesicle capacitance was prevented by the calcineurin inhibitor deltamethrin. Endocytotic fission pore kinetics were accelerated by Ca2+ in both the intact cells and isolated membrane patches; however, the effect in this case was neither bimodal nor deltamethrin sensitive. Membrane retrieval can therefore be upregulated by a Ca2+-dependent increase in endocytotic vesicle size and acceleration of membrane fission in insulin-secreting INS-1 cells.

Animals↗

PubMed1997

Fluid-phase uptake by macropinocytosis in Dictyostelium.

To study fluid-phase endocytosis in living cells and its relationship to changes in the cell cortex, we have used a green fluorescent protein (GFP)-tagged version of coronin, an actin-associated protein that localises to dynamic regions of the Dictyostelium cell cortex. In the confocal microscope, internalisation of fluorescently labelled dextran as a fluid-phase marker can be recorded simultaneously with the recruitment of the coronin-GFP fusion-protein from the cytoplasm of the phagocyte. At crown-shaped surface protrusions, extracellular medium is taken up into vesicles with an average diameter of 1.6 microns, which is significantly larger than the 0.1 microns diameter of clathrin-coated pinosomes. The observed frequency of macropinosome formation can account for a large portion, if not all, of the fluid-phase uptake. The redistribution of coronin-GFP strongly resembles cytoskeletal rearrangements during phagocytosis. Scanning-electron micrographs indicate that crown-shaped cell-surface extensions can undergo shape changes, without a particle bound, that are similar to shape changes that occur during phagocytosis. In quantitative assays, the uptake of particles and fluid are about equally dependent on F-actin and coronin.

Actins↗

PubMed1992

Freeze-fracture and lanthanum studies of the retinal microvasculature in diabetic rats.

To see whether or not blood-retinal barrier breakdown during diabetes was associated with breakdown of the endothelial cell tight junctions or with other membrane alterations in the cells comprising the wall of the retinal microvasculature, streptozotocin-induced diabetic rat retinas were studied using lanthanum tracer and freeze-fracture electron microscopic morphometry. This study showed that endothelial cell tight junction permeability to lanthanum and luminal surface area were normal in these diabetic rats. However, freeze-fracture morphometry showed several alterations in the diabetic retinal microvessels. First, the endothelial cell membranes had abnormally large (80-120 nm) plasmalemmal vesicles not evident in the control retinas, suggesting that membrane turnover was abnormal. Second, endothelial cell P-face membranes at the blood front contained more larger particles than those in the control rats (P less than 0.05), implying an alteration in endothelial cell luminal membrane composition. Third, endothelial cell P-face membranes in areas of close apposition with pericyte membranes showed abnormal areas of particle clearing not seen in the control animals, suggesting a change in pericyte-endothelial cell interactions. Finally, pericyte membranes facing the neural retina contained increased numbers of plasmalemmal vesicles compared with control membranes (P less than 0.05). Moreover, the association of these vesicles with collagen fibrils in the extracellular space suggested an alteration in extracellular matrix turnover.

Animals↗

PubMed1987

Cytosolic free calcium levels in sickle red blood cells.

In this study, we used a recently developed nuclear magnetic resonance (NMR) technique to measure ionized calcium in sickle erythrocytes. The NMR technique, which involves 19F NMR studies of a fluorinated calcium chelator quinMF, [2-(2-amino-4-methyl-5-fluorophenoxy)methyl-8-aminoquinoline-N,N,N',N'- tetraacetic acid] provides a novel approach to the study of ionized calcium in erythrocytes since the presence of hemoglobin precludes the use of fluorescent calcium indicators. The mean value for ionized calcium in oxygenated sickle erythrocytes was 18 +/- 2 nmol/L (SE). Experiments with normal RBCs gave a mean value of 21 +/- 2 nmol/L (SE). After 1 hour of deoxygenation, mean values for ionized calcium in sickle erythrocytes did not increase as compared with values obtained under oxygen. To investigate whether deoxygenation stimulated endocytosis, sickle erythrocytes were deoxygenated for 1 hour in the presence of impermeant FBAPTA (1,2 bis-(2-amino-5-fluorophenoxy) ethane N,N,N',N'-tetraacetic acid). Cells were then separated from the extracellular medium and assayed for the presence of FBAPTA; they had incorporated significant quantities of the extracellular FBAPTA. This incorporation was not observed with normal erythrocytes. These data are consistent with at least a portion of the elevation in total cell calcium in sickle erythrocytes arising as a consequence of an endocytotic process in which extracellular calcium ions are incorporated into vesicles. Additional experiments show that these intracellular vesicles accumulate Ca2+ on further deoxygenation, consistent with a transient increase in ionized cell calcium. These studies represent the first use of NMR spectroscopy to evaluate endocytotic processes.

Aminoquinolines↗

PubMed1988

A new long-term in vitro invasion assay using fibrous connective tissue matrices maintaining architectural characteristics of connective tissue.

A long-term invasion assay using fibrous connective tissue matrices was developed. The matrices were prepared by treating murine skin or human dura mater with 25 mM ammonium hydroxide containing proteinase inhibitors at 4 degrees C for 7 days. They could be maintained almost indefinitely without the degeneration and necrosis. Electron micrographs of them revealed the preservation of native collagen fibers, and sequential enzyme digestion showed the presence of glycoprotein in the matrices. Local dissolution of extracellular matrix by cultured human rectal adenocarcinoma cell line, RCM-1, was observed morphologically and confirmed by a quantitative assay using radiolabeled matrices. The destruction of extracellular matrix occurred associated with membrane vesicle-shedding from the cells. Both the advantages and disadvantages of this assay were discussed.

Animals↗

PubMed1972

The cellular pathology of experimental hypertension. VI. Alterations in retinal vasculature.

The ultrastructure of retinal arterial vessels from rats with severe renal hypertension has been studied. The permeability of retinal vessels has also been examined by means of vascular labeling technics utilizing horseradish peroxidase and microperoxidase as tracer substances. Small retinal arteries and arterioles exhibit foci of smooth muscle necrosis characterized initially by fragmentation of medial smooth muscle cells, and subsequently by loss of myofilaments and release of free vesicles, vacuoles and other cytoplasmic organelles extracellularly. Evidence for increased permeability is observed occasionally in retinal capillaries and less frequently in arteries and arterioles. The enzymatic tracers penetrate the tight junctions of the endothelial cells and are found in the basement membranes adjacent to endothelial and smooth muscle cells, as well as in expanded extracellular spaces around the capillaries. The alterations in the ultrastructure and permeability of retinal vessels in experimental hypertension have been compared with that of visceral and cerebral cortical vessels.

Animals↗

PubMed1983

The role of nucleoside triphosphate pyrophosphohydrolase in in vitro nucleoside triphosphate-dependent matrix vesicle calcification.

Nucleoside triphosphate pyrophosphohydrolase (EC 3.6.1.8) activity is associated with matrix vesicles purified from collagenase digests of fetal calf epiphyseal cartilage. This enzyme hydrolyzes nucleoside triphosphates to nucleotides and PPi, the latter inducing precipitation in the presence of Ca2+ and Pi. An assay for matrix vesicle nucleoside triphosphate pyrophosphohydrolase is developed using beta, gamma-methylene ATP as substrate. The assay is effective in the presence of matrix vesicle-associated ATPase, pyrophosphatase, and alkaline phosphatase activities. A soluble nucleoside triphosphate pyrophosphohydrolase is obtained from matrix vesicles by treatment with 5 mM sodium deoxycholate. The solubilized enzyme induced the precipitation of calcium phosphate in the presence of ATP, Ca2+, and Pi. Extraction of deoxycholate-solubilized enzymes from matrix vesicles with 1-butanol destroys nucleoside triphosphate pyrophosphohydrolase activity while enhancing the specific activities of ATPase, pyrophosphatase, and alkaline phosphatase. In solutions devoid of ATP and matrix vesicles, concentrations of PPi between 10 and 100 microM induce calcification in mixtures containing initial Ca2+ X P ion products of 3.5 to 7.9 mM2. This finding plus the discovery of nucleoside triphosphate pyrophosphohydrolase in matrix vesicles supports the view that these extracellular organelles induce calcium precipitation by the enzymatic production of PPi. Nucleoside triphosphate pyrophosphohydrolase is more active against pyrimidine nucleoside triphosphates than the corresponding purine derivatives. The pH optimum is 10.0 and the enzyme is neither activated nor inhibited by Mg2+ or Ca2+ ions or mixtures of the two. Vmax at pH 7.5 for beta, gamma-methylene ATP is 0.012 mumol of substrate hydrolyzed per min per mg of protein and Km is below 10 microM. The enzyme is irreversibly destroyed at pH 4 and is stable at pH 10.5.

Animals↗

PubMed1994

Cultures of human embryonic osteoblasts. A new in vitro model for biocompatibility studies.

Cell populations derived from human embryonic bone were isolated according to the ability of osteoblasts to migrate from bone onto glass fragments. Morphologic and biochemical assays showed (1) osteoblast-like appearance; (2) elevated alkaline phosphatase 1,25(OH)2D3 responsive activity associated with plasma membranes and matrix vesicles; (3) production of a thick extracellular matrix, mainly composed of Type I collagen, which mineralized in the presence of 10 mM beta-glycerophosphate; and (4) higher growth rate and viability when compared with their mature counterpart. Cultures of embryonic cells were challenged with particles of several biomaterials, and their effects on morphology, vitality, and osteogenic capacity of the cultured cells were tested. Stainless steel, titanium alloy, Co-Cr-Mo alloy (vitallium), carbon fiber-reinforced polybutylene terephtalate, ultra-high molecular weight polyethylene, ceramic, calcium phosphate, and hydroxyapatite did not exert any significant deleterious effects on the cultured human osteoblasts.

Alkaline Phosphatase↗

PubMed1996

Comparison of neurodegenerative pathology in transgenic mice overexpressing V717F beta-amyloid precursor protein and Alzheimer's disease.

Overexpression of mutated human amyloid precursor protein (hAPP717V-->F) under control of platelet-derived growth factor promoter (PDAPP minigene) in transgenic (tg) mice results in neurodegenerative changes similar to Alzheimer's disease (AD). To clarify the pathology of these mice, we studied images derived from laser scanning confocal and electron microscopy and performed comparisons between PDAPP tg mice and AD. Similar to AD, neuritic plaques in PDAPP tg mouse contained a dense amyloid core surrounded by anti-hAPP- and antineurofilament-immunoreactive dystrophic neurites and astroglial cells. Neurons were found in close proximity to plaques in PDAPP tg mice and, to a lesser extent, in AD. In PDAPP tg mice, and occasionally in AD, neuronal processes contained fine intracellular amyloid fibrils in close proximity to the rough endoplasmic reticulum, coated vesicles, and electron-dense material. Extracellular amyloid fibrils (9-11 nm in diameter) were abundant in PDAPP tg and were strikingly similar to those observed in AD. Dystrophic neurites in plaques of PDAPP tg mouse and AD formed synapses and contained many dense multilaminar bodies and neurofilaments (10 nm). Apoptotic-like figures were present in the tg mice. No paired helical filaments have yet been observed in the heterozygote PDAPP tg mice. In summary, this study shows that PDAPP tg mice develop massive neuritic plaque formation and neuronal degeneration similar to AD. These findings show that overproduction of hAPP717V-->F in tg mice is sufficient to cause not only amyloid deposition, but also many of the complex subcellular degenerative changes associated with AD.

Alzheimer Disease↗

PubMed1996

[Biochemical approaches to neuromuscular junction disorders].

In view of the conformation-dependent B-cell epitope and the MHC class II-restricted T-cell epitope, myasthenogenic sites in the molecular structure of acetylcholine receptor alpha-subunit were defined by the induction of a rat model of myasthenia gravis using synthetic peptides, alpha 183-200, alpha 125-147 and alpha 67-76 with alpha 107-116. Structural model for the acetylcholine receptor synthetic peptides were designed to obtain a conformation that was highly antigenic compared with that of natural sequence peptides. To search for antigenic sites in the molecular structure of P/Q type of voltage-gated calcium channel (alpha 1) in Lambert-Eaton myasthenic syndrome (LEMS), LEMS patients' sera were studied by immunoprecipitation assay using antigens synthesized referring to its sequential residue map. Peptides corresponding to extracellular region (S5 S6 linker) of domains II and IV were specifically reactive with LEMS antibodies; their titers respectively correlated with those of anti-P/Q type calcium channel (human cerebellum). The extracellular region of synaptotagmin, a synaptic vesicle protein that acts as a calcium sensor in release of acetylcholine, was also immunogenic in the induction of a rat model of LEMS; a proportion of LEMS patients' sera reacted with recombinant synaptotagmin in immunoblot.

Animals↗

PubMed1998

[Structural differentiation of dissociated skeletal embryonal myocytes of frog under conditions of cell culture].

The development of membrane structures, providing E-C coupling, and the contractile apparatus organization were investigated in frog skeletal myocytes cultured for 1 to 10 days in conditions preventing both myocyte division and fusion. Ruthenium red was used to determine the membranous structures being in contact with the extracellular environment. The marked membrane structures (vesicles and short tubules) appeared to be near the cell membrane on the first days of culturing. The increase in the ratio of the surface area of all internal membranous structures, marked by Ruthenium red, to the external membrane area with aging was proven by morphometric calculations, that means a progressive development. Contractile filaments were found near the cell membrane on the first days of development. Bundles of filaments with initial signs of sarcomere organization were observed on the 3rd-4th days, and myofibrils with highly organized sarcomeres occupied the main part of the sarcoplasm on the 6th day of culturing. The triads appeared also on the sixth day, being regularly inserted into the sarcomere structure. Degenerative signs in the myocytes (sarcomere disorganization and T-tubule swelling) were observed on the 8-10th days, but the area occupied by contractile elements was increased. These results show that the myocyte fusion into myotubules is not a necessary condition for either sarcomere formation, or the formation of all membranous structures providing the E-C coupling.

Animals↗

PubMed1988

Alzheimer's disease: coated vesicles, coated pits and the amyloid-related cell.

The amyloid-related cell (ARC) of the neuritic plaques of Alzheimer's disease revealed numerous cytoplasmic projections surrounding extracellular amyloid material. It is proposed that ARC-coated vesicles fuse with the cell membrane, forming coated pits, which may empty their secretory material into the extracellular space where polymerization of amyloid filaments could occur.

Alzheimer Disease↗

PubMed1988

Effect of alpha-latrotoxin on the frog neuromuscular junction at low temperature.

1. alpha-Latrotoxin (alpha-LTx) was applied to frog cutaneous pectoris muscles bathed at 1-3 degrees C in either Ringer solution, Ca2+-free Ringer solution with 1 mM-EGTA and 4 mM-Mg2+ or Ringer solution plus 4 mM-Mg2+, and its effects on miniature end-plate potential (MEPP) frequency, nerve terminal ultrastructure and uptake of horseradish peroxidase (HRP) were studied. 2. Large concentrations (2 micrograms/ml) of alpha-LTx increased MEPP rates to levels above 100/s at all junctions, but the time course of the increases depended upon the divalent cation content of the bathing solution. However, similar numbers of MEPPs (0.3-0.7 x 10(6] were recorded at all junctions during 2 h of secretion. 3. Nerve terminals exposed to alpha-LTx for 2 h lost 60-75% of their synaptic vesicles and were swollen; their presynaptic membranes were deeply infolded and they often contained many large vesicular structures. Terminals in Ringer solution retained the largest number of synaptic vesicles; terminals in Ringer solution plus Mg2+ swelled the least and contained the largest number of coated vesicles. The average number of synaptic vesicles lost was approximately equal to the average number of MEPPs recorded. 4. Few vesicles became loaded with HRP when this extracellular tracer was present in the bathing solution and the muscles were fixed near the peak of secretion. 5. When the terminals were warmed to 20 degrees C, those in the Ca2+-free solution with Mg2+ secreted additional quanta and lost almost all their residual vesicles; those in Ringer solution without Mg2+ secreted few additional quanta and retained most of their residual vesicles. 6. These results suggest that recycling was blocked at these terminals and that for each quantum secreted a vesicle became permanently incorporated into the axolemma.

Action Potentials↗

PubMed1996

An ultrastructural study of the relationship between sensory trigeminal nerves and odontoblasts in rat dentin/pulp as demonstrated by the anterograde transport of wheat germ agglutinin-horseradish peroxidase (WGA-HRP).

Because the ultrastructure of the trigeminal sensory nerves in dentin, especially in relation to odontoblasts, remains to be clarified, we investigated the relationship between the trigeminal sensory nerves and the odontoblast processes using the anterograde axonal transport technique by injecting wheat germ agglutinin-horseradish peroxidase (WGA-HRP) into the rat trigeminal ganglion. Light microscopically, the nerves labeled with WGA-HRP were mainly concentrated at the pulpal horn, forming a nerve plexus at the subodontoblastic region and penetrating the predentin/dentin about 50 to 70 microns. Ultrastructurally, HRP reaction products were observed intra-axonally in the myelinated (A delta) and unmyelinated (C) axons in the subodontoblastic region. Most nerves lost the Schwann sheath and were naked in the predentin/dentin. The labeled varicosities were close to the odontoblast processes in the dentinal tubules. No synaptic structures could be detected between the varicosities and the odontoblasts, but a gap about 20 nm wide was found between them. One type of varicosity was a rich mitochondria-containing varicosity, while the other was a rich vesicle-containing (large dense core vesicles and small clear vesicles) one. The reaction products were also found in the extracellular spaces surrounding the axons. Sometimes the reaction products were seen in the coated pits or the endocytotic vesicles of the odontoblast processes. The present study demonstrated that nerve endings (varicosities) derived from the trigeminal ganglion were present in the dentinal tubules, and that WGA-HRP extracellularly extruded from the sensory nerves in the odontoblastic layer or predentin/dentin. These findings thus suggest that sensory nerves may have some (e.g., trophic) effect on either odontoblasts or the environment around the sensory nerves in the dentin/pulp.

Afferent Pathways↗

PubMed2005

Genes encoding ADP-ribosylation factors in Arabidopsis thaliana L. Heyn.; genome analysis and antisense suppression.

Vesicle trafficking delivers proteins to intracellular and extracellular compartments, cellulose synthase to the plasma membrane, and non-cellulosic polysaccharides to the cell wall. The Arabidopsis genome potentially encodes 19 proteins with sequence similarities to ARFs (ADP-ribosylation factors) and its relatives such as ARLs (ARF-like proteins). ARFs are essential for vesicle coating and uncoating in all eukaryotic cells, while ARLs play more diverse roles. Nine proteins, six of them highly similar, are possible ARFs, three are putative ARL orthologues and the remainder were designated ARF-related proteins. The functions of the six highly similar, putative ARFs in whole plant development were probed by suppressing their expression with antisense. Antisense plants were severely stunted because cell production rate and final cell size were both reduced. Changed time-to-flowering, apical dominance, and fertility may reflect alterations to hormonal and other signalling pathways with which ARFs may interact. No gross changes in targeting or compartmentalization were seen in antisense plants containing GFP targeted to the ER and Golgi and changes in cell wall composition were limited to increases in some non-cellulosic polysaccharides and a relatively small decrease in cellulose. The reasons why these effects are less drastic than the effects on endomembranes and wall composition that are seen in short-term experiments with brefeldin A and with dominant negative ARF mutants are discussed.

ADP-Ribosylation Factors↗

PubMed2006

Wild-type huntingtin participates in protein trafficking between the Golgi and the extracellular space.

Huntington disease (HD) is an autosomal dominant neurodegenerative disease caused by an expanded CAG trinucleotide repeat in the first exon of the HD gene, which results in a toxic polyglutamine stretch within huntingtin, the protein it encodes. Understanding the normal function of this essential protein is vital to understanding the root of the disease, yet despite more than a decade of investigation, its role in the cell remains elusive. Identifying the subcellular localization of huntingtin and understanding its effects on global gene expression are critical to this endeavor. While most reports agree that huntingtin is predominantly a cytoplasmic protein, conflicting distribution patterns have been demonstrated at the subcellular level. Here, we examine wild-type huntingtin's localization in cultured cells by expressing the full-length human protein tagged with enhanced green fluorescent protein (EGFP) within its unspliced genomic context. In fibrosarcoma and neuroblastoma cells, huntingtin shows discrete punctate, perinuclear localization overlapping largely with the trans-Golgi and cytoplasmic clathrin-coated vesicles, implicating huntingtin in vesicle trafficking. To determine whether huntingtin is involved in trafficking a specific subset of proteins, we measured changes in global transcription levels in embryonic stem cells and neurons lacking huntingtin. Huntingtin null neurons exhibit a significant reduction in transcripts encoding proteins destined for the extracellular space, many of which are components of the extracellular matrix or involved in cellular adhesion, receptor binding and hormone activity. Together, these findings support a role for huntingtin in the intracellular trafficking of proteins required for the construction of the extracellular matrix.

Animals↗