Studies On Plasma Membranes

1967 ◽  
Vol 2 (4) ◽  
pp. 499-512
Author(s):  
E. L. BENEDETTI ◽  
P. EMMELOT
Keyword(s):  
Sialic Acid ◽  
Tight Junctions ◽  
Plasma Membranes ◽  
Iron Hydroxide ◽  
Chelating Agent ◽  
Colloidal Iron ◽  
Outer Leaflet ◽  
Liver Membranes ◽  
Junctional Complexes ◽  
Pre Treatment

Plasma membranes were isolated from rat liver and a transplanted rat hepatoma of the hepatocellular type. After glutaraldehyde fixation the membranes were treated with colloidal iron hydroxide (CIH) at pH 1.7, which was found to react specifically with the neuraminidase-sensitive sialic acid of the liver membranes. The CIH-reactive, neuraminidase-sensitive sialic acid, comprising 70% of the membrane-bound sialic acid, was exclusively located in the outer leaflet of the liver membranes as shown by the rather regular distribution of electron-dense CIH granules. This granular, asymmetric type of staining was also observed in the hepatoma membranes, which contained some 50% more sialic acid than did the liver membranes. In addition, the hepatoma membranes showed an intense and uniform staining by CIH of short segments of both membrane leaflets; the latter type of staining was but little impaired by neuraminidase pre-treatment. None of the junctional complexes of the liver membranes was stained by CIH. Tight junctions were very rarely observed in the hepatoma membrane preparations, and the desmosomes and intermediate junctions of these membranes not infrequently exhibited a loosened appearance exposing CIH-reactive neuraminidase-sensitive sialic acid at their opposite plates. This aspect could be induced in the desmosomes and intermediate junctions, but not in the tight junctions, by pre-treatment of the liver membranes with the chelating agent ethylenediaminetetra-acetate.

Visualization of Charged Groups on the Surface of Rat Liver Nuclei

1974 ◽  
Vol 16 (3) ◽  
pp. 665-675
Author(s):  
ISMO VIRTANEN ◽  
JORMA WARTIOVAARA
Keyword(s):  
Rat Liver ◽  
Plasma Membranes ◽  
Iron Hydroxide ◽  
Nuclear Surface ◽  
Colloidal Iron ◽  
Rat Liver Nuclei ◽  
Ph Values ◽  
Liver Nuclei ◽  
Pre Treatment ◽  
Anionic Groups

Anionic groups on the outer surfaces of isolated rat liver nuclei were rendered visible in the electron microscope by staining with colloidal iron hydroxide at different pH values. At pH 1.8 the nuclei did not adsorb particles of stain, although plasma membranes left in the same preparation showed heavy labelling. After pretreatment with neuraminidase at pH 6 the plasma membranes were no longer stained. At pH 3.0 the nuclear surfaces also stained intensely. The staining pattern acquired at this pH did not appear to be changed by neuraminidase pre-treatment. With the staining method used, rat liver nuclear surfaces seemed to have no exposed sialic acid under isolation conditions which preserve the nuclear membranes and leave the ribosomes attached to the nuclear surface. However, at higher pH values other anionic groups seem to become dissociated and are stained with colloidal iron hydroxide.

Histochemical and Biochemical Demonstration of Sialic Acid and Sulphate in Vesicles and Membranes Isolated from Nerve Endings of Rat Brain

1973 ◽  
Vol 13 (1) ◽  
pp. 237-255
Author(s):  
RUTH MARX ◽  
ELKE GRAF ◽  
W. WESEMANN
Keyword(s):  
Sialic Acid ◽  
Rat Brain ◽  
Sulphuric Acid ◽  
Nerve Ending ◽  
Synaptic Vesicles ◽  
Iron Hydroxide ◽  
Acid Methyl Ester ◽  
Nerve Endings ◽  
Uniform Size ◽  
Colloidal Iron

The reaction of colloidal iron hydroxide (CIH) with acidic groups was applied for an ultra-structural study of the distribution of sulphuric acid monoesters and sialic acid in synaptic vesicles and external nerve ending membranes isolated from rat brain. At pH 1.7 CIH was precipitated as electron-dense granules with a uniform size of 6-7 nm specifically labelling the carboxyl group of sialic acid and the sulphate group of monoesters of sulphuric acid. The differentiation of these 2 groups was achieved by treatment with neuraminidase and methylation followed by saponification. After preincubation with neuraminidase, which released 90-100% of the sialic acid from the membranes of the synaptic vesicles and the nerve endings, the electron-dense deposits marked the reaction sites of sulphate with CIH. The sulphate groups which were present at a concentration of 2.3 and 2.2 µmol/mg protein for the synaptic vesicle and nerve ending membrane preparations, respectively, were rendered soluble as methyl monosulphate by trans-esterification with acid/methanol and quantitatively removed from the structures. By this treatment membrane-bound sialic acid was blocked as sialic acid methyl ester and partly split off by acid hydrolysis. About 55% of the sialic acid found in the nerve ending membranes remained attached to the structure as compared with about 35% of the sialic acid of the vesicles. The acid-resistant proportion of the sialic acid could be localized with CIH after saponification of the esterified preparations. The method described allows the electron-microscopical demonstration of acid-resistant, neuraminidase-sensitive sialic acid in synaptic structures and the discrimination from sulphated mucopolysaccharides.

Intramembranous Fibrils at Tight Junctions

1970 ◽  
Vol 28 ◽  
pp. 108-109 ◽  
Author(s):  
Ronald S. Weinstein ◽  
N. Scott McNutt ◽  
Surl L. Nielsen ◽  
Vivian W. Pinn
Keyword(s):  
Tight Junctions ◽  
Plasma Membranes ◽  
Central Plane ◽  
Thin Sections ◽  
Membrane Complex ◽  
Outer Leaflet ◽  
Membrane Surfaces ◽  
Zonula Occludens ◽  
Extracellular Compartment ◽  
Fibrillar Component

The tight junction (zonula occludens) forms a linear region of contact between plasma membranes of adjacent epithelial cells near their juxtaluminar borders. This junction is a seal that prevents free diffusion between luminal and adluminal aspectsoof epithelia. At tight junctions in thin sections, the two apposed 75 Å plasma membranes join to form a membrane complex 140 Å in thickness. There is apparent “fusion” of the membranes with loss of outer leaflet material of one or both of the membranes. Previous interpretations of freeze-cleaved (and etched) tight junctions have been based on the Moor-Muhlethaler hypothesis that true membrane surfaces are revealed by cleaving. However, there is now compelling evidence that the surfaces revealed by cleaving are generated by splitting frozen membranes along a central plane thereby creating two new surfaces: Face A oriented toward the extracellular compartment and Face B directed toward the cytoplasm. We now describe a distinctive intramembranous fibrillar component in plasma membranes at tight junctions as revealed by membrane splitting.

The Fine Structure of Glycocalyx in Human Spermatozoa. A High Resolution Cytochemical Study

1973 ◽  
Vol 31 ◽  
pp. 640-641
Author(s):  
P. Hernández-Jáuregui ◽  
A. Sosa ◽  
A. González Angulo
Keyword(s):  
Negative Charge ◽  
Iron Hydroxide ◽  
Human Spermatozoa ◽  
Surface Coat ◽  
Cell Surfaces ◽  
Colloidal Iron ◽  
Cytochemical Study ◽  
Specific Permeability ◽  
Extracellular Glycoprotein ◽  
Mammalian Spermatozoa

Glycocalyx is the name given by Bennett to the extracellular glycoprotein coat present in some cell surfaces. It appears to play an important role in cell properties such as antigenicity, cell adhesivity, specific permeability, and ATP ase activity. In the sperm this coat can be directly related to such important phenomena as capacitation and fertilization. The presence of glycocalyx in invertebrate spermatozoa has already been demonstrated. Recently Yanagimachi et al. has determined the negative charges on sperm surfaces of mammalian spermatozoa including man, using colloidal iron hydroxide. No mention was made however of the outer surface coat as composed of substances other than those confering a negative charge. The purpose of this work was therefore to determine the presence of a glycocalyx in human spermatozoa using alcian blue and lanthanum staining.

Freeze-etch observations on the tight junctions of sertoli cells grown in organ culture with FSH

1978 ◽  
Vol 36 (2) ◽  
pp. 340-341
Author(s):  
Rita Meyer ◽  
Zoltan Posalaky ◽  
Dennis Mcginley
Keyword(s):  
Organ Culture ◽  
Tight Junctions ◽  
Sertoli Cell ◽  
Germ Cells ◽  
Sertoli Cells ◽  
Barrier Function ◽  
Cell Junction ◽  
Intramembranous Particles ◽  
Junctional Complexes ◽  
Oriented Parallel

The Sertoli cell tight junctional complexes have been shown to be the most important structural counterpart of the physiological blood-testis barrier. In freeze etch replicas they consist of extensive rows of intramembranous particles which are not only oriented parallel to one another, but to the myoid layer as well. Thus the occluding complex has both an internal and an overall orientation. However, this overall orientation to the myoid layer does not seem to be necessary to its barrier function. The 20 day old rat has extensive parallel tight junctions which are not oriented with respect to the myoid layer, and yet they are inpenetrable by lanthanum. The mechanism(s) for the control of Sertoli cell junction development and orientation has not been established, although such factors as the presence or absence of germ cells, and/or hormones, especially FSH have been implicated.

scholarly journals Characterization of the binding of plasminogen activators to plasma membranes from human liver

1992 ◽  
Vol 287 (3) ◽  
pp. 911-915 ◽  
Author(s):  
G Nguyen ◽  
S J Self ◽  
C Camani ◽  
E K O Kruithof
Keyword(s):  
Human Liver ◽  
Plasma Membranes ◽  
Gel Filtration ◽  
Mannose Receptor ◽  
Hepatoma Cells ◽  
Tissue Type ◽  
High Affinity ◽  
Liver Membranes ◽  
Membrane Bound ◽  
Pai 1

The binding of tissue-type plasminogen activator (t-PA) to membranes prepared from human liver was investigated, and a specific, saturable, high-affinity binding site (Kd = 3.4 nM) was identified. The binding of t-PA to liver membranes was not affected by an excess of D-mannose or D-galactose, or by active urokinase (u-PA), whereas binding of t-PA to membranes prepared from human HepG2 hepatoma cells was inhibited by u-PA. HepG2-membrane-bound t-PA was fully complexed to PA inhibitor 1 (PAI-1), whereas liver-membrane-bound t-PA was not complexed. Gel filtration on Sephacryl S300 of membrane proteins solubilized in deoxycholate revealed that high-affinity t-PA binding activity elutes at an apparent molecular mass of 40 kDa. Monoclonal antibodies specific for the growth factor and the kringle 2 domains inhibited the binding of t-PA to liver membranes and the catabolism of t-PA by rat hepatoma cells. Human liver membranes also bound u-PA; binding was inhibited by pro-u-PA, the N-terminal fragment of u-PA, but not by the 33 kDa form of u-PA or by t-PA. Our results show that human liver membranes contain a specific 40 kDa binding protein for t-PA that is different from the PAI-1-dependent receptor described on HepG2 cells and the mannose receptor isolated from human liver.

scholarly journals Negative charge distribution and density on the surface of oxygenated normal and sickle red cells

Blood ◽  
1981 ◽  
Vol 57 (4) ◽  
pp. 675-678
Author(s):  
LJ Clark ◽  
LS Chan ◽  
DR Powars ◽  
RF Baker
Keyword(s):  
Charge Distribution ◽  
Electron Micrographs ◽  
Negative Charge ◽  
External Surface ◽  
Iron Hydroxide ◽  
Red Cells ◽  
Glutaraldehyde Fixation ◽  
Colloidal Iron ◽  
Membrane Area ◽  
Significant Difference

Negative charges on the external surface of red cells were visualized by colloidal iron hydroxide labelling of 50% of the membrane area after osmotic hemolysis and glutaraldehyde fixation. Counts were made over randomly selected areas on electron micrographs at 350,000 x magnification. Statistical analyses showed that at the 95% level of confidence there was no significant difference between oxygenated normal (AA) and sickle (SS) cells in either the distribution or the density of negative charges.

Expression of anionic sites on tumour cells at different stages of tissue invasion in vivo: a comparative study by X-ray microanalysis

1989 ◽  
Vol 94 (3) ◽  
pp. 561-566
Author(s):  
P.M. Evans ◽  
D.K. Suker ◽  
I. ap Gwynn
Keyword(s):  
Iron Hydroxide ◽  
Tumour Cells ◽  
Ehrlich Carcinoma ◽  
Similar Degree ◽  
Anionic Sites ◽  
Colloidal Iron ◽  
Cell Behaviour ◽  
X Ray ◽  
Invasion Stage

Quantification of colloidal iron hydroxide (CIH) labelling by X-ray microanalysis was used to investigate anionic sites at the surface of Ehrlich carcinoma cells from different locations in the mouse host. Individual tumour cells from peritoneal ascites suspensions (pre-invasion stage) varied up to threefold in their ability to bind CIH and a similar degree of intra-tumour heterogeneity was observed in different experimental animals. Pretreatment of the cells with neuraminidase confirmed that binding was at least partly due to surface sialic acid. Invasive cells isolated from mesenteric tumour nodules were also heterogeneous with regard to the availability of surface anionic sites, as were tumour cells adhering to the surface of the mesentery; however, in both these populations CIH binding was significantly greater on average than for free ascites tumour cells. The results suggest that surface anionic sites are determinants of the invasiveness of malignant cells in vivo, and that both the number and topography of these sites may be important in modulating tumour cell behaviour.

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