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At least 1,189 records · Page 66Linked to original sources
PubMed1985

Aspects of splenic hypofunction in old age.

The weight of the human spleen decreases in old age. The percentage of erythrocytes with pits observed by interference contrast microscopy was significantly higher in 39 elderly subjects (median 4.2%, range 1.2%-14.6%) than in 34 younger controls (median 1.4%, range 0.6%-6.2%). Further evidence of splenic hypofunction was obtained from 7 elderly people with increased pit counts: the removal from circulation and accumulation into the spleen of heat-damaged autologous erythrocytes was abnormally slow. However, no difference in the spleen area of the same subjects measured by scintillation scanning was observed in comparison with normals. Thus, the weight reduction of spleen in old age is accompanied by a decrease in splenic function.

Adolescent↗
PubMed1986

Membrane potentials and intracellular Cl- activity of toad skin epithelium in relation to activation and deactivation of the transepithelial Cl- conductance.

The potential dependence of unidirectional 36Cl fluxes through toad skin revealed activation of a conductive pathway in the physiological region of transepithelial potentials. Activation of the conductance was dependent on the presence of Cl or Br in the external bathing solution, but was independent of whether the external bath was NaCl-Ringer's, NaCl-Ringer's with amiloride, KCl-Ringer's or choline Cl-Ringer's. To partition the routes of the conductive Cl- ion flow, we measured in the isolated epithelium with double-barrelled microelectrodes apical membrane potential. Va, and intracellular Cl- activity, acCl, of the principal cells identified by differential interference contrast microscopy. Under short-circuit conditions, Isc = 27.0 +/- 2.0 microA/cm2, with NaCl-Ringer's bathing both surfaces, Va was -67.9 +/- 3.8 mV (mean +/- SE, n = 24, six preparations) and acCl was 18.0 +/- 0.9 mM in skins from animals adapted to distilled water. Both Va and acCl were found to be positively correlated with Isc (r = 0.66 and r = 0.70, respectively). In eight epithelia from animals adapted to dry milieu/tap water Va and acCl were measured with KCl Ringer's on the outside during activation and deactivation of the transepithelial Cl- conductance (GCl) by voltage clamping the transepithelial potential (V) at 40 mV (mucosa positive) and -100 mV. At V = 40 mV; i.e. when GCl was deactivated, Va was -70.1 +/- 5.0 mV (n = 15, eight preparations) and acCl was 40.0 +/- 3.8 mM. The fractional apical membrane resistance (fRa) was 0.69 +/- 0.03. Clamping to V = -100 mV led to an instantaneous change of Va to 31.3 +/- 5.6 mV (cell interior positive with respect to the mucosal bath), whereas neither acCl nor fRa changed significantly within a 2 to 5-min period during which GCl increased by 1.19 +/- 0.10 mS/cm2. When V was stepped back to 40 mV, Va instantaneously shifted to -67.8 +/- 3.9 mV while acCl and fRa remained constant during deactivation of GCl. Similar results were obtained in epithelia impaled from the serosal side. In 12 skins from animals adapted to either tap water or distilled water the density of mitochondria-rich (DMRC) cells was estimated and correlated with the Cl current (ICl through the fully activated (V = -100 mV) Cl conductance). A highly significant correlation ws revealed (r = -0.96) with a slope of -2.6 nA/m.r. (mitochondria-rich cell and an I-axis intercept not significantly different from zero.(ABSTRACT TRUNCATED AT 400 WORDS)

Amiloride↗
PubMed1975

Studies on the brush border membrane of the mouse duodenum. I. Membrane isolation and analysis of protein components.

Brush border membranes have been isolated from villus epithelial cells of the adult Swiss mouse duodenum. Preparations of these membranes are not contaminated by other organelles as judged from electron-micrographs of sectioned pellets of brush borders. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of proteins from brush borders solubilized in Tris-sodium dodecyl sulfate buffer reveals a reproducible Coomassie Brilliant Blue pattern of 17 bands. By comparing the brush border protein band positions with those of standard proteins run concurrently on sodium dodecyl sulfate-polyacrylamide gel slabs it is estimated that the 17 brush border proteins and subunits have molecular weights ranging from over 250,000 to around 16,000. Periodate-fuchsin sulfite staining shows that the five more slowly migrating, high molecular weight proteins are glycoproteins. The two proteins of smallest molecular size react positively with Oil Red O but have very small amounts of lipophilic amino acid residues, which indicates that the lipid extractable from the gels in these areas is a contaminant and is not bound to the proteins.

Animals↗
PubMed1985

Method for testing adherence of Haemophilus influenzae to human buccal epithelial cells.

A method for testing adherence of Haemophilus influenzae strains to buccal mucosal cells is described. Bacteria grown in broth for 4 h were mixed with buccal mucosal cells. After elimination of unattached bacteria by repeated cycles of centrifugation and resuspension in PBS, the number of attached bacteria was counted microscopically. Optimal results were obtained with an early log-phase bacterial culture at a concentration of 10(9) bacteria/ml mixed with 2 X 10(4) cells/ml and incubated at 37 degrees C for 60 min. This assay showed an at least ten times higher rate of adherence for Haemophilus influenzae than previous studies. Nontypeable strains attached in higher numbers than strains with the type b capsule. Adherence was related to the frequency of nontypeable strains rather than to the site of isolation or type of infection. Thus all the isolates from middle ear fluid were nontypeable, and all but one adhered. The results suggest a difference in virulence mechanisms between type b and nontypeable Haemophilus influenzae strains.

Adhesiveness↗
PubMed2002

Capsule-substrate contact deformation: determination of adhesion energy.

A study is reported of a cellular entity (liquid-filled microcapsule) adhered on a flat glass substrate in response to changes in osmotic pressure and temperature. High-resolution reflection interference contrast microscopy (HR-RICM) and phase-contrast microscopy were developed for probing the adhesion contact area, capsule-substrate separation profile and adhesion energy of the adhering microcapsule. The new technique increased the detection limit of the measured capsule wall-substrate separation in the cohesive zone from 1 to 4.5 microm and improved the spatial resolution of the heterogeneous contact zones. A theoretical model was applied to correlate quantitatively the adhesion energy to the area of the contact zone. The work demonstrated the possibility of ascertaining the quantitative interfacial adhesion energy of a liquid-filled microcapsule using the present technique and represents the first step in extending this novel approach to study more complicated systems, such as cell-substrate interactions, in the future.

Animals↗
PubMed1976

A longitudinal study of parotid secretory kinematics by cinemicrography and computer analysis.

The kinematic responses of parotid gland acini to isoproterenol were studied in vitro by time-lapse cinemicrography. It was shown that once secretion begins, granule depletion accelerates to a maximum rate in 8-10 min, continues at a steady rate for 5-8 min and then slows, depletion becoming essentially complete 25-30 min after stimulation. The continuum of morphologic change manifested by the acinar lumen during secretion was quantified by computer analysis of sequential cine-frames representing a time resolution of 0.1 min. Interpretation of seventh-order and eleventh-order polynominal non-linear least squares trend curves shows that the lumen enlarges from an initial cross-sectional area of approximately 96 mum2 at 14.9 min. The shape of these curves suggests that after an initial partial enlargement the lumen shrinks (or remains steady) before climbing to the maximum. The basic pattern resembles distortion curves characteristic of two-phase viscoelastic compliance systems. It is also shown that luminal plasma membranes undergo a very active ebb and flow during the egestive phase of secretion.

Animals↗
PubMed1992

An in vitro model giving access to adhesion plaques.

A new approach was investigated to study the interaction between integrins and actin via intracytoplasmic proteins. Because intracellular processes are hampered by the limiting plasma membrane, we developed an in vitro model with cells perforated by a bacterial toxin, streptolysin O. The specific conditions for the use of permeabilized cells to study the intramolecular associations occurring at adhesion plaques are described. The two cell types used, HUVEC and CHO, showed that the choice of the perforation method is of great importance. After perforation of cells in a monolayer, 75 +/- 10% of the cells remained adherent to a fibronectin substrate; after perforation of cells in suspension, only 25 +/- 10% of the cells readhered. Specific conditions were required however to maintain these adhesive properties up to 4 h: the presence of 1 mM Mg++ in the medium was crucial, and it was necessary to layer the cells on a specific coat rather than a substitute such as gelatin. Immunofluorescence investigations of actin, talin and vinculin, and Normarsky differential interference contrast microscopy showed retention of focal adhesion plaques in perforated cells. Moreover, in perforated cells antibodies directed against actin led to actin disorganization, showing that our model of perforated cells in a monolayer can give new insight to adhesion study.

Actins↗
PubMed1995

Expression levels of B-50/GAP-43 in PC12 cells are decisive for the complexity of their neurites and growth cones.

To study the role of the protein B-50/GAP-43 in NGF-induced neurite outgrowth, a number of stable PC12 subclones with either very low or considerably enhanced expression levels of the protein were selected. Cell bodies of subclones with suppressed B-50 expression (-B2, -B5, or -B12) possessed a relative small spherical shape and, on NGF-treatment for 7 d, developed processes that were virtually devoid of branches and that mostly bore short or blunt-ended growth cones. Cells of subclones with overexpression of B-50 (+B3, +B4, or +B11), on NGF treatment, acquired a flattened, spiky appearance with highly branched neurites possessing extended and complex growth cones. Confocal microscopy with immunofluorescence for B-50 and F-actin revealed that in neurites and growth cones of the B-50-deficient subclone -B2, no detectable B-50 and reduced amounts of filamentous F-actin were present, whereas in overexpressing +B3 cells, cell membranes, neurites, and complex growth cones were intensively stained for B-50 and exhibited numerous spikes, in which B-50 was strikingly colocalized with F-actin. These data suggest that, under normal conditions of neuritogenesis, the expression level of B-50 in PC12 cells is decisive for the complexity of neurites and growth cones.

Animals↗
PubMed1983

On spatial periodicity in the formation of cell adhesions to a substrate.

Interference reflection microscopy reveals a fairly regular array of marginal focal contacts in early states of cell spreading. A theoretical explanation of the onset of patchiness is given, based on the positively cooperative binding to the substrate of laterally diffusing receptor molecules.

Animals↗
PubMed2001

Imaging transmitter release. I. Peeking at the steps preceding membrane fusion.

Over the recent year we have witnessed considerable advances in the study of neurotransmitter release. This progress has been severalfold as different techniques have allowed us to characterise many steps along the process of exocytosis, membrane fusion, formation of the fusion pore, and have given insight in the kinetics of release and membrane re-uptake. Patch clamping provided quantitative measurements of the capacitance changes as the membrane of the secretory vesicle is added to the surface of the cell during secretion, and the change in the opposite direction when membrane is retrieved back into the cell during exocytosis. Carbon-fibre microelectrodes have measured electrochemically the release of oxidisable transmitters into the extracellular space. Differential interference contrast microscopy has given us spatially resolved images of the cell surface during exocytosis; real-time images that are suggestive of bubbles breaking the surface of a boiling pot of water. The interest in novel techniques stems from the fact that existing approaches can provide only indirect evidence on the steps preceding membrane fusion. The vesicular dynamics just beneath the plasma membrane are out of the reach of capacitance measurements or amperometric detection. What we have needed is a tool that would allow us to look just below the cell surface. This much-needed tool appears to be evanescent-wave microscopy. This review describes how laser microscopy can be used to study exocytosis at single-vesicle resolution. A companion paper deals with the practical aspects of evanescent-wave imaging.

Electrochemistry↗
PubMed1996

Hydraulic properties of MDCK cell epithelium.

The water permeability of the apical and basolateral cell membranes and the compliance of the lateral intercellular spaces (LIS) of MDCK monolayers were measured on confluent cultures grown on permeable supports. Cell membrane water permeabilities were determined, using quantitative differential interference light microscopy, from the rate of cell volume decrease after exposure to a hyperosmotic bathing solution. Both membranes exhibited osmotic water permeabilities (POSM) of approximately 10 microm/sec, comparable to that of unmodified lipid bilayers. The compliance of the cell membranes forming the lateral intercellular space (LIS) between cells was determined from the pressure-volume relation. Confocal microscopy of fluorescent labeling of the basolateral cell membranes was used to delineate the LIS geometry as transepithelial hydrostatic pressure was varied. The LIS were poorly deformable as a function of transepithelial hydrostatic pressure until a pressure of >/=8 cm H2O (basolateral > apical) was reached where catastrophic failure of intercellular connections occurred. The compliance of the LIS was calculated from the geometry changes at pressures <8 cm H2O and ranged from 0.05-0.11 cm H2O-1, comparable to that previously predicted in mathematical models of the rat proximal tubule.

Animals↗
PubMed2002

Decorated surfaces by biofunctionalized gold beads: application to cell adhesion studies.

We describe a simple but versatile method to decorate solid surfaces randomly with colloidal gold particles to which ligands of cell receptors can be coupled to generate local attraction sites for the control of cell adhesion. A self-assembled monolayer of (3-mercaptopropyl)trimethoxysilane was deposited on glass slides. Gold beads were anchored to the functionalized surface through the sulfur group. We characterized the gold bead distribution on the functionalized surface with reflection interference contrast microscopy. The gold beads were functionalized with a disulfide-coupled cyclic pentapeptide containing an arginine-glycine-aspartic acid (RGD) tripeptide sequence which is selectively recognized by integrin receptors alpha(V)beta(3) of endothelial cells. A blocking layer of bovine serum albumin was adsorbed onto the surface to prevent non-specific binding of the cells. We demonstrate that the RGD-functionalized colloidal gold beads act as local attraction centers, mediating rapid cell anchoring on a substrate impeding cell adhesion in the absence of attraction centers. Surprisingly, microinterferometry shows that after a time delay of about 1 h, the regions of the cell surface between the gold beads form close contacts with the substrate, which is attributed to strong van der Waals attraction after escape of repeller molecules from the contact surface.

Biophysics↗
PubMed2003

A new labyrinthulid isolate, which solely produces n-6 docosapentaenoic acid.

A labyrinthulid strain, L59, was isolated from a leaf floating on seawater collected at the coastal area of Hokkaido Prefecture, Japan. Strain L59 contained only n-6 docosapentaenoic acid ( n-6 DPA) among all the long-chain polyunsaturated fatty acids. The proportion of n-6 DPA in the total fatty acids was 48.1% and the total fatty acids content in the cell dry weight was 26.6%. Many oil bodies were observed in the cell, mostly in the vicinity of cell membranes. The strain had spindle-shaped cell bodies and all cells were surrounded by ectoplasmic net elements. It was also clearly classified in the labyrinthulid group by phylogenetic analysis. In the optimum culture condition, using soybean oil and peptone as carbon and nitrogen sources, 0.53 g of n-6 DPA/l was produced at 20 degrees C in 7 days.

DNA, Fungal↗