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Biomedical subjects

S Ohki

Publications and source records attributed to S Ohki.

At least 127 records · Page 7Linked to original sources
PubMed1983

Isolation and characterization of large (0.5 - 1.0 micron) cytoskeleton-free vesicles from human and rabbit erythrocytes.

Large (0.5 - 1.0 micron) cytoskeleton-free vesicles were obtained, by 'budding', from fresh human and rabbit erythrocytes incubated at 45 degrees C and titrated with EDTA and CaCl2. This process occurs without hemolysis. The isolated vesicles maintain their cytoplasmic integrity and normal membrane orientation, and are resistant to hemolysis over the pH range 5.0 - 11.0 and temperature range 4-50 degrees C. The only membrane proteins detected in vesicles from human erythrocytes were band 3 region polypeptides and bands PAS-1, PAS-2 and PAS-3. Vesicles obtained from rabbit erythrocytes were similarly simple. Because of their size and stability these vesicles are amenable to both kinetic and quantitative analysis using the same experimental techniques employed in studies of synthetic lipid membranes. The results obtained in this study indicate that these vesicles are essentially markedly simplified biological cells, and thus may be useful as a biologically relevant model membrane system for examining the molecular interactions which occur within, across and between cell membranes.

Animals↗
PubMed1982

A mechanism of divalent ion-induced phosphatidylserine membrane fusion.

A mechanism for the divalent cation-induced membrane fusion of phosphatidylserine membranes is proposed. Fusion was followed by the Tb/DPA (dipicolinic acid) assay, monitoring the fluorescent intensity for mixing of the internal aqueous contents of unilamellar lipid vesicles, and the threshold concentrations required for various divalent cations to induce membrane fusion were determined from the fluorescence spectrum of the lipid vesicle suspension with respect to various concentrations of divalent ions. Also, the surface tension of monolayers made of the same lipids as used in the fusion experiments was measured with respect to the variation of divalent cation concentrations. The surface tension increase in the monolayer, induced by changing divalent ion concentrations from zero to a concentration which corresponded to its threshold concentration to induce vesicle membrane fusion, was the same (approx. 8 dyn/cm) for all divalent ions used. From these experimental data and theory concerning ion binding to the membrane, it is deduced that the main cause of divalent cation-induced membrane fusion of phosphatidylserine membranes is the degree of increased hydrophobicity (surface tension increase) of the membrane surface, which results from the binding of cations to acidic phospholipid membrane surfaces. Some discussion on the molecular mechanism of phospholipid membrane fusion is given.

Calcium↗
PubMed1982

Effects of proteins on phospholipid vesicle aggregation and lipid vesicle-monolayer interactions.

The effects of proteins on divalent cation-induced phospholipid vesicle aggregation and phospholipid vesicle-monolayer membrane interactions (fusion) were examined. Glycophorin (from human erythrocytes) suppressed the membrane interactions more than N-2 protein (from human brain myelin) when these proteins were incorporated into acidic phospholipid vesicle membranes. The threshold concentrations of divalent cations which induced vesicle aggregation were increased by protein incorporation, and the rate of vesicle aggregation was reduced. A similar inhibitory effect by the proteins, incorporated into lipid vesicle membranes, was observed for Ca2+-induced lipid vesicle-monolayer interactions. However, when these proteins were incorporated only in the acidic phospholipid monolayers, the interaction (fusion) of the lipid vesicle-monolayer membranes, induced by divalent cations, was not appreciably altered by the presence of the proteins. In contrast to these two proteins, the presence of synexin in the solution did enhance the Ca2+-induced aggregation of phosphatidylserine vesicles, but did not seem to affect the degree of Ca2+-induced fusion between phosphatidylserine/phosphatidylcholine (1:1) and phosphatidylserine vesicles and monolayer membranes.

Animals↗
PubMed1982

[TAC-278. A new 5-FU derivative, by the oral route for its efficacy and safety in 36 patients with advanced cancers].

Tumor regression was observed in only one patient. Twelve (33%) of the 36 patients reported adverse effects: gastrointestinal symptoms in 8, CNS symptoms in 3, and others in 1. The antitumor efficacy of TAC-278 was insufficient while incidence of adverse effects was similar to other 5-FU derivatives in man and the clinical usefulness of the drug was hardly found. TAC-278 has been developed with an aim to give higher plasma concentrations of 5-FU in man. Even at a dose causing adverse effects, however, the clinical efficacy of TAC-278 was unsatisfactory. The study is a meaning tal eince the results suggest that there might be a more important factor than blood concentration of 5-FU to enhance an antitumor effect of 5-FU derivatives.

Adult↗
PubMed1981

Surface potential and surface charge density of the cerebral-cortex synaptic vesicle and stability of vesicle suspension.

Using a microelectrophoresis instrument employing the Lazer-Zee system, the electrophoretic mobility of synaptic vesicles isolated from Guinea-pig brain cortex was measured under conditions. The mobility was found to depend on both pH and ionic concentration of the solution. The surface of the synaptic vesicle was shown to be negatively charged under physiological conditions. The isoelectric point was observed at pH 4.0 in 0.01 M NaCl solution. Effects of divalent cations were examined and reversal of surface charge was observed in 0.1 M CaCl2 solution. Interaction of vesicles was also considered on the basis of the DLVO theory of colloid stability by using calculated values of surface charge density and surface potential of the synaptic vesicle.

Animals↗
PubMed1981

Surface potential of phosphatidylserine monolayers. II. Divalent and monovalent ion binding.

Ion binding constants for phosphatidylserine membranes have been derived from the variation of the surface potential of phosphatidylserine monolayers with divalent cation concentrations in the presence of various monovalent salts in the aqueous subphase. The observed surface potential data for the monolayers, analyzed by use of the Gouy-Chapman diffuse potential theory, together with a simple binding reaction formula, yield, for Ca2+, Mg2+, Na+ and (Me)4N+ binding constant values of 30 M-1, 10 M-1, 0.6 M-1 and 0.05 M-1, respectively. The effect of pH on surface potential of phosphatidylserine monolayers was found to be dependent upon ionic species other than H+ in the subphase solution. The distinction between apparent and intrinsic dissociation constants of H+ for biomolecules was made in terms of ion binding due to other ions at the same site as for H+ in biomolecules.

Animals↗
PubMed1981

Permeability of axon membranes to local anesthetics.

The permeability of the neutral form of tertiary amine local anesthetics across squid axon membranes was studied by utilizing three different experimental methods: (1) narcotic action of axon excitability was measured by monitoring the time derivative of action potential and the results were analyzed in terms of a diffusion reaction equation of local anesthetics to obtain their permeabilities; (2) the influx of local anesthetic into the axon was measured by use of the radioisotope tracer technique; and (3) the desorption rates of the neutral form of local anesthetics from lipid monolayers were measured and the desorption rate was correlated with permeability. The relative permeabilities obtained for procaine, lidocaine and tetracaine by the above three methods were comparable. The order of relative permeabilities was procaine greater than lidocaine greater than tetracaine, and had an inverse correlation with the partition coefficients of anesthetics at oil/water phases. Some discussion concerning the concept of permeability is made when the partition coefficient of a permeant molecule is high.

Action Potentials↗
PubMed1981

Correlation between membrane expansion and temperature-induced membrane fusion.

For each phospholipid membrane, there is a characteristic phase transition temperature, and for each phospholipid spherical membrane, there is a specific 'fusion' temperature. In order to examine the possible correlation between temperature-induced membrane fusion and membrane expansion, the relationship between the physical states of phospholipid membranes at both temperatures have been investigated by the use of the monolayer system. Monolayer expansion studies have indicated that the increase in area per lipid molecule, caused by increasing the temperature from the phase transition to the fusion temperature, is approximately the same for five different phospholipids used. With the same temperature increase, phospholipid monolayers containing cholesterol did not expand appreciably. This correlates qualitatively with the greater inhibition of membrane fusion seen in the spherical phospholipid membrane systems when cholesterol was incorporated in the membrane. The effect of pH on the expansion of phosphatidylserine monolayers was also studied in relation to membrane fusion phenomena. The shift in fusion temperature of the spherical phospholipid membranes due to the change of pH is explained by the shift in phase transition temperatures of lipid membranes. The expanded area per molecule in the monolayer caused by increasing the temperature from the phase transition to the fusion temperature was approximately the same irrespective of surface charge densities.

Cholesterol↗
PubMed1981

Fusion of small unilamellar liposomes with phospholipid planar bilayer membranes and large single-bilayer vesicles.

Small unilamellar phosphatidylserine/phosphatidylcholine liposomes incubated on one side of planar phosphatidylserine bilayer membranes induced fluctuations and a sharp increase in the membrane conductance when the Ca2+ concentration was increased to a threshold of 3--5 mM in 100 mM NaCl, pH 7.4. Under the same ionic conditions, these liposomes fused with large (0.2 micrometer diameter) single-bilayer phosphatidylserine vesicles, as shown by a fluorescence assay for the mixing of internal aqueous contents of the two vesicle populations. The conductance behavior of the planar membranes was interpreted to be a consequence of the structural rearrangement of phospholipids during individual fusion events and the incorporation of domains of phosphatidylcholine into the Ca2+-complexed phosphatidylserine membrane. The small vesicles did not aggregate or fuse with one another at these Ca2+ concentrations, but fused preferentially with the phosphatidylserine membrane, analogous to simple exocytosis in biological membranes. Phosphatidylserine vesicles containing gramicidin A as a probe interacted with the planar membranes upon raising the Ca2+ concentration from 0.9 to 1.2 mM, as detected by an abrupt increase in the membrane conductance. In parallel experiments, these vesicles were shown to fuse with the large phosphatidylserine liposomes at the same Ca2+ concentration.

Animals↗
PubMed1981

Membrane potential, surface potential, and ionic permeabilities.

To clarify contributions to cellular transmembrane potential concentration potentials of phospholipid bilayer membranes and surface potentials of phospholipid monolayers were measured with respect to salt concentrations and various surface charge densities. For highly charged membranes of symmetrical surface charge density, it was found that the observed transmembrnae potentials are due mostly to the difference between the surface potentials on each side of the membrane. For uncharged membranes, it was found that the transmembrane potential is due to the ion diffusion potential across the membrane. Also discussed are ion permeabilities through the membrane in relation to transmembrane potential.

Cell Physiological Phenomena↗
PubMed1980

Binding of Streptomyces pepsin inhibitor (acetyl-pepstatin) with chymosin (Rennin).

Chymosin (Rennin) was effectively purified using an AH-Sepharose 4B column. Binding of Streptomyces pepsin inhibitor (acetul-pepstatin) with chymosin was studied spectroscopically. The binding caused ultraviolet difference and CD spectral changes suggesting microenvironmental changes around tryptophan and/or tyrosine residue(s) in chymosin. The fluorescence intensity of a hydrophobic probe, 2-p-toluidinylnaphthalene-6-sulfonate, increased in the presence of chymosin and was further amplified when Streptomyces pepsin inhibitor was added to the chymosin-2-p-toluidinylnaphthalene-6-sulfonate solution. The binding and dissociation-rate constants between chymosin and the inhibitor were determined using 2-p-toluidinylhnaphthalene-6-sulfonate as a probe. The binding constant was determined from the binding and dissociation-rate constants, to be 3.1 . 10(7) M-1 at 25 degrees C, pH 5.5.

Chymosin↗
PubMed1980

Hydrophobic interaction between the monomer of mitochondrial malate dehydrogenase and phospholipid membranes.

Porcine mitochondrial malate dehydrogenase (EC 1.1.1.37) dissociates into subunits on dilution. The enzyme monomer caused large increases in the surface pressure of monolayers of 1:1 phosphatidylserine/phosphatidylcholine at air/water and oil/water interfaces. The monomer increased the permeability of phospholipid vesicles to 22Na+. Both effects were significantly greater than the corresponding effects of ribonuclease A, cytochrome c and the dimeric form of malate dehydrogenase. Changes in the circular-dichroism spectra of the enzyme indicated that conformational changes may be associated with dimer formation or when monomer interacts with lysophosphatidyl-choline. Similar interactions to those described may occur in situ when mitochondrial malate dehydrogenase is transported to the mitochondrial matrix from its site of synthesis on cytosolic ribosomes.

Animals↗