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Biomedical subjects

N Uyesaka

Publications and source records attributed to N Uyesaka.

34 records · Page 2Linked to original sources
PubMed1988

Flow cytometric studies of the binding of monoclonal antibodies OKT3, OKT4 and OKT8.

The binding of monoclonal antibodies (OKT3, OKT4 and OKT8) to human T cells was investigated by flow cytometry. A flow cytometer was calibrated with standard fluorescence microspheres, which permitted quantitation of the number of bound antibody molecules. Considerable care was taken to perform the flow cytometric assay at a constant temperature and the effect of temperature on the binding reaction was examined. The binding of OKT3, OKT4 and OKT8 exhibited saturation kinetics. The maximum binding varied with temperature. Kinetic analysis according to the Hill equation revealed that the value of the Hill coefficient for OKT3 changed from 1.8 to 1.0 when the temperature was raised from 12 degrees C to 36 degrees C, whereas the corresponding values for OKT4 and OKT8 did not vary with temperature. Thermodynamic functions obtained from the Van't Hoff plot showed that the binding of OKT3 was exothermic whereas the binding of OKT4 and OKT8 were endothermic.

Animals↗
PubMed1988

Effect of water-soluble iodinated contrast media on pressure-flow relationship of red cell suspension.

The effect of radiological contrast media on blood flow through a vascular network was investigated, taking physical and physiological conditions such as osmolality into account. The perfusion of the bullfrog's hind limbs was performed, with a slight modification of the vertical tube method. The effect of contrast media on red cell deformability was studied by perfusion with erythrocyte suspensions in glutaraldehyde-fixed hind limbs. The echinocytic shape change induced by metrizamide and hypertonic iothalamate solutions caused a marked increase in resistance to flow. When the perfusion with erythrocyte suspension was performed using intact hind limbs, the pressure-flow relationship was influenced by contrast media effects on both red cell deformability and the vascular bed. Ioxaglate had less rheologic effect on the pressure-flow relationship than metrizamide or iothalamate. It could be concluded that contrast media should be isotonic, of low viscosity and chemotoxicity, and that ioxaglate was preferable to metrizamide and iothalamate at equal iodine content.

Animals↗
PubMed1988

Pressure-flow relationship of erythrocyte suspension in perfusion of nuclepore membrane and red cell deformability.

The hemodynamic characteristics of Nuclepore (NP) membrane filtration were evaluated from a pressure (P)-flow rate (Q) relationship of erythrocyte suspension obtained by the vertical-tube method (Nichol et al., 1951) with a slight modification. It became evident that the vertical-tube method was a more quantitative and simple method than the conventional steady flow perfusion method using a variable speed pump. The P-Q relationship of erythrocyte suspensions in the perfusion of NP membrane consisted of a smooth curve convex to the P-axis at low P tending to a straight line at high P. The extrapolated linear segments of P-Q curves of erythrocyte suspension did not converge to the same point on the negative Q-axis, that is, the value of the negative intercept increased as flow decreased. This observation indicated clearly an obstruction or plugging of the pores by erythrocytes. It was made clear that flow rate measured with NP membrane filtration was influenced not only by intrinsic red cell deformability but also by several different factors, namely, the different distribution of pore size of NP membrane, microdust contaminated inevitably in red cell suspension, total volume of cell suspension through NP membrane, stagnation or retention of erythrocyte suspension and the hematocrit value of suspension. In conclusion, the present study shows that the precise definition of deformability depends on the method or rheological conditions, such as hematocrit value of suspension, used to measure it, even if confined within NP membrane filtration.

Blood Flow Velocity↗
PubMed1985

A new standard fluorescence microsphere for quantitative flow cytometry.

A standard fluorescence microsphere has been prepared for quantitative flow cytometry. A uniformly sized (6.53 +/- 0.34 micron in diameter) and perfectly spherical plastic microsphere, Micropearl-FT, was covalently conjugated with fluorescein isothiocyanate (FITC). The characteristic of this microsphere to lyse completely in strong alkaline solutions made it possible to determine the amount of bound FITC easily and reproducibly. The amount of bound FITC depended on the concentration of FITC and the incubation time at pH 9.5 and 37 degrees C. In flow cytometry, the fluorescence intensity of the FITC-conjugated microsphere (FCMS) showed linear dependence on the amount of bound FITC. The fluorescence intensity of FCMS was stable for at least 3 months even in aqueous solutions. Using FCMS, an Epics V was calibrated and the minimum measurable fluorescence intensity was examined. From analysis using this system, the number of FITC-conjugated OKT3 molecules bound to human T lymphocytes was shown to average 4.4 X 10(4) per cell at saturation.

Adult↗
PubMed1976

Calcium-binding of synaptosomes isolated from rat brain cortex. IV. Effects of ruthenium red on the Co-operative nature of calcium-binding.

Ruthenium red combines with isolated synaptosomes, resulting in strong inhibition of their Ca2+-binding. In isotonic saline media, however, the dye-induced inhibition of Ca2+-binding is significantly greater than that expected for the amount of bound dye and Hill's exponent of the Ca2+-binding decreases to 1 with an increase in the amount of the dye bound. On the other hand in isotonic mannitol-sucrose solution, inhibition of synaptosmal Ca2+-binding brought about by the dye is proportional to the amount of dye bound. Based on these results, the effects of the dye on the co-operative nature of synaptosomal Ca2+-binding is discussed.

Animals↗
PubMed1976

Lanthanum and some other cation-induced changes in fluidity of synaptosomal membrane studied with nitroxide stearate spin labels.

Using nitroxide fatty acid spin labels, the effects of some cations such as La3+, Cd2+ and Hg2+ on synaptosomal membranes were studied by observing changes in their ESR spectra. The labels were incorporated almost instantaneously into synaptosomes isolated from rat brain cortex. ESR spectra of the spin-labeled synaptosomes were significantly braodened immediately upon adding La3+, Ce3+, Cd2+ or Hg2+ but hardly affected by Ca2+, Sr2+ and Ba2+. The magnitude of the change in the separation of the outer two peaks in ESR spectra (2T') depends on the number (n) of methylene units between the polar head group and the spin-label (nitroxide) group; that is, it increases with decreasing n. Among these ions, the effect of La3+ was the greatest and appeared to be in parallel with the amount of La3+ bound with the synaptosomes. On the other hand, K+, Rb+ or Li+ causes hardly any significant changes.

Animals↗
PubMed1975

Calcium-binding of synaptosomes isolated from rat brain cortex, III. Binding with some divalent heavy metal ions and calcium-binding sites.

Free ion concentration of some divalent heavy metal ions such as Mn2+, Co2+, Ni2+, Cd2+ and Zn2+ in the synaptosomal suspension was measured to determine binding with synaptosomes isolated from rat brain cortex. A dual wavelength spectrophotometer was utilized to monitor the absorbance changes of murexide raised by stepwise addition of these ions (as chloride salts). Such titration experiments of the synaptosomal suspension revealed that a part of the added divalent cation such as Mn2+, Co2+ or Ni2+ was almost instantaneously bound to synaptosomes in isotonic NaCl media. Our previous study (Kamino, Uyesaka & Inouye, J. Membrane Biol. 17:13, 1974) demonstrated that raised external K+ resulted in a specific noncompetitive inhibition of synaptosomal Ca-binding. Just like the Ca-binding, Mn-, Co- or Ni-binding was almost completely depressed by high external K+ or ruthenium red when the free concentration of the cations was 10 mum or less, while at higher concentrations the binding was not affected. The present results indicate that tested divalent cations bind with both "Ca-binding sites" and "non-Ca-binding sites" of synaptosomal membrane, the nature of the binding sites of both being quite different: the former is sensitive to high external K+ and to ruthenium red but the latter is not.

Animals↗
PubMed1975

Calcium-binding of Synaptosomes isolated from rat brain cortex. II. Inhibitory effects of magnesium ions and some other cations.

As in our previous report (Kamino, Uyesaka & Inouye, J. Membrane Biol. 17:13 1974), the absorbance changes of murexide caused by Ca2+ and followed up by a dual wavelength spectrophotometer were applied to measure synaptosomal Ca2+-binding in the presence of cations such as Rb+, Mn2+ or La3+. All the cations tested showed a significant inhibition of synaptosomal Ca2+-binding except Li+. The inhibitory effects could be divided into the following three categories: (1) noncompetive, co-operative K+-type, which includes alkali metal ions. The potency of inhibition is K+ greater than Rb+ greater than Cs+ greater than Li+, Na+ =0; (2) competitive Mn2+ -type which includes many divalent cations. The inhibitory potency was found to be in the following order: Mn2+ greater than Sr2+ greater than Cd2+, Ba2+ greater than Mg2+; (3) nonspecific, noncompetitive La3+ -type; among the cations tested, La3+ and Ce3+ were found to markedly reduce the Ca-binding capacity of synaptosomal particles, resulting in a noncompetitive inhibition, at least in the range of Ca2+ concentration used.

Animals↗
PubMedDate not supplied

Effects of superoxide anions on red cell deformability and membrane proteins.

The effect of superoxide anions (O2-) on red blood cells (RBC) deformability and membrane proteins was investigated using hypoxanthine-xanthine oxidase system. Exposure of RBC to O2- caused a marked decrease in RBC deformability with a concomitant increase in cell volume and shape changes. The RBC exposed to O2- also displayed pronounced degradation of membrane proteins such as band 3 protein and spectrin; new bands of low molecular weight products appeared as the original membrane proteins tended to diminish, without the appearance of high molecular weight products. Since the membrane proteins are involved in processes regulating membrane properties such as permeability and viscoelasticity, the decreased deformability induced by O2- may be attributable to changes in membrane proteins. Interestingly, resealed ghosts exposed to O2- did not show any significant change in membrane proteins, which suggests the existence of further generation of O2- and subsequent production of other active oxygen species mediated by O2(-)-initiated autoxidation of hemoglobin in intact RBC. Furthermore, electrophoretic analysis suggested that active oxygens increased the endogenous proteolytic susceptibility of RBC. In conclusion, a close linkage was suggested between RBC deformability and the membrane proteins.

Anion Exchange Protein 1, Erythrocyte↗
PubMedDate not supplied

Impaired deformability of Heinz body-forming red cells.

Although a decrease in the deformability of red blood cells (RBCs) has been suspected in Heinz body-forming hemolytic anemia, it remains uncertain whether the formation of Heinz bodies themselves impairs RBC deformability or not. To elucidate this question, we used RBCs treated with phenylhydrazine and RBCs from patients with unstable hemoglobin (Hb) disease (Hb Yokohama) to investigate the effect of Heinz body formation on deformability in terms of RBC filterability through nickel mesh and viscosity of the RBC suspension. The phenylhydrazine-treated RBCs exhibited a marked decrease in deformability in a dose-dependent manner. The Heinz body-forming RBCs from the patients also showed a marked decrease in deformability. Thus we confirmed that Heinz body formation impairs RBC deformability. Further, both phenylhydrazine-treated RBCs and RBCs from the patient showed a degradation of spectrin without any cross-linking of membrane proteins, thereby suggesting that the impaired deformability is associated with the oxidative degradation of the cytoskeletal framework. In summary, this study supports the conclusion that RBC deformability is impaired by the presence of Heinz bodies as well as the related oxidative damage involved in their formation.

Anemia, Hemolytic↗