Biomedical subjects
Publications and source records attributed to M Ota.
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The addition of vanadyl ribonucleoside complex (VRC), a potent inhibitor of RNase, to the transformed 4.5S androgen receptor from rat submandibular gland caused an increase in the sedimentation coefficient to 7.0S. Moreover, VRC decreased the DNA-cellulose binding of the transformed receptor; 50% inhibition of the DNA-cellulose binding was achieved at 1.8 mM VRC. On the other hand, agents related to VRC and oxoanions of transient metals, such as ribonucleoside, vanadate, molybdate, tungstate and arsenate, exerted no effect on the DNA-cellulose binding ability of the receptor. These findings suggest that VRC binds to the transformed androgen receptor at the DNA-binding site and that both oxovanadium ion and ribonucleoside are indispensable for the binding of VRC to the transformed androgen receptor.
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The transformed glucocorticoid receptor (GR) from rat liver precipitated at 30% saturation of ammonium sulfate and sedimented at 4.3 S on glycerol gradient centrifugation, whereas the nontransformed GR precipitated at higher concentrations of ammonium sulfate (40-50% saturation) and sedimented at 8.6 S on a gradient. Sodium dodecyl sulfate polyacrylamide gel electrophoresis showed that heat shock protein 90 (hsp 90) precipitated at 40-50% saturation of ammonium sulfate. Moreover, hsp 90 and the nontransformed GR were eluted from DEAE high performance ion-exchange chromatography at similar salt concentrations (0.22-0.23 M NaCl), whereas the transformed GR was eluted at 0.1 M NaCl. Therefore, hsp 90 seems to be responsible for the surface charge characteristics of the nontransformed GR.
The use of a novel amide surfactant, N-methyl oleoyl taurate (Igepon T-77), has been examined for the separation of amines on reversed-phase chromatographic material. This reagent was found to partition onto the C18 material in a partially irreversible and concentration independent manner. When the stationary phase is saturated with this surfactant, the loaded column performs as a strong cation exchanger. Novel separations are possible as a result of secondary hydrogen-bonding effects which modify classical retention order for primary, secondary and tertiary amines. Sensitive and selective applications of these separations are demonstrated for catecholamine determinations in blood plasma, cerebrospinal fluid, urine and brain tissue. Additional sensitivity is obtained for epinephrine by taking advantage of the pH-dependent intramolecular cyclization and on-column concentration of large injection volumes.
Goblet cell mucin in 39 human colons was studied by methods specific for various sugar residues, including staining with three lectins, Dolichos biflorus agglutinin (DBA, specific for blood group A antigen), Griffonia simplicifolia agglutinin-I (GSA-I, B) and peanut agglutinin (PNA, T antigen), and immunostaining for A, B, H and T. Isoantigens A, B or H were found only in the right colon. GSA-I reactive goblet cells occurred in the right colon of both blood group A and B patients and possibly contained isoantigens. However DBA reactive cells were found in all cases. Prior neuraminidase digestion imparted anti-A, GSA-I and DBA reactivities to the cells lining the lower crypts in all cases. This pretreatment also imparted PNA and anti-T reactivities to goblet cells, only the latter reactivity being eliminated by galactose oxidase. Goblet cell mucin in transitional mucosa revealed decreased A and B, and increased H antigens. Enhanced galactose oxidase-Schiff (GOS) and anti-T reactivities were also noted. The present results revealed that some lectin reactions of goblet cells might be related to blood group antigens but others were not, and that different techniques for demonstrating reputedly the same sugar residues produced different results, indicating a need for proper evaluation of their specificity.
The fetal erythrocyte membranes were partially solubilized with Triton X-100 at the low concentration (0.5%). The localizations of Rh1(D), 2(C), 3(E), 4(c), 5(e) and 25(LW) were investigated. Using hemagglutination inhibition assay, Rh1(D) antigen activity was observed in the Triton-treated membrane (Triton shell) containing mainly band 1, 2 (spectrin), band 5 (actin), band 4.1 and a part of band 3, while Rh2(C), 3(E), 4(c), 5(e) and 25(LW) antigens were detected in the supernatant containing band 3, 6, 2.2, 2.3 and 4.2. It is suggested that: Rh1(D) antigen would associate with cytoskeleton matrix of fetal erythrocyte membranes; Rh1(D) and Rh25(LW) antigens might be integral membrane proteins, while Rh2(C), 3(E), 4(c) and 5(e) antigens would be surface membrane proteins which are easily released from membranes by EDTA, mercaptoethanol and alkaline treatments.
The nontransformed glucocorticoid receptor (GR) and an 88-kDa protein in rat liver cytosol were selectively adsorbed on protamine- and arginine-Sepharose. The 88-kDa protein was purified from rat liver cytosol to homogeneity by precipitation with protamine sulfate, followed by DEAE-ion exchange chromatography, gel chromatography, and DEAE ion-exchange high-performance liquid chromatography. The 88-kDa protein appeared to be present as a dimer at both low and high salt concentrations. The physicochemical properties and the amino acid composition of the 88-kDa protein were almost the same as those of heat-shock protein 90 of HeLa cells and yeast. Preincubation of the GR with the polyclonal antibody raised against the 88-kDa protein increased the sedimentation coefficient of the nontransformed GR, but did not change that of the transformed GR. These results indicate that heat-shock protein 90 is associated with rat liver nontransformed GR and is responsible for the interaction of GR with protamine.
The purpose of this study was to examine the effect of glucocorticoid on human neoplastic salivary duct epithelial cell line (HSG). Dexamethasone was found to inhibit cell growth and to increase cell size and the ratio of protein content to DNA content in a cell. The inhibition of cell growth was dose-dependent; in comparison to the control (33.8 +/- 3.1 h), the population doubling time was 1.57-fold longer in 10(-5) M dexamethasone (P less than 0.01, N-K test). [3H] thymidine incorporation was inhibited in 45.5% of the control at 10(-5) M. Plating efficiency was 20.5 +/- 3.0% in 10(-5) M and 47.0 +/- 4.4% in the absence of dexamethasone. Cell diameters increased 1.29 fold in 10(-5) M dexamethasone in comparison to the control size (16.0 +/- 2.1 micron). The ratio of total protein content of DNA content increased 1.46 fold in 10(-5) M dexamethasone-treated cells on the seventh day of cultivation. Scatchard plot analysis using [6, 7-3H]-triamcinolone revealed that the HSG cells had apparent cytosolic glucocorticoid receptors with an equilibrium dissociation constant (Kd value) of 6.48 nM, whose number of binding sites (NBS) was 57.8 fmol/mg protein.
This study is the first to report on the use of x-ray photoelectron spectroscopy (XPS or ESCA) for studying the surface films (less than 10 nm thick) of aged amalgams. The concentrations and electron binding energies of the elements on the surfaces of four different amalgams aged for 20 min, one day, seven days, and 30 days were determined quantitatively. For comparison, the bulk compositions of the amalgams aged for seven days were also determined after removal of approximately 5 nm of material from the surface by argon-ion-sputtering. The XPS data revealed that the surface films of aged zinc-containing amalgams were not a simple oxide but were primarily composed of a (hydrated) tin and zinc oxy-hydroxide, whereas, in the zinc-free amalgams, the surface films were primarily a tin oxide. The concentration of mercury in this thin surface film after aging was depleted. This suggests that tin and/or zinc preferentially diffused to the surface and combined with oxygen, forming a surface film and diluting the mercury concentration in the surface. Another probable explanation for the depleted mercury is that a minimal amount of mercury in the surface film evaporated during the aging.
The activities of p-tosyl-L-arginine methyl ester (TAMEase) and cytosolic and nuclear androgen receptors in the submandibular glands of male and female mice were determined at various developmental stages. In males, the activity of TAMEase was detectable at four weeks after birth, and thereafter it increased rapidly. Cytosolic androgen receptor increased gradually with age, whereas nuclear androgen receptor, which was minimal one week after birth, increased remarkably four weeks after birth. In females, the minimal activity of TAMEase was detectable at five weeks after birth and increased very slowly with age. Cytosolic receptor increased with age, but nuclear receptor level was unchanged. These findings suggest that the appearance of TAMEase was in accordance with an elevation of nuclear androgen receptor in mice submandibular glands.
Two familial cases of dilated cardiomyopathy were evaluated by HL-A typing. In the case of the first family, the mode of inheritance is likely to be an autosomal dominant trait. Only the affected individuals carried the identical HL-A haplotype (A2, Bw54, Cw1, DR4, DQw3), while the unaffected members do not share this pattern. In the second family case, the disease is probably inherited by autosomal recessive traits. All of the family members examined shared the identical HL-A haplotype (A24, Bw52, DR2, DQw1), but only the affected individuals were homozygous for this haplotype.
Female mice were used to examine the process of depletion and replenishment of cytosolic androgen receptors in submandibular glands, and to investigate the effects of cycloheximide and actinomycin D on these processes. The dose-dependence of receptor depletion and replenishment in the cytosolic fraction, and that of receptor accumulation in the nuclear fraction were investigated. Almost 100% depletion was revealed 1 h after the injection of testosterone propionate at a dose of 500 or 50 micrograms testosterone/100 g body weight. With a 5 micrograms dose, depletion of cytosolic receptors was not complete and replenishment proceeded rapidly compared with that which occurred with the 50 or 500 micrograms dose. The nuclear receptor level increased 1 h after injection of testosterone, and the raised level was gradually reduced to the pretreatment level with all doses. However, the time required for this return to pretreatment level was dependent on the dose of testosterone. The change in the levels of cytosolic and nuclear androgen receptors following injection of testosterone was parallel to the level of circulating androgen. To determine the requirements for transcriptional and translational events in the replenishment process, cycloheximide and actinomycin D were given in vivo. The process of replenishment of cytosolic receptors was inhibited by the injection of cycloheximide. However, actinomycin D exerted no inhibitory effect on receptor replenishment. Neither cycloheximide nor actinomycin D had any effect on the nuclear receptor level until 6 h after the injection of testosterone. Cycloheximide or actinomycin D alone had no effect on the cytosolic or nuclear receptor level. These results suggest that receptor replenishment involves protein synthesis.
A paternity test is presented in which a father and his two children possessed an extremely rare amorphic gene R-29 (r,---). One of the children was determined to be illegitimate at the first trial as her Rh phenotype was R2R2(ccDEE) and the father's phenotype was R1R1(CCDee). At the Court of Appeal, however, the rare Rh gene r(---) was shown to be inherited from the father to the appellant child through extended tests including her brother whose phenotype was also R2R2(ccDEE). She was acknowledged to be legitimate.