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Biomedical subjects

K Takeda

Publications and source records attributed to K Takeda.

At least 649 records · Page 36Linked to original sources

Cytotoxic NK1.1 Ag+ alpha beta T cells with intermediate TCR induced in the liver of mice by IL-12.

Systemic administration of IL-12 greatly reduced the hepatic metastases of i.v.-injected liver metastatic EL4 tumor cells in C57BL/6 +/+ and nu/nu mice. Cytotoxic assay in vitro revealed that administration of IL-12 greatly enhanced cytotoxicity of hepatic mononuclear cells (MNC) against various NK- sensitive and -resistant tumor targets, including EL4 cells, whereas only slight or moderate augmentation of the cytotoxicity was observed in splenocytes in normal and nude mice. After IL-12 administration, hepatic MNC increased in number and showed vigorous proliferation in vitro. Hepatic MNC of control C57BL/6 +/+ mice contain alpha beta T cells with intermediate TCR (TCRint) as well as alpha beta T cells with bright TCR, whereas hepatic MNC of nu/nu mice have only TCRint cells. These TCRint cells are found to be NK1.1 Ag+ (NK1+ TCRint). Systemic administration of IL-12 into normal and nude mice markedly augments the NK1 expression of NK1+ TCRint cells (NK1high TCRint), which is comparable to or brighter than that of NK cells in the liver, whereas alpha beta T cells with bright TCR or gamma delta T cells in the liver are NK1-. Depletion of either NK1.1+ or CD3+ cells, but not CD8+ cells, of hepatic MNC from IL-12-treated normal mice by respective Abs and C in vitro abrogate their cytotoxicity. These results revealed that TCRint cells are potent cytotoxic effector cells and suggest that NK1high TCRint cells are the main antimetastatic population in the liver, and that TCRint cells are functionally different from regular T cells with bright TCR.

Animals↗

Differential control of the metal-mediated activation of the human heme oxygenase-1 and metallothionein IIA genes.

Heme oxygenase-1, an essential enzyme in heme catabolism, and metallothionein IIA, a small metal-binding protein with clusters of cysteins, are remarkably induced in HeLa cells following the treatment with cadmium or zinc. Both proteins are considered to be involved in the defense system against metal toxicity. Here we showed by transient expression assays that the cadmium-responsive element (CdRE) of the human heme oxygenase-1 gene is not responsive to zinc, whereas a metal-regulatory element (MRE) of the human metallothionein IIA gene is able to respond to either cadmium or zinc. The CdRE is recognized by a certain nuclear protein(s) which is however unable to bind to an MRE of the metallothionein IIA gene. These results suggest that the metal-selective activation of each gene promoter is mediated by a separate mechanism.

Base Sequence↗

Value of a dynamic MR scan in predicting vascular ingrowth from free vascularized scapular transplant used for treatment of avascular femoral head necrosis.

We have treated three patients with avascular osteonecrosis using vascularized scapular bone graft. To predict blood perfusion in both the diseased femoral head and the transferred bone, all the hips were followed up using dynamic magnetic resonance (MR) scans, performed 1 and 7 months after surgery. In the present cases, it was shown that conventional enhanced MR imaging sometimes depicts increased intensity in bone marrow without blood perfusion due to the leakage of gadolinium-DTPA (Gd-DTPA) from the capillaries surrounding the avascular tissue. It was found that Gd-DTPA remaining in the dead bone marrow resulted in a false-positive image. By contrast, the dynamic MR scan evaluated only those images taken before the leakage. This is one of the advantages of the dynamic study, which reflects actual blood flow in the bone. The fast rise in the time-intensity curve following bolus injection of Gd-DTPA indicates that there is fast blood perfusion in the bone. The dynamic MR scan has demonstrated that there is little blood perfusion in the diseased bone 1 month after the operation and that vascular ingrowth from the transferred bone flap proceeds gradually between 1 and 7 months after surgery. These findings indicate that the dynamic MR scan is very useful in demonstrating vascular ingrowth after surgery in avascular necrosis of the femoral head and can be a reliable monitoring technique for anastomotic patency of the vascularized bone flap.

Adolescent↗

Inhibition of nicotinic acetylcholine receptor channels in bovine adrenal chromaffin cells by Y3-type neuropeptide Y receptors via the adenylate cyclase/protein kinase A system.

The effect of neuropeptide Y [NPY(1-36)] and related peptides on the voltage-dependent currents and the nicotinic acetylcholine receptor (nAChR) currents (IACh) of bovine adrenal chromaffin cells was investigated using the whole-cell patch clamp technique. Catecholamine release from single chromaffin cells was measured by means of fast cyclic voltammetry. The potency order of these peptides in inhibiting IACh evoked by nicotine was NPY(1-36), NPY (16-36) > peptide YY(PYY) > [Leu31, Pro34]NPY. NPY(16-36) produced a similar degree of inhibition, irrespective of whether nicotine or an equipotent concentration of acetylcholine was used to evoke IACh. NPY(16-36) failed to alter voltage-dependent inward or outward currents. Intracellular cAMP, and extracellular dibutyryl-cAMP, produced a slowly developing increase in IACh. Intracellular cAMP, extracellular 8-Br-cAMP or dibutyryl-cAMP, and an inhibitor of cyclic nucleotide phosphodiesterases 3-isobutyl-1-methyl-xanthine (IBMX), decreased the inhibitory effect of NPY(16-36) on IACh. Although the intracellular application of the cAMP-dependent protein kinase A inhibitor [PKI(14-24)amide] alone did not alter IACh, it potentiated the effect of NPY(16-36) in interaction experiments. While the NPY(16-36)-induced inhibition of IACh was reversed on washout of the peptide, the slightly shorter C-terminal fragment NPY(18-36) caused a long-lasting depression of both IACh and catecholamine secretion evoked by nicotine. This depression was smaller in the presence of extracellular 8-Br-cAMP than in its absence. NPY(18-36) did not alter the secretory activity induced by a high concentration of potassium. It appears that, by activating Y3-receptors, NPY inhibits nAChR-current and the resulting secretion of catecholamines from bovine chromaffin cells. This process may involve a G protein-mediated decrease in intracellular cAMP with a subsequent decrease in the degree of phosphorylation of the nAChR-channel.

Acetylcholine↗

The expression of Na+/K(+)-ATPase beta-subunit cRNA injected into Xenopus oocytes is affected by coinjection with alpha-subunit cRNA.

The cRNA for Torpedo californica Na+/K(+)-ATPase beta-subunit (cRNA beta) was injected into Xenopus oocytes alone or with the cRNA for the Na+/K(+)-ATPase alpha-subunit (cRNA alpha). When cRNA beta was injected alone, the amount of the beta-subunit that accumulated in oocytes increased with increasing amounts of injected cRNA beta. When cRNA beta and cRNA alpha were injected simultaneously, less beta-subunit accumulated than when cRNA beta was injected alone, whereas the Na+/K(+)-ATPase activity increased markedly. The decrease in the accumulation of the beta-subunit was dose-dependent upon the cRNA alpha. The mutant beta-subunit unable to assemble with the alpha-subunit accumulated in oocytes independently of cRNA alpha, suggesting that post-translational control mechanisms may serve to reduce the accumulation of the beta-subunit.

Animals↗

Calcium-sensitivity of the plasmalemmal delayed rectifier potassium current suggests that calcium influx in pulvinar protoplasts from Mimosa pudica L. can be revealed by hyperpolarization.

Isolated protoplasts from pulvinar motor cells of Mimosa pudica were studied using conventional whole-cell patch clamp techniques. With internal solutions weakly buffered for Ca2+ (0.2 mM EGTA), a rundown of the outward delayed rectifier K+ current was induced by hyperpolarizing the holding potential, and this effect was strongly promoted by high external Ca2+ concentrations. This rundown could be reversed by coming back to less hyperpolarized holding potentials or by lowering the external [Ca2+]. Such rundown was absent when pipette internal solutions strongly buffered (10 mM EGTA) for Ca2+ were used. Ionomycin induced rundown of the K+ current with internal solutions containing 0.2 mM but not 10 mM EGTA. The hyperpolarization-associated rundown was reversibly blocked by Gd3+ and La3+.

Bepridil↗

Neuropeptide Y2-type receptor-mediated activation of large-conductance Ca(2+)-sensitive K+ channels in a human neuroblastoma cell line.

We have proposed recently that a pertussistoxin-insensitive Ca2+ influx stimulated by Y2-type receptor activation in CHP-234 human neuroblastoma cells underlies increases in intracellular free Ca2+ concentration ([Ca2+]i) induced by neuropeptide Y (NPY), which were strictly dependent on extracellular Ca2+ and independent of internal Ca2+ stores. We describe here the actions of NPY in these same cells, using the activity of Ca(2+)-activated K+ channels as an indicator of [Ca2+]i. The elementary slope conductance of these channels was 110 +/- 3 pS (with an asymmetrical K+ gradient), their activity was greatly increased by application of ionomycin, and they were reversibly blocked by 1 mM tetraethylammonium (TEA) and 100 nM charybdotoxin. Application of 100 nM NPY, in the presence but not in the absence of extracellular Ca2+, increased the channel open probability. ATP applied in the absence of external Ca2+ caused rises both in channel open probability and [Ca2+]i. Inositol trisphosphate production was stimulated by ATP but not by NPY. In outside-out patches, NPY increased channel open probability, indicating that NPY-associated Ca2+ influx does not require all the intracellular machinery present in intact cells. Channel activation by NPY was unaffected by the replacement of guanosine 5'-triphosphate (GTP) by (guanosine 5'-O-(2-thiodiphosphate) (GDP[ beta S]), a non-hydrolysable GDP analogue, in the pipette internal solution, consistent with the lack of involvement of G-proteins in the coupling of Y2-type receptors to Ca2+ influx in CHP-234 cells.

Adenosine Triphosphate↗

Role of [Ca2+]i in lethal oxidative injury in rat cultured inner medullary collecting duct cells.

Reactive oxygen metabolites have been implicated in the pathogenesis of toxic, ischaemic and immunologically mediated renal injury. An increase in the cytosolic free Ca2+ concentration ([Ca2+]i) has been proposed as a mechanism of oxidative stress-induced cell injury. We used a fluorescence spectrometer and a fluorescence probe to measure the [Ca2+]i and viability of rat primary cultured inner medullary collecting duct (IMCD) cells during oxidative stress induced by 5 mM tert-butyl hydroperoxide (TBHP). Initially, this oxidative stress evoked a small increase in [Ca2+]i which was followed by a slower sustained increase from the resting level of 170.8 +/- 38.8 nM to 1490.5 +/- 301.7 nM after 60 min, and this preceded the loss of plasma membrane integrity, measured by the propidium iodide fluorescence method. The elimination of extracellular Ca2+ from the culture medium prevented the TBHP-induced [Ca2+]i increase and improved cell viability. Restoration of extracellular Ca2+ resulted in an immediate and large increase in [Ca2+]i and extensive cell death. Verapamil, a Ca2+ channel blocker, inhibited the [Ca2+]i increase and afforded significant protection against cellular injury following exposure to TBHP-induced oxidative stress. Extracellular acidosis also prevented the increase in [Ca2+]i and cell death caused by this oxidative stress. These results are consistent with the hypothesis that oxidative stress-induced IMCD cellular injury may be the result of increased [Ca2+]i caused, in part, by activation of voltage-dependent Ca2+ channels.

Animals↗

Conformational changes of alpha-lactalbumin induced by the stepwise reduction of its disulfide bridges: the effect of the disulfide bridges on the structural stability of the protein in sodium dodecyl sulfate solution.

Four disulfide bridges of bovine alpha-lactalbumin (alpha-lact) were selectively reduced to obtain its derivatives with three, two, and zero disulfide bridges (designated as 3SS, 2SS, and 0SS alpha-lact, respectively). The original helicity was almost maintained in 3SS alpha-lact missing only the Cys6-Cys120 bridge. Upon the reduction of both Cys28-Cys111 and Cys6-Cys120 bridges, various changes occurred in the protein. In particular, the maximum fluorescence of 1-anilinonaphthalene-8-sulfonic acid was observed in this stage. Upon the reduction of all disulfide bridges, the hydrophobic box of the protein, formed by Trp60, Ile95, Tyr103, and Trp104, was disrupted and an internal helical structure was destroyed. The conformation of each derivative was examined mainly in a solution of sodium dodecyl sulfate. In the surfactant solution, the helicity increased from 33% to 37% in 3SS alpha-lact, from 26% to 31% in 2SS alpha-lact, and from 18% to 37% in 0SS alpha-lact, as against from 34% to 44% in intact alpha-lact. On the other hand, the tryptophan fluorescence of each derivative was affected in very low surfactant concentrations, suggesting that the tertiary structure considerably changed prior to the secondary structural change in the surfactant solution.

Anilino Naphthalenesulfonates↗

Heterotopic gastric mucosa of the gallbladder.

A case of heterotopic gastric mucosa in the fundus of the gallbladder is reported. A 23-year-old man, who had been healthy and asymptomatic, visited our hospital because of abnormal findings in a liver enzyme test given during a routine health screening. Ultrasonography demonstrated a highly echogenic polypoid mass in the fundus of the gallbladder. The gallbladder mass was confirmed by both computed tomography and intravenous cholangiogram. After a 10-month follow up, laparoscopic cholecystectomy was performed. Intraoperative touch smear cytology of this lesion revealed class II cells. The surgical specimen revealed a 15 x 10 x 5 mm polypoid lesion in the fundus, with no gallstones in the gallbladder. Histologically, the polypoid lesion consisted of both fundic type and pyloric type gastric glands located in the mucosa of the gallbladder. In the literature, 42 cases of heterotopic gastric mucosa of the gallbladder have been reported, only 3 of which, including this present case, were found incidentally, with no apparent symptoms.

Adult↗

Coherent potential approximation approach to electronic structure of DNA.

The electronic structure of DNA is theoretically investigated by use of the coherent potential approximation. Even when the sequence of the four kinds of bases is nonperiodic, guanine block forms the persistent highest valence band edge, and adenine block forms the persistent lowest conduction band edge state. According to the calculated joint density-of-states energy profiles, the site first attacked by the lowest excitation is adenine block. After this excitation, electrons are generated at adenine sites, and holes are generated at guanine sites. The resulting electronic structures of the valence band and conduction band suggest that the base complementarity in DNA produces the complementarity in the density-of-states divergences of the excited electrons and holes. This complementarity lowers the excitation instability in the DNA chains.

DNA↗

Word length and error types in Japanese left-sided neglect dyslexia.

Seven patients who showed left-sided neglect in reading single words were examined. Neglect errors typically involved omission of initial letters in our patients. Two patients with enough errors had a tendency that longer words were more susceptible to errors than short words. Most of the patients were less affected than the patients in the previous reports. Japanese people read strings of letters written not only from left to right but also from right to left. The seven patients continued to misread the letters on the left end when asked to read words from right to left. The two patients showed a significant deficit in reading words from left to right compared to reading from right to left.

Adult↗

A comparative study of whole-blood platelet aggregation in laboratory animals: its species differences and comparison with turbidimetric method.

We undertook a systematic comparison of whole-blood platelet aggregation concerning species difference of commonly used laboratory animals (rabbits, guinea pigs, rats and dogs), agonist difference (collagen, ADP and platelet activating factor [PAF] and technical difference against a traditional method of turbidimetry. Collagen-induced aggregation with whole-blood aggregometry was less variable among species tested and was similar to the results obtained by turbidimetry, which measures platelet aggregation in citrated plasma. In contrast, whole-blood aggregometry in responses to ADP and PAF showed marked species difference, being different from turbidimetry. Our results provide species difference among laboratory animals and suggest that the modulations of platelet aggregation by other blood elements probably differ in each species and with stimulating agonists.

Animals↗

Expression of prostaglandin E receptor subtypes in bone: expression of EP2 in bone development.

Prostaglandin (PG) E2 displays physiological and pharmacological action in various tissues including bone. It increases intracellular Ca, and stimulates or inhibits cAMP production through the PGE receptor subtypes EP1, EP2, and EP3, respectively. These receptor subtypes have been recently cloned. In the present study, we investigate the expression of these receptor subtypes in bone tissue. RT-PCR revealed that EP1, EP2, and EP3 were expressed in rat calvariae and that osteoblastic cells (MC3T3-E1) expressed EP1 and EP2. In situ hybridization analysis using cryosection of neonatal calvariae revealed that EP2 was expressed by osteoblasts and cells not in contact with bone, probably including preosteoblasts. EP2 expression was observed at an early stage in calvarial development, at 14 days prenatal. EP2 expression was also observed at day 3 in rat bone marrow cell culture in which bone-like mineralized nodules are formed at day 8. It has been established that PGE2 response accompanying cAMP production is one of the characteristics of osteoblasts. The present results indicate that this phenotype appears at an early stage of osteoblastic differentiation and bone development.

3T3 Cells↗

Distribution of immunoreactive 2',5'-oligoadenylate synthetase in mouse digestive tract.

2',5'-Oligoadenylate synthetase (2-5OAS), an enzyme induced by interferon (IFN), has also been found in various "normal" animals that had not been treated with IFN. The distribution of this enzyme in the digestive tracts of normal healthy mice was studied by western blotting and by an immunohistochemical method using a specific monoclonal antibody. On western blotting, the antibody to 42 kD 2-5OA synthetase reacted with extracts from the stomach and intestines (small and large intestine), but not with extracts from the esophagus. Immunohistochemically, the 42 kD 2-5OAS was localized on the following cells: surface mucous and parietal cells of fundic glands in the stomach; surface epithelial cells in the intestines; and some enteric nervous cells in the esophagus, stomach, and intestines. Treatment with IFN- alpha/beta did not essentially change the distribution of the enzyme in the tissues, although a small amount of the enzyme was detected in the esophagus by western blotting. Expression of the 42 kD 2-5OAS in the digestive tract may be responsible for the prevention of viral infections.

2',5'-Oligoadenylate Synthetase↗

Functional consequences of substitution of the disulfide-bonded segment, Cys127-Cys150, located in the extracellular domain of the Na,K-ATPase beta subunit: Arg148 is essential for the functional expression of Na,K-ATPase.

The Cys127-Cys150 disulfide-bonded loop (L1) of the Torpedo californica Na,K-ATPase beta 1 subunit was substituted with the corresponding loop of the rat beta 1, mouse beta 2, or pig H,K-ATPase beta subunit. All the substituted mutant beta subunits assembled with the Na,K-ATPase alpha subunit in a trypsin-resistant manner. The mutants with L1 from the Na,K-ATPase beta subunit isoforms (rat beta 1 and mouse beta 2) each formed a functional complex with the Na,K-ATPase alpha subunit. On the other hand, the complex of the alpha subunit with the mutant beta subunit that was substituted with the pig H,K-ATPase beta subunit L1 was inactive as to ATP hydrolysis. Ser131 and Phe148 located within L1 of the pig H,K-ATPase beta subunit-substituted mutant were back-mutated to Pro131 and Arg148, respectively. The Phe148 to Arg mutation restored the ability of the mutant beta subunit substituted with the H,K-ATPase beta subunit L1 to form a functional complex with the alpha subunit. These results suggested that the Cys127-Cys150 loop of the Na,K-ATPase beta 1 subunit, especially Arg148, plays a critical role in the functional expression of Na,K-ATPase.

Amino Acid Sequence↗