Biomedical subjects

T Hatakeyama

Publications and source records attributed to T Hatakeyama.

At least 55 records · Page 3Linked to original sources

PubMed1998

Brainstem auditory evoked potentials during brainstem ischemia and reperfusion in gerbils.

To evaluate the reversibility of neural function in the brainstem following ischemia, we investigated the effect of transient brainstem ischemia on the brainstem auditory evoked potential in gerbils. Brainstem ischemia was produced by bilateral extracranial occlusion of vertebral arteries. Local cerebral blood flow was measured by quantitative autoradiography after 5 min of ischemia and was reduced to less than 3 ml/100 g per min in the pons and lower midbrain, indicating severe and reproducible brainstem ischemia. During brainstem ischemia, brainstem auditory evoked potential waveforms disappeared completely. After a brief ischemic insult (5 min), all four brainstem auditory evoked potential components recovered to normal. After longer ischemic insults (10-30 min), brainstem auditory evoked potential components never recovered to normal. Microtubule-associated protein 2 immunoreactivity revealed differential vulnerability of the acoustic relay nuclei in the brainstem. Neurons in the lateral lemniscus were most vulnerable, followed in order by neurons in the trapezoid body, the superior olive and the cochlear nucleus. We also demonstrated a close relationship between the reversibility of ischemia-induced changes on brainstem auditory evoked potential and ischemic lesions of these relay nuclei. These data may be useful for evaluating the therapeutic window of thrombolytic therapy during acute vertebrobasilar occlusion.

Animals↗

PubMed1998

Interaction of alpha-helical peptides with phospholipid membrane: effects of chain length and hydrophobicity of peptides.

To investigate the interaction of amphiphilic alpha-helical peptides with phospholipid membranes, we synthesized Ac-(Leu-Ala-Arg-Leu)3-NHCH3 (4[3]) and three derivatives, in which the chain length and the size of the hydrophobic region of the peptides were different from each other. These peptides formed an alpha-helical structure in the presence of vesicles. In the membrane-perturbation measurement, only 43 showed a strong membrane-perturbation activity below phase-transition temperature (25 degrees C), but above phase-transition temperature (50 degrees C), most peptides showed similar strong activities. On the other hand, in membrane-fusion measurement the long peptides, e.g., Ac-(Leu-Ala-Arg-Leu)3-(Leu-Arg-Ala-Leu)3-NHCH3, had strong activities at low peptide concentrations at 25 degrees C. The present study indicated a parallel relationship did not always exist between membrane fusion and perturbation caused by peptides.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed1998

Chemical modification of the hemolytic lectin CEL-III by succinic anhydride: involvement of amino groups in the oligomerization process.

CEL-III is a Ca(2+)-dependent lectin from a marine invertebrate, Cucumaria echinata, which shows strong hemolytic activity toward human and rabbit erythrocytes. After binding to carbohydrate receptors, CEL-III oligomerizes in the erythrocyte membrane to form ion-permeable pores, leading to the colloid osmotic rupture of the cells. Since hemolysis was greatly increased in the alkaline pH, especially above pH 9, involvement of amino groups of CEL-III in its hemolytic activity was evaluated using chemical modification by succinic anhydride. After modification of 7 amino groups per protein molecule, the hemolytic activity of CEL-III was reduced to 23% of the native protein, but hemagglutinating and carbohydrate-binding activities were only slightly affected even after modification of 14 amino groups. A circular dichroism spectrum of modified CEL-III showed almost no change in the secondary structure from that of the native protein, indicating that the decrease of hemolytic activity was not caused by partial unfolding of the protein. Immunoblotting analysis of the erythrocyte membrane treated with modified CEL-III showed a decrease in the formation of CEL-III oligomer in the membrane in parallel with the decrease in hemolytic activity. These results suggest that amino groups of CEL-III are involved in its oligomerization in the cell membrane, and their modification leads to inactivation of the protein without much influence on the carbohydrate-binding activity.

Amino Acids↗

PubMed1998

Studies on the carbohydrate binding sites of the hemolytic lectin CEL-III isolated from the marine invertebrate Cucumaria echinata.

The binding of carbohydrates to the hemolytic lectin CEL-III isolated from the marine invertebrate Cucumaria echinata was studied. Equilibrium dialysis data suggest that CEL-III has two carbohydrate-binding sites with equal affinity. The binding of specific carbohydrates to CEL-III induces a decrease in the fluorescence intensity at 339 nm and the shift of the fluorescence emission maximum to a wavelength shorter by 3 nm, owing to the change in the environment of tryptophan. By analyzing the change in the fluorescence intensity at 339 nm as a function of the concentration of carbohydrates, the association constants for binding of individual carbohydrates to CEL-III were calculated. The results indicate that GalNAc, lactulose, and lactose are bound by CEL-III with fairly high affinity among the carbohydrates tested. The pH-dependence profile of the association constant of lactose suggests that CEL-III binds carbohydrates with highest affinity around pH 5.0. Modification of CEL-III with N-bromosuccinimide produces an oxidized derivative, in which four tryptophan residues/mol were oxidized and had no hemolytic activity. However, two out of these four tryptophans escaped from the modification in the presence of specific saccharides and the resulting derivative retained fairly high hemolytic activity.

Animals↗

PubMed1998

Usefulness of neuroendoscopy in treating supracollicular arachnoid cysts--case report.

A 12-year-old girl presented with a supracollicular arachnoid cyst manifesting as a compressive headache. Neurological examination on admission revealed no deficit except bilateral papilledema. Stereotactic cyst puncture failed to perforate the cyst wall. The wall was then punctured using microforceps under neuroendoscopic guidance, followed by cystoperitoneal shunting. Her headache disappeared immediately after surgery. Neuroendoscopy is useful in treating a deep-seated arachnoid cyst.

Arachnoid Cysts↗

PubMed1998

[The intrarenal distribution of prostaglandin E2 and thromboxane A2 in the rat with unilateral ureteral obstruction or unilateral nephrectomy].

BACKGROUND: We evaluated the intrarenal distribution of prostaglandin E2 (PGE2) and thromboxane B2 (TxB2) on the rats that underwent unilateral ureteral obstruction (UUO), unilateral nephrectomy (UNX) or sham operation. METHODS: Male Sprague-Dawley rats were divided into three groups; left ureteral obstruction (UUO), left nephrectomy (UNX) and sham-operation (Control). They were sacrificed at 1, 3, 6, 12, 24 hours and Day 2, Day 3, Day 5, Day 7 and Day 9 after surgery. Intrarenal distribution of eicosanoids were immunohistochemically detected on both kidneys of UUO rats, and on right kidneys of UNX and Control rats. RESULTS: PGE2: In the obstructed kidneys, immunostained PGE2 increased in medullary interstitium at one hour to 6 hours, and in glomeruli and cortical interstitium at 6 hours. An increase of immunostained PGE2 was observed again in cortical interstitium at Day 3 to 5, and in medullary interstitium at Day 2 to 5. In the intact opposite kidneys, expression of immunostained PGE2 increased in glomeruli at Day 5 to 7, and in medullary interstitium at Day 3 to 5. In UNX, immunostained PGE2 increased in the medullary interstitium of the remnant kidneys at 3 hours and Day 3 to 7. On the other hand, an increase of immunostained PGE2 observed in glomeruli and cortical interstitium of these kidneys at Day 5 to 7. TxB2: In the obstructed kidneys, immunostained TxB2 increased in glomeruli and cortical interstitium at 6 hours, and in medullary interstitium at 3 to 12 hours. Predominant expression of TxB2 was observed in medullary interstitium at 3 hours compared to PGE2. We also observed an increase of immunostained TxB2 in cortical interstitium at Day 3 to 5, and in medullary interstitium at Day 2 to 5. In the intact opposite kidneys, immunostained TxB2 increased in medullary interstitium at 3 hours and Day 3. In the remnant kidneys of UNX, an increase of immunostained TxB2 was demonstrated in glomeruli at 6 hours and Day 7, and in medullary interstitium at 3 to 6 hours and Day 3 to 7. CONCLUSION: In the obstructed kidneys, imbalance between PGE2 and TxA2 may contribute to the progression of renal injuries. The fact that expression patterns of these eicosanoids in the opposite kidneys of UUO different from that of the remnant kidneys of UNK, even though both were similarly associated with functional loss of contralateral kidneys, suggested that the opposite kidneys of UUO were affected by any additional factors different from that responsible for the remnant kidneys of UNK.

Animals↗

PubMed1997

Helicobacter pylori vacuolating cytotoxin binds to the 140-kDa protein in human gastric cancer cell lines, AZ-521 and AGS.

To investigatie a potential mechanism of how Helicobacter pylori establishes infection, we purified a lot of vacuolating toxin (VacA) from supernatant of H. pylori ATCC49503 (tox+ strain 60190). We used an antibody which was prepared by immunizing rabbits with a synthetic peptide consisting of 16 amino acids reflecting a portion (Glu69-Arg83) of amino acid sequence of Vac A. VacA caused vacuoles in human gastric cancer cell lines AZ-521 AGS, and monkey kidney cell line COS-7, but not human promyeloblastic cell line HL-60. By immunoprecipitation analysis using anti VacA antibody, a biotinylated cell surface protein of 140kDa (p140) was precipitated only when the lysates of VacA-susceptible cells were incubated with VacA but not with inactivated VacA, indicating the association of p140 with VacA.

Animals↗

PubMed1997

Small-angle X-ray scattering study on CEL-III, a hemolytic lectin from Holothuroidea Cucumaria echinata, and its oligomer induced by the binding of specific carbohydrate.

Hemolytic lectin CEL-III from a marine invertebrate Cucumaria echinata forms an oligomer upon binding of specific carbohydrate such as lactose at high pH values and in the presence of high concentrations of salt. In this study, using small-angle X-ray scattering, we characterized CEL-III and its oligomer induced by the binding of lactose. The molecular mass of the oligomer was determined as 1019 kDa from its forward scattering value, compared with 47,490 Da for the monomer. This oligomer size is much larger than that estimated using SDS-polyacrylamide gel electrophoresis (SDS-PAGE, 270 kDa). The monomer has a 24.6 A radius of gyration and can be approximated by a rod which has a 20 A radius and a height of 75 A, while the oligomer has a 101.4 A radius of gyration. Together with the comparison of the radii of gyration and the forward scattering of the cross-section of the monomer and oligomer, it is suggested that in aqueous solution the oligomer comprises three or four molecules of a smaller unit which was observed by SDS-PAGE (270 kDa), held by a relatively weak interaction. The scattering profile also suggests that the oligomer has a hole in its central axis which might be associated with the formation of ion-permeable pores in the erythrocyte membrane by CEL-III during the hemolytic process.

Animals↗

PubMed1997

The enhancing effect of anionic alpha-helical peptide on cationic peptide-mediating transfection systems.

A peptide consisting of 12 amino acids including 3 glutamic acids (LAEL-LAEL-LAEL; 4(3)E) underwent pH-dependent conformational change from random coil to alpha-helix when the pH was decreased from 7.4 to 5.0 in the presence of egg PC. This alpha-helical 4(3)E had higher membrane-perturbation activity at acidic conditions compared with neutral conditions. When 4(3)E was incorporated with plasmid DNA-cationic peptide complex that utilizes an endocytosis pathway for uptake into cultured cells, high transfection efficiency was observed, indicating that 4(3)E can enhance the transfection activity of cationic peptide. It is likely that 4(3)E in the multi-complex of the plasmid DNA and the cationic peptide effectively disrupts the endosomal membrane and increases the population of the complex which could transfer to cytosol. The small lysosome-disruptive peptide is very probably useful as the enhancer molecule for the gene transfer techniques mediated by the endocytosis pathway.

Amino Acid Sequence↗

PubMed1997

Endovascular treatment for acute thrombotic occlusion of the middle cerebral artery: local intra-arterial thrombolysis combined with percutaneous transluminal angioplasty.

We report our experience in treating 15 patients with acute thrombotic occlusion of the M1 or M2 segment of the middle cerebral artery who underwent intra-arterial thrombolytic therapy alone or in combination with percutaneous transluminal angioplasty (PTA). The results were compared with those of 30 patients with acute embolic occlusion of the same artery. Intra-arterial thrombolysis was performed in 10 patients and thrombolysis combined with PTA in 5 in whom symptoms reappeared due to restenosis or reocclusion, or in whom recanalisation was not successfully accomplished by thrombolysis alone. In the patients with embolism recanalisation was observed in 28 (93 %) and there was no patient with reocclusion. In the patients with thrombosis recanalisation immediately after thrombolysis alone was observed in 9 of 15 (60%). Restenosis, with reappearance of symptoms, occurred in 2 of these (22 %). In the patients who also underwent PTA, angiography after 1 month did not demonstrate any restenosis or reocclusion. Thrombolysis combined with PTA for acute thrombotic stroke may provide an effective procedure for restoring patency and preventing reocclusion of the occluded artery.

Aged↗

PubMed1997

Carbohydrate-binding properties of the hemolytic lectin CEL-III from the holothuroidea Cucumaria echinata as analyzed using carbohydrate-coated microplate.

The carbohydrate-binding properties of the hemolytic lectin CEL-III from the Holothuroidea Cucumaria echinata were studied using the microplate assay system which we have recently developed [Hatakeyama et al. (1996) Anal. Biochem. 237, 188-192]. When the binding of CEL-III to lactose covalently immobilized on a microplate was examined using colloidal gold solution, the binding was detected with as little as 1 microgram/ml protein. Affinity of several carbohydrates to CEL-III was assessed by means of an inhibition experiment using the lactose-coated plate and it was found that N-acetylgalactosamine has the highest affinity for CEL-III, followed by lactose and lactulose. Examination of the binding of CEL-III to the lactose-coated plate at various pH values and temperatures revealed that the affinity is higher in the acidic pH region and at lower temperatures. From the Ca(2+)-dependence profile for the binding of CEL-III to the lactose-coated plate, the apparent dissociation constant for Ca2+ was estimated to be 2.3 mM. These results suggested that the carbohydrate-binding properties of CEL-III are closely related to its hemolytic activity, although an additional interaction between the protein and the lipid bilayer, which is enhanced in the alkaline pH region, also seems to be necessary for its hemolytic action.

Animals↗

PubMed1997

Temperature- and pH-dependent cytotoxic effect of the hemolytic lectin CEL-III from the marine invertebrate Cucumaria echinata on various cell lines.

We investigated the cytotoxicity of CEL-III, one of four Ca2+-dependent galactose/N-acetylgalactosamine (GalNAc)-binding lectins from the marine invertebrate Cucumaria echinata. Among six cell lines tested, MDCK cells showed the highest susceptibility to CEL-III cytotoxicity and its LD50 was estimated to be 53 ng/ml, while no significant cytotoxicity of CEL-III was observed in CHO cells up to 10,000 ng/ml. In the presence of 0.1 M lactose, the cytotoxicity of CEL-III was strongly inhibited. The binding studies using FITC-labeled CEL-III revealed that the amount of CEL-III bound to MDCK cells was about 2-fold greater than that in the case of CHO cells. The cytotoxicity of CEL-III increased with decreasing temperature. The surviving fractions of Vero cells exposed to CEL-III at 4 degrees C were immediately decreased, and more than 90% of exposed cells were killed within 20 min, whereas at 37 degrees C much longer exposure period (more than 10 h) was required to kill 50% of the cells. CEL-III induced the release of carboxyfluorescein (CF) from CF-loaded MDCK cells and this activity was markedly increased at alkaline pH (pH 10) and at lower temperature (4 degrees C). Even in CHO cells, considerable CF release was induced by CEL-III at 4 degrees C and at pH 10 but not at pH 7.5 at both temperatures. In agreement with these results, CHO cells exposed to CEL-III at 4 degrees C and at pH 10 were killed in a dose-dependent manner. These results suggest that CEL-III exhibits cytotoxicity through damaging the plasma membrane by pore-formation in a temperature- and pH-dependent manner. Different susceptibility of each cell line to CEL-III cytotoxicity may be due to differences in the processes leading to pore-formation after binding to cell-surface carbohydrates.

Animals↗

PubMed1997

Adults and children with high imagery show more pronounced perceptual priming effect.

36 children in Grade 5 and 59 university students, all native speakers of Japanese, studied three types of priming stimuli in a mixed list: words written in hiragana (Japanese syllabary used in writing), words written in kanji (Chinese characters also used in writing), and pictures. They were then given a task involving completion of hiragana-word fragments: the task involved studied and nonstudied items. For both children and university students, words in hiragana produced the largest priming effects, that is, the words that had appeared in hiragana in the preceding study phase were generated more often in the test phase of word completion than the other two types of priming stimuli. This confirms that the perceptual priming effect depends much on data-driven processing. For both age groups, words in kanji produced nearly half the priming effects seen for hiragana-words. On the other hand, pictures had no priming effect for children but they had a similar effect to kanji-words for students. The discrepancy between kanji-words and pictures for children suggests that the former force the subject to read the words, which, possibly, activates the hiragana-words, while the latter do not necessarily force labelling the pictures. Among three kinds of imagery tests, the Verbalizer-Visualizer Questionnaire predicted priming scores for children and the Questionnaire upon Mental Imagery did so for students, but the Test of Visual Imagery Control did not predict the scores for either age group. This shows that children reporting habitual use of imagery and adults reporting vivid imagery have more pronounced perceptual priming effects. We conclude that the imagery ability based on self-judgments reflects real characteristics of the perceptual representation system of Tulving and Schacter (1990).

Adult↗

PubMed1997

[Autologous peripheral blood stem cell transplantation for CHOP-resistant mediastinal diffuse large B-cell lymphoma].

We report two cases of CHOP-resistant mediastinal diffuse large B-cell lymphoma (MDLCL) treated by autologous peripheral blood stem cell transplantation (auto-PBSCT). The patients were 17 and 28 year old female, and they had anterior mediastinal bulky masses at presentation. The clinical stage was IV in both patients. They were initially treated with 6 courses of bi-weekly CHOP therapy following by irradiation of the mediastinal tumors. During the last few cycles of CHOP therapy, their disease had become progressive. They underwent auto-PBSCT after high-dose chemotherapy consisting of busulfan, cyclophosphamide and etoposide, but failed to achieve remission. They died within one year after the auto-PBSCT because of the disease progression. Our experience suggests that this conditioning regimen is not effective for CHOP-resistant MDLCL, and it is necessary to develop new conditioning regimens.

Adolescent↗

PubMed1996

Quantitative determination of cardiac glycosides in Digitalis lanata leaves by reversed-phase thin-layer chromatography.

An analytical method for the determination of cardiac glycosides in Digitalis lanata leaves by reversed-phase thin-layer chromatography (RP-TLC) was developed. The procedure consisted of extraction of dry leaf powder with 50% methanol and clean-up by Sep-Pak cartridges prior to RP-TLC analysis. RP-TLC was performed on an octadecylsilyl bonded silica gel plate, using a developing solvent of acetonitrile-methanol-0.5 M NaCl (1:1:1) for primary glycosides and acetonitrile-methanol-0.5 M NaCl (12:7:9) for secondary glycosides. The plate was scanned with a reflectance densitometer at 225 nm. The quantitation was carried out by the internal standard method. The present method is reliable and relatively simple for the determination of cardiac glycosides in Digitalis lanata leaves.

Cardiac Glycosides↗

PubMed1996

Oligomerization of the hemolytic lectin CEL-III from the marine invertebrate Cucumaria echinata induced by the binding of carbohydrate ligands.

The hemolytic lectin CEL-III is a Ca2+-dependent, galactose/GalNAc-specific lectin purified from the marine invertebrate Cucumaria echinata (Holothuroidea). We found that this lectin forms ion-permeable pores in erythrocyte and artificial lipid membranes that have specific carbohydrate ligands on the surface. The hemolytic activity of CEL-III exhibited characteristic pH dependence; activity increased remarkably with pH in the alkaline region, especially above pH 9. When rabbit erythrocyte membrane was examined by immunoblotting using anti-CEL-III antiserum after treatment with CEL-III, the irreversible binding of the CEL-III oligomer increased with pH, indicating that the increase in hemolytic activity at higher pH is associated closely with the amount of oligomer irreversibly bound to the membrane. Surface hydrophobicity of CEL-III, as measured by the fluorescent probe 8-anilino-1-naphthalenesulfonate, increased markedly with the binding of specific ligands such as lactose, lactulose, and N-acetyllactosamine at pH 9-10 in the presence of 1 M NaCl. The enhancement of surface hydrophobicity induced by the binding of carbohydrates was also accompanied by the formation of a CEL-III oligomer, which was found to be the same size on sodium dodecyl sulfate-polyacrylamide gel electrophoresis as the oligomer that formed in CEL-III-treated erythrocyte membranes. Far-UV circular dichroism spectra of CEL-III and the oligomer revealed a definite difference in secondary structure. These data suggest that the binding of CEL-III to specific carbohydrate ligands on the erythrocyte surface induces a conformational change in the protein, leading to the exposure of a hydrophobic region which triggers oligomerization and the irreversible binding of the protein to the membrane.

Animals↗

PubMed1996

An assay for lectin activity using microtiter plate with chemically immobilized carbohydrates.

A simple microtiter plate assay for lectins or carbohydrate-binding proteins was developed. The method utilizes carbohydrates immobilized in the wells of the microtiter plate containing primary amino groups on their surface. After incubation of the lectins, bound proteins are measured by the protein assay using the colloidal gold solution. When the binding of Ricinus communis agglutinin, concanavalin A, and wheat germ agglutinin was measured using the microtiter plate wells coated with lactose, mannose, or N-acetylglucosamine, binding of the lectins according to their known specificity was observed. Inhibition experiments with various carbohydrates also demonstrated that the specificity of lectins for different carbohydrates could be determined quantitatively. Since there is no need for modification of the lectins, such as biotinylation or conjugation with marker enzymes, the carbohydrate-binding ability of intact proteins can be easily determined by this method. When gel filtration fractions from the extract of the marine invertebrate Cucumaria echinata were subjected to this assay, different carbohydrate-binding activities were observed with different elution profiles, suggesting that this assay could also be widely applicable for the simultaneous detection of lectins from various sources.

Animals↗

PubMed1996

A strategy for selective anti-cancer drug concentration increase in rat glioma tissue with Ca(2+)-channel blocker co-administration: calcium kinetics in intra-glioma arteriolar smooth muscle cells.

A rat glioma model was employed to estimate the Ca2+ kinetics in the tumor arteriolar smooth muscle cells. Electron microcytochemistry revealed that the density of intracellular Ca2+ deposits in the intra-tumor arteriolar smooth muscle cells was significantly greater, with slightly higher membrane Ca(2+)-adenosine triphosphatase (ATPase) activity, compared to the contralateral cerebral arterioles. Furthermore, the administration of tyrphostin, a tyrosine kinase inhibitor, specifically increased only the intra-tumor blood flow. These findings suggest that the condition of the intra-tumor arteriole alters the susceptibility to contraction by the accelerated Ca2+ influx into the cytoplasm mediated through the tyrosine kinase pathway. After the administration of diltiazem, which also has a blocking effect on the Ca(2+)-channel mediated through this pathway, the local intra-tumor blood flow showed an increase of 39% with a marked decrease of intracellular Ca2+ concentration of the arteriolar smooth muscle cells in the tumor, while the blood flow in the basal ganglia increased by only 8%. The intra-tumor concentration of Nimustine-HCl (ACNU) with co-administration of diltiazem was significantly increased compared to that without the co-administration. Co-administration of diltiazem may be a valuable strategy in chemotherapy for glioma in affording the selective increase of intra-tumor concentration of the anti-cancer drug.

Animals↗