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Biomedical subjects

K Sakai

Publications and source records attributed to K Sakai.

At least 685 records · Page 38Linked to original sources
PubMed1994

[A problem of quantitative measurement of regional cerebral blood flow using microsphere model and N-isopropyl-p-[123I]iodoamphetamine (IMP): comparison with 133Xe SPECT and sequential dynamic 123I-IMP SPECT].

Regional cerebral blood flow (rCBF) was measured in 13 patients with various cerebral disease by N-isopropyl-p-[123I]iodoamphetamine (IMP) SPECT with the microsphere model (rCBF (CbST)), which was compared with 133Xe inhalation SPECT (rCBFXe). Continuous withdrawal of arterial blood was performed for 5 minutes and SPECT image was obtained from 25 min to 55 min after the tracer injection. The reconstructed counts at 5 min was corrected using entire cerebral counts collected at 5 min, 20 min, and 60 min after the tracer injection. As compared with rCBFXe, rCBF (CbST) was underestimated in high flow areas and overestimated in low flow areas. Regional CBF was also measured using one minute short time SPECT images at 5 min (rCBF (Cb5)), 20 min (rCBF (Cb20)), and 60 min (rCBF (Cb60)) after the injection. Regional CBF tended to be underestimated with time from the tracer injection due to the wash-out from brain tissue to blood. In comparison with rCBF (Cb5), rCBF (CbST) was underestimated in high flow areas and overestimated in low flow areas. This error was considered to be due to the correction of reconstruct counts using entire cerebral counts, because of the kinetic behavior of 123I-IMP was different in each region of the brain.

Adult↗
PubMed1993

Goniodomin A induces modulation of actomyosin ATPase activity mediated through conformational change of actin.

Goniodomin A, one of the polyether macrolides, isolated from the dinoflagellate Goniodoma pseudogoniaulax stimulated skeletal actomyosin ATPase activity 2-3 times, with a peak at 3 x 10(-8) to 3 x 10(-7) M. The effect of goniodomin A was dependent on the concentration of actin, but not of myosin. The actomyosin ATPase activity was increased by pretreatment of actin (but not of myosin) with goniodomin A. Goniodomin A induced a sustained and concentration-dependent increase in the fluorescence intensity (excitation wavelength, 277 nm; emission wavelength, 329 nm) of actin. The maximum response was obtained with concentrations of goniodomin A in the 10(-5) to 10(-4) M range in the presence of 5 microM F-actin. However, the ATPase activity and fluorescence intensity of myosin were not changed by goniodomin A at concentrations from 10(-8) to 10(-5) M. Interestingly, goniodomin A induced a remarkable but transient increase in the fluorescence intensity of actomyosin in a concentration-dependent manner, with a peak at 3 x 10(-7) M. This profile was quite similar to that found in the stimulation of the actomyosin ATPase activity induced by goniodomin A. To investigate further the effect of goniodomin A, actin was labeled with N-(1-pyrenyl)iodoacetamide. Goniodomin A at 10(-6) M had no effect on the fluorescence intensity of pyrenyl-actin (excitation wavelength, 365 nm; emission wavelength, 407 nm), but increasing concentrations of goniodomin A to 3 x 10(-6) M remarkably decreased its intensity. This effect was potentiated by heavy meromyosin. Actin molecules treated with goniodomin A were completely sedimented by mild centrifugation (for 15 min at 12,000 x g). Electron microscopic observations suggest that actin filaments associate with each other to form a gel in the presence of 3 x 10(-6) M goniodomin A. The conformational change of actin molecules, resulting from stoichiometric binding of goniodomin A to actin monomers in filaments, may modify the interaction between actin and myosin.

Actins↗
PubMed1993

Xenogeneic dopaminergic grafts reverse behavioral deficits induced by 6-OHDA in rodents: effect of 15-deoxyspergualin treatment.

Foetal rat mesencephalic dopamine-containing tissue was transplanted into the lateral ventricle of mice previously subjected to a 6-OHDA lesion of dopaminergic nerve terminals in the corpus striatum. The graft recipients were immunosuppressed by subcutaneous injections of 15-deoxyspergualin (DSG). Four weeks postgrafting, all DSG-treated mice showed partial or complete functional compensation in amphetamine-induced motor asymmetry. The immunohistochemical staining of tyrosine hydroxylase (TH) revealed large numbers of surviving dopamine neurons and abundant fibers in the grafted animals. In contrast, all grafts in non-DSG-treated animals were rejected and functional compensation was lacking. It is concluded that DSG treatment promotes xenogeneic intracerebral graft survival, recovery of function and reduce the histological sign of rejection.

Amphetamine↗
PubMed1993

Suppression of immunorejection against rat fetal dopaminergic neurons in a mouse brain by 15-deoxyspergualin.

In this study, the immunosuppressive effect of 15-deoxyspergualin (DSG) on the survivability of rat embryonic dopaminergic neurons grafted into the lateral ventricle of adult mice was investigated. DSG was administered daily in dose of 5 mg/kg for 2 weeks postgrafting, commencing on the day of transplantation, in the immunosuppressant-treated groups. Animals were then allowed to survive for 2 to 4 weeks after transplantation. Histological analysis revealed that most of transplanted rat fetal neural tissue was destroyed and scavenged in 2 weeks after transplantation in the non-immunosuppressed group. However, a large number of tyrosine hydroxylase (TH)-positive grafted neurons survived and grew postgrafting in the brain of the host administered with DSG even if they suffered from T lymphocyte infiltrations visualized by cytotoxic T cell immunohistochemistry. The results thus indicated that DSG is an potent immunosuppressant in neural transplantation as well as in transplantation of other organs in animals. It seems to be able to block, at least in part, the ability of mature specific cytotoxic T cells to lyse their targets.

Animals↗
PubMed1993

Spectral analyses of PGO-on neurons during paradoxical sleep in freely moving cats.

We have investigated the power spectral density (PSD) characteristics of spontaneous neuronal activities during paradoxical sleep (PS), including phasically firing PGO-on and tonically discharging type I-R neurons in the pontine tegmentum. The PSD of lateral geniculate PGO waves and blood pressure (BP) were also examined. During PS, the PSD of PGO-on neurons and PGO waves showed an almost white noise-like pattern, while that of TI-R neurons and BP exhibited a 1/f spectrum. It appears, therefore, that the neuronal and autonomic slow fluctuations found in the 1/f spectrum during PS are not driven by the PGO wave system.

Animals↗
PubMed1993

Pulmonary surfactant is a potential endogenous inhibitor of proteolytic activation of Sendai virus and influenza A virus.

The pathogenicities of influenza viruses and paramyxoviruses have been proposed to be primarily determined by a host cell protease(s) that activates viral infectivity by proteolytic cleavage of the envelope glycoproteins. We recently isolated a trypsin-type endoprotease, named tryptase Clara, from rat bronchial and bronchiolar epithelial Clara cells, which is secreted into the airway lumen and activates Sendai virus and influenza A virus proteolytically. We report here that surfactant in the bronchial fluid inhibited tryptase Clara specifically, having a Ki value of 0.13 microM, and inhibited the proteolytic activations by tryptase Clara in vitro and in organ cultures of rat lung. Intranasal infection of rats with Sendai virus was shown to stimulate secretion of tryptase Clara without changing the amount of surfactant in the bronchial lumen, resulting in a preferable condition for proteolytic viral activation and multiplication.

Amino Acid Sequence↗
PubMed1993

Transmembrane calcium influx associated with von Willebrand factor binding to GP Ib in the initiation of shear-induced platelet aggregation.

We found that the binding of multimeric vWF to GP Ib under a shear force of 108 dynes/cm2 resulted in the transmembrane flux of Ca2+ ions with a two- to three-fold increase in their intracellular concentration ([Ca2+]i). The blockage of this event, obtained by inhibiting the vWF-GP Ib interaction, suppressed aggregation. In contrast, the blockage of vWF binding to GP IIb-IIIa, as well as the prevention of activation caused by increased intracellular cAMP levels, inhibited aggregation but had no significant effect on [Ca2+]i increase. A monomeric recombinant fragment of vWF containing the GP Ib-binding domain of the molecule (residues 445-733) prevented all effects mediated by multimeric vWF but, by itself, failed to support the increase in [Ca2+]i and aggregation. These results suggest that the binding of multimeric vWF to GP Ib initiates platelets aggregation induced by high shear stress by mediating a transmembrane flux of Ca2+ ions, perhaps through a receptor-dependent calcium channel. The increase in [Ca2+]i may act as an intracellular message and cause the activation of GP IIb-IIIa; the latter receptor then binds vWF and mediates irreversible aggregation.

Biological Transport↗
PubMed1993

Phosphorylation sites of myelin basic protein by a catalytic fragment of non-receptor type protein-tyrosine kinase p72syk and comparison with those by insulin receptor kinase.

Myelin basic protein has been used as a model substrate for determination of substrate recognition motif of various protein kinases. In this report phosphorylated sites of bovine brain myelin basic protein were studied with a catalytic fragment of protein-tyrosine kinase p72syk. Three of four tyrosine residues in myelin basic protein were phosphorylated by this kinase. Major phosphorylated site was 134Y and minor phosphorylated sites were 68Y and 127Y. As the phosphorylation site by p56lck was only 68Y, the recognition motif of p72syk was quite different from that of p56lck. Furthermore, the fact that elution pattern on HPLC of the phosphopeptides obtained by insulin receptor kinase was different from that by p72syk suggested that the recognition motif of p72syk was also different from that of insulin receptor kinase. These results may suggest that each protein-tyrosine kinase has a specific substrate recognition motif.

Amino Acid Sequence↗
PubMed1993

Effects of putative cognitive function-enhancing drugs and dopaminergic agents on somatostatin and neuropeptide Y in rat brain.

Reduced levels of somatostatin and neuropeptide Y (NPY) have been demonstrated in the brain of patients with some organic mental disorders. We designed the present study to test whether drugs thought to be effective in improving cognitive functions in these disorders increased the levels of these two peptides. The drugs were given to normal rats for 14 days to examine chronic effects on regional brain somatostatin and NPY levels. Amantadine and bifemelane increased these peptide levels. Idebenone and indeloxazine had little effect. The effect of dopaminergic agents on peptide levels was also tested in rats. 1-Dihydroxyphenylalanine increased somatostatin levels in whole brain, and hypothalamic NPY levels, and reduced NPY levels in the cerebral cortex, hippocampus and striatum. Sulpiride increased the levels of NPY in the striatum and brainstem. 6-Hydroxydopamine decreased cortical somatostatin levels, and increased striatal NPY levels. These findings indicate that some agents used to improve cognitive function impairment in organic mental disorders may increase somatostatin and NPY content in the brain of rats. Also, it is suggest that these peptides are under different influences of the dopaminergic system in the brain.

Animals↗
PubMed1993

Clinical observations on adoptive immunotherapy with vaccine-primed T-lymphocytes secondarily sensitized to tumor in vitro.

The adoptive immunotherapy of human malignancy requires reliable methods to sensitize and expand patients' T-cells reactive to autologous tumors. In animal studies, we have generated therapeutic effector cells against a poorly immunogenic tumor by a two-step procedure: vaccination of the host followed by the secondary stimulation of vaccine-primed lymph node (LN) cells by in vitro sensitization (IVS) with tumor in the presence of interleukin 2 (IL-2). Based on these observations, we performed a clinical trial in patients with advanced cancer to evaluate the antitumor efficacy of vaccine-primed LN cells which were similarly activated in vitro. Patients were vaccinated with irradiated autologous tumor admixed with Bacillus Calmette-Guérin and had draining LN excised 10 days later for IVS culture. During IVS culture, LN cells expanded up to 14-fold (average of 8.4-fold). A mean of 6.7 x 10(9) cells was infused in ten patients (seven melanoma, three renal cell cancer) along with the concomitant i.v. administration of IL-2 (180,000 IU/kg every 8 h for 5 days). Phenotype analysis of IVS-LN cells revealed 78 +/- 4% CD3+ T-cells which were predominantly CD4+ (67 +/- 5%) with expression of HLA-DR and IL-2 receptor. IVS-LN cells displayed relative specificity of autologous tumor lysis in four of ten cases compared to zero of seven IVS-peripheral blood lymphocytes derived from the same patients as measured by the 51Cr release assay. One mo after therapy, seven of nine patients treated with IVS-LN cells and IL-2 developed delayed-type hypersensitivity reactivity to autologous tumor compared to zero of nine patients treated with tumor vaccination and IL-2 only (P < 0.002). These observations suggest that antitumor reactivity was passively transferred with the IVS-LN cells. Major toxic side effects including fever, hepatic dysfunction, and weight gain associated with the capillary leak syndrome were associated with exogenous IL-2 administration. Tumor vaccination and cell transfer were well tolerated without significant complications. Of the ten patients treated with IVS-LN cells and IL-2, there were one partial and one minor response, and one patient has had stable disease for 27+ mo. There was no evidence of tumor response in ten patients treated with tumor vaccination and IL-2 only. Further clinical studies evaluating the antitumor reactivity of vaccine-primed LN cells are warranted.

Adult↗
PubMed1993

Cloning of two isozymes of Trichoderma koningii glyceraldehyde-3-phosphate dehydrogenase with different sensitivity to koningic acid.

Koningic acid inhibits glyceraldehyde-3-phosphate dehydrogenase (GAPDH) by binding to the SH group in the active center. The fungus Trichoderma koningii, the producer of koningic acid, contains two GAPDH isozymes (GAPDHs I and II). GAPDH I is inhibited 50% by 1.1.10(-3) M koningic acid, while GAPDH II is inhibited 50% at 6.8 x 10(-6) M. cDNAs of the two isozymes were cloned from T. koningii and their nucleotide sequences were determined. The sequence of coding region and codon usage in both clones were compared with each other and with those of the gene for Aspergillus nidulans GAPDH (enzyme activity is inhibited 50% by 2.7 x 10(-7) M koningic acid). Results indicated that GAPDH II is more closely related to A. nidulans GAPDH than GAPDH I. All essential amino acid residues, except 174 and 181, which are implicated in catalysis and binding of NAD and substrates, were conserved among A. nidulans GAPDH and GAPDHs I and II. Residues 174 and 181 are threonine in both A. nidulans GAPDH and GAPDH II, but alanine and serine, respectively, in GAPDH I. The side-chain of alanine-174 in GAPDH I can not replace threonine-174 functionally as threonine-174 side-chain forms a hydrogen bond with the catalytically essential histidine-176.

Amino Acid Sequence↗
PubMed1993

Protein-tyrosine kinase p72syk is activated by thrombin and is negatively regulated through Ca2+ mobilization in platelets.

Activation of platelets by thrombin results in a dramatic increase in tyrosine phosphorylation on multiple cellular proteins (Ferrell, J. E., and Martin, G. S. (1988) Mol. Cell. Biol. 8, 3603-3610; Golden, A., and Brugge, J. S. (1989) Proc. Natl. Acad. Sci. U. S. A. 86, 901-905; Nakamura, S., and Yamamura, H. (1989) J. Biol. Chem. 264, 7089-7091). However, none of the responsible protein-tyrosine kinase has been reported so far. We report here that p72syk, one of the non-receptor-type protein-tyrosine kinases, is activated following thrombin stimulation in blood platelets. Washed porcine platelets were stimulated by thrombin, and the activation of p72syk was assessed in an immunoprecipitation kinase assay. The activity of p72syk increased within 5 s, reached a maximum at 10 s, and decreased to a basal level within 60 s after 0.5 unit/ml thrombin stimulation. The amount of immunoprecipitated p72syk was not altered throughout the time course. This activation was greatly enhanced in a dose-dependent manner and was completely canceled by the pretreatment of platelet suspension with hirudin, a specific antagonist of thrombin. In the Ca(2+)-depleted condition both extra- and intracellularly, the activation of p72syk was still persistent; in contrast, the deactivation process was completely abrogated even at 120 s after thrombin stimulation. In addition, the replenishment of Ca2+ resulted in a similar deactivation pattern as seen in the Ca(2+)-rich condition. Furthermore, this deactivation was also canceled by the pretreatment of platelets with W7, a calmodulin antagonist, as well as ML9, a myosin-light-chain kinase inhibitor. These results indicate that p72syk can be a responsible enzyme to the protein-tyrosine phosphorylation events following the platelet activation by thrombin and may be negatively regulated by Ca2+ in a calmodulin-dependent manner, inter alia myosin light-chain kinase, in thrombin-stimulated platelets.

Animals↗
PubMed1993

Origin of intracellular Ca2+ elevation induced by in vitro ischemia-like condition in hippocampal slices.

Microfluorometry was used to investigate the origin of intracellular Ca2+ ([Ca2+]i) elevation in field CA1 of gerbil hippocampal slices perfused with a glucose-free physiological medium equilibrated with a 95% N2/5% CO2 gas mixture (standard in vitro ischemia-like condition). Large [Ca2+]i elevation was detected about 4 min after the beginning of standard in vitro ischemia-like condition, which was accompanied with a negative shift of extracellular DC potential. When slices were perfused with Ca(2+)-free in vitro ischemia-like medium, large [Ca2+]i elevation was observed about 3.5 min after the beginning of Ca(2+)-free in vitro ischemia-like condition, however, the increase in [Ca2+]i was more gradual and of a lesser extent compared with that detected in the slices perfused with the standard in vitro ischemia-like medium that contained Ca2+. When slices were perfused with the Ca(2+)-free in vitro ischemia-like medium that contained dantrolene (50 microM) which is known to prevent Ca(2+)-induced Ca2+ release from intracellular Ca2+ stores, the increase in [Ca2+]i was more gradual and of a lesser extent compared with that detected in the slices perfused with the Ca(2+)-free in vitro ischemia-like medium that did not contain dantrolene. These results indicate that large [Ca2+]i elevation induced by in vitro ischemia-like condition in field CA1 of the hippocampus was caused by both Ca2+ influx from extracellular space and Ca2+ release from intracellular Ca2+ stores, and that a part of the Ca2+ release was due to Ca(2+)-induced Ca2+ release from intracellular Ca2+ stores.

Animals↗
PubMed1993

Analysis of autoantibody binding to 52-kd paraneoplastic cerebellar degeneration-associated antigen expressed in recombinant proteins.

A 52-kd neural antigen was reported to be recognized by anti-Purkinje cell antibodies in serum of a patient with paraneoplastic cerebellar degeneration associated with uterine carcinoma. In this study, we demonstrate that this neural antigen is recognized by antibodies known as anti-Purkinje cell antibody type I (PCAb Type I) and anti-YO. The latter's antigen is reported to be specific for the 62- to 64-kd antigen CDR62. Assuming that the 52-kd and 62- to 64-kd antigens share a common epitope(s) recognized by all of these antibodies, we examined the antigenic region on the 52-kd protein by immunoblots with deletion fragment proteins of the recombinant 52-kd protein. A major epitope was localized in the region of amino acid residues 94 to 133 of the 52-kd protein, which is the site of a leucine zipper motif. The potential pathogenicity of PCAb Type I is discussed.

Amino Acid Sequence↗
PubMed1993

Porous polyurethane tubes as vascular graft.

A vascular graft with the inner diameter of about 3 mm was prepared from segmented poly (ether urethane) with an extrusion technique. To make the wall of the vascular grafts porous, NaCl salts were added to the polyurethane solution to be extruded and removed with water extraction after evaporating the solvent in the extruded tube. The wall was reinforced with elastic fiber to prevent dilation. The compliance of the vascular graft measured with the method of Hayashi et al. ranged from 0.2 to 0.3% mmHg -1. The initial Young's modulus was close to that of canine carotic artery, to which the porous polyurethane graft 4-cm long was anastomosed. Vascular grafts were occluded within 2 weeks after implantation, when their pore size was 0, 1.7, or 4.4 mum, whereas those with the pore size of 5.5, 7.4, and 30 mum were patent for longer than 4 weeks. When the vascular graft with the pore size of 30 mum was implanted for 6 months, the luminal surface was covered with neointima, but the endothelium-like cells appearing in the middle of the intima of the vascular graft were immature and sometimes had a very big nucleus. In addition, spindle-shaped, modified smooth muscle cells were noticed in the deep layer of the neointima, especially in the tissue where anastomotic intimal hyperplasia occurred.

Animals↗