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K Shimada

Publications and source records attributed to K Shimada.

At least 181 records · Page 10Linked to original sources
PubMed2000

Biophysical properties of ClC-3 differentiate it from swelling-activated chloride channels in Chinese hamster ovary-K1 cells.

ClC-3 is a highly conserved voltage-gated chloride channel, which together with ClC-4 and ClC-5 belongs to one subfamily of the larger group of ClC chloride channels. Whereas ClC-5 is localized intracellularly, ClC-3 has been reported to be a swelling-activated plasma membrane channel. However, recent studies have shown that native ClC-3 in hepatocytes is primarily intracellular. Therefore, we reexamined the properties of ClC-3 in a mammalian cell expression system and compared them with the properties of endogenous swelling-activated channels. Chinese hamster ovary (CHO)-K1 cells were transiently transfected with rat ClC-3. The resulting chloride currents were Cl(-) > I(-) selective, showed extreme outward rectification, and lacked inactivation at positive voltages. In addition, they were insensitive to the chloride channel blockers, 5-nitro-2-(3-phenylpropylamino)-benzoic acid (NPPB) and 4, 4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) and were not inhibited by phorbol esters or activated by osmotic swelling. These properties are identical to those of ClC-5 but differ from those previously attributed to ClC-3. In contrast, nontransfected CHO-K1 cells displayed an endogenous swelling-activated chloride current, which was weakly outward rectifying, inactivated at positive voltages, sensitive to NPPB and DIDS, and inhibited by phorbol esters. These properties are identical to those previously attributed to ClC-3. Therefore, we conclude that when expressed in CHO-K1 cells, ClC-3 is an extremely outward rectifying channel with similar properties to ClC-5 and is neither activated by cell swelling nor identical to the endogenous swelling-activated channel. These data suggest that ClC-3 cannot be responsible for the swelling-activated chloride channel under all circumstances.

Animals↗
PubMed2000

Serotonin increases interleukin-6 synthesis in human vascular smooth muscle cells.

BACKGROUND: Interleukin-6 (IL-6) is a key molecule in chronic inflammation and has been implicated in the progression of atherosclerosis. Serotonin (5-hydroxytryptamine; 5-HT) causes vascular contraction and proliferation, but its role in atherogenesis has not been clarified. We investigated the effects of 5-HT on IL-6 synthesis in human vascular smooth muscle cells (VSMCs). METHODS AND RESULTS: IL-6 levels in the culture medium of VSMCs were determined by ELISA. IL-6 mRNA accumulation was determined by use of a Quantikine mRNA colorimetric quantification kit. NF-kappaB activation was tested by gel retardation assay. 5-HT induced IL-6 production by VSMCs in a time- and dose-dependent manner, with increased IL-6 mRNA accumulation and nuclear factor-kappaB activation. The effect of 5-HT on IL-6 production was significantly inhibited by the 5-HT(2) receptor antagonist ketanserin and the selective 5-HT(2A) receptor antagonist sarpogrelate. Conversely, the 5-HT(2) receptor agonist alpha-methyl-5-HT increased IL-6 production. The protein kinase C (PKC) inhibitor calphostin C, but not the protein kinase A inhibitor KT5720, suppressed 5-HT-induced IL-6 production. The effect of 5-HT was also abolished in PKC-depleted VSMCs after pretreatment with phorbol 12-myristate 13-acetate for 24 hours. CONCLUSIONS: 5-HT acts on 5-HT(2A) receptors and increases IL-6 synthesis in human VSMCs at least partially through a PKC-dependent pathway. These results suggested that 5-HT may contribute to inflammatory activation of the vessels during atherogenesis.

Cells, Cultured↗
PubMed2000

On-line automated high-performance liquid chromatographic determination of total riboflavin phosphates using immobilized acid phosphatase as a pre-column reactor.

An automated chromatographic detection system for the determination of total riboflavin phosphates using immobilized sweet potato acid phosphatase as a pre-column reactor is reported on. An immobilized enzyme reactor, incorporated in the on-line analytical system, hydrolysed riboflavin phosphates to riboflavin, and then lipophilic riboflavin was concentrated at the top of an ODS trap column. Enzymatically hydrolysed riboflavin was back-eluted from the trap column using a mobile phase containing methanol, and then subsequently chromatographed on an ODS analytical column. The effluents were monitored by UV absorption at 280 nm. The calibration graph for total riboflavin phosphates, determined by this method, was linear over the range 0.5-500 nmol/ml, with a correlation coefficient of 0.9999. The detection limit at a signal-to-noise ratio of 3 was 25 pmol/ml. The average conversion rate of riboflavin phosphates to riboflavin was estimated at 97%. The relative standard deviations of the intra- and inter-assay precision were 1.2 and 2.6%, respectively.

Acid Phosphatase↗
PubMed2000

Biologically active oligodeoxyribonucleotides. Part 12: N2-methylation of 2'-deoxyguanosines enhances stability of parallel G-quadruplex and anti-HIV-1 activity.

2'-Deoxyguanosine residues of a 3',5'-end-modified hexadeoxyribonucleotide (R-95288) with anti-HIV-1 activity were substituted with N2-methyl-2'-deoxyguanosine (m2dG). These modified oligodeoxyribonucleotides (ODNs) showed a 2-fold higher activity than R-95288. Also, the CD spectra of these ODNs indicated that the m2dG modification stabilized the tertiary structure of the G-quadruplex.

Anti-HIV Agents↗
PubMed2000

Studies on neurosteroids XII. Determination of enzymatically formed catechol estrogens and guaiacol estrogens by rat brains using liquid chromatography-mass spectrometry-mass spectrometry.

Enzymic formations of catechol- and guaiacol-estrogens by rat brains have been investigated using classical- and catechol-estrogens as substrates, respectively. The incubation mixtures were pretreated with liquid-liquid and/or solid-phase extraction, and the products were identified by comparison with authentic samples using liquid chromatography-mass spectrometry (-mass spectrometry) [LC-MS (-MS)]. The enzymic activities were also determined by measuring the formed products with LC-MS.

Animals↗
PubMed2000

Common polymorphism in the promoter of the CD14 monocyte receptor gene is associated with acute myocardial infarction in Japanese men.

We investigated whether C(-260)-->T polymorphism in the promoter of the CD14 monocyte receptor gene predisposed to coronary atherosclerosis and acute myocardial infarction (AMI) in Japanese men. The frequencies of T allele and T/T homozygotes in patients with AMI were significantly higher than in controls and in patients with angina without prior AMI, suggesting that the CD14 promoter polymorphism is associated with AMI rather than with coronary atherosclerosis, and this polymorphism may be one of the genetic risk factors for AMI in Japanese men.

Aged↗
PubMed2000

Adoptive immunotherapy to lower postsurgical recurrence rates of hepatocellular carcinoma: a randomised trial.

BACKGROUND: Postsurgical recurrence of hepatocellular carcinoma (HCC) is frequent and fatal. Adoptive immunotherapy is active against HCC. We assessed whether postoperative immunotherapy could lower the frequency of recurrence. METHODS: Between 1992 and 1995, we did a randomised trial in which 150 patients who had undergone curative resection for HCC were assigned adoptive immunotherapy (n=76) or no adjuvant treatment (n=74). Autologous lymphocytes activated vitro with recombinant interleukin-2 and antibody to CD3 were infused five times during the first 6 months. Primary endpoints were time to first recurrence and recurrence-free survival and analyses were by intention to treat. FINDINGS: 76 patients received 370 (97%) of 380 scheduled lymphocyte infusions (mean cell number per patient 7.1x10(10) [SD 2.1]; CD3 and HLA-DR cells 78% [16]), and none had grade 3 or 4 adverse events. After a median follow-up of 4.4 years (range 0.2-6.7), adoptive immunotherapy decreased the frequency of recurrence by 18% compared with controls (45 [59%] vs 57 [77%]) [corrected] patients. Time to first recurrence in the immunotherapy group was significantly longer than that in the control group (48% [37-59] vs 33% [22-43] at 3 years, 38% [22-54] vs 22% [11-34] at 5 years; p=0.008). The immunotherapy group had significantly longer recurrence-free survival (p=0.01) and disease-specific survival (p=0.04) than the control group. Overall survival did not differ significantly between groups (p=0.09). INTERPRETATION: Adoptive immunotherapy is a safe, feasible treatment that can lower recurrence and improve recurrence-free outcomes after surgery for HCC.

Adult↗
PubMed2000

Evaluation of the effects of alpha-phenyl-N-tert-butyl nitrone pretreatment on the neurobehavioral effects of methamphetamine.

A relationship between formation of reactive oxygen species (ROS) and energy depletion has been proposed to play an important role in mediating methamphetamine (METH)-induced neurotoxicity. To evaluate this relationship, we examined the effect of the spin-trap agent, alpha-phenyl-N-tert-butyl nitrone (PBN) on hyperthermia and self-injurious behavior (SIB) and striatal dopamine (DA) depletion produced by METH (4 injections of 4 mg/kg, 2 hr intervals, s.c.) in BALB/c mice. Repeated administration of METH induced hyperthermia, incidence of SIB and striatal DA depletion (84% after 3 days). Pretreatment with PBN (4 injections of 60 or 120 mg/kg, i.p.) reduced METH-induced hyperthermia, but did not significantly attenuate METH-induced SIB or the striatal DA depletion. On the other hand, pretreatment with high doses of PBN (4 injections of 180 or 240 mg/kg, i.p.) protected against METH-induced hyperthermia and SIB, and PBN (180 mg/kg) also completely protected against the acute striatal DA depletion 60 min after the last injection of the drug. However, the long-lasting striatal DA depletion was only attenuated by 52 or 56%, respectively. These results indicate that METH-induced hyperthermia contributes to, but is not solely responsible for METH-induced neurotoxicity, and supports a role for formation of ROS and other mechanisms in the generation of METH-induced striatal dopaminergic neurotoxicity. In addition, the difference in the efficacy of PBN to protect against the acute or long-lasting striatal DA depletion induced by METH may indicate that both ROS formation and other mechanisms are required for METH-induced neurotoxicity to develop.

Animals↗
PubMed2000

Mediation by the protein-tyrosine kinase Tec of signaling between the B cell antigen receptor and Dok-1.

A variety of growth factor receptors induce the tyrosine phosphorylation of a nonreceptor protein-tyrosine kinase Tec as well as that of a Tec-binding protein of 62 kDa. Given the similarity in properties between this 62-kDa protein and p62(Dok-1), the possibility that these two proteins are identical was investigated. Overexpression of a constitutively active form of Tec in a pro-B cell line induced the hyperphosphorylation of endogenous Dok-1. Tec also associated with Dok-1 in a phosphorylation-dependent manner in 293 cells. Tec mediated marked phosphorylation of Dok-1 both in vivo and in vitro, and this effect required both the Tec homology and Src homology 2 domains of Tec in addition to its kinase activity. Expression of Dok-1 in 293 cells induced inhibition of Ras activity, suggesting that Dok-1 is a negative regulator of Ras. In the immature B cell line Ramos, cross-linking of the B cell antigen receptor (BCR) resulted in tyrosine phosphorylation of Dok-1, and this effect was markedly inhibited by expression of dominant negative mutants of Tec. Furthermore, overexpression of Dok-1 inhibited activation of the c-fos promoter induced by stimulation of the BCR. These results suggest that Tec is an important mediator of signaling from the BCR to Dok-1.

B-Lymphocytes↗
PubMed2000

Determination of incorporated amounts of poly(ethylene glycol)-derivatized lipids in liposomes for the physicochemical characterization of stealth liposomes.

We describe a method for determining incorporated amounts of poly(ethylene glycol) (PEG)-derivatized lipids in liposomes for the physicochemical characterization of PEG-coated liposomes. This method is based on the spectrophotometric determination of complexes of polyethers with sodium ions after their extraction as picrates into 1,2-dichloroethane, developed by Favretto for measuring levels of polyoxyethylene alkylphenyl-ether non-ionic surfactants in waste water. The same assay was applied to the estimation of PEG-derivatized lipids in liposomes and percent incorporation of PEG-derivatized lipids into liposomes was successfully determined. To prevent the interference from liposomal lipids other than PEG-derivatized lipids in this assay, liposomal samples were diluted at least to a concentration of less than 0.2 mM. The percent incorporation of PEG-lipids varied, depending on the molecular weight of PEG and anchor acyl chain length in PEG-lipids and it was suggested that the percent incorporation of PEG-lipids into liposomes would be a good parameter of quality control of PEG-liposomes in manufacturing facility and the picrate method used in the present study allows for the determination of this parameter without the need for hazardous radioisotopes.

Calibration↗
PubMed2000

Circadian component influences the photoperiodic induction of diapause in a drosophilid fly, Chymomyza costata.

The last-instar larvae of a drosophilid fly, Chymomyza costata enter diapause in response to the dark-phases longer than 9 h (Yoshida, T., Kimura, M.T., 1995. The photoperiodic clock in Chymomyza costata. Journal of Insect Physiology 41, 217-222). In order to switch the developmental programming of the sensitive larvae from continuous development to diapause, after they were transferred from the short (8 h) to the long (14 h) dark-phase, significantly less time (1-2 days) was required when the dark-phase was abruptly and asymmetrically extended into the evening, than when it was extended symmetrically into both morning and evening (2-3 days), or asymmetrically into the morning hours (4-6 days). Diapause was also induced in 40-70% of sensitive larvae that were reared under the gradually shortening light-phase (from 16 h to 2 h, by 1 h in each cycle), despite that the dark-phase remained constant and short (8 h). Larvae developed continuously, however, when reared under the gradually extending light-phase (from 16 h to 24 h) and a constantly short dark-phase. We interpret such results, with the help of the two-oscillator model of circadian rhythmicity (Pittendrigh, C.S., Daan, S., 1976. A functional analysis of circadian pacemakers in nocturnal rodents. V. Pacemaker structure: A clock for all seasons. Journal of Comparative Physiology A 106, 333-355), as indicating that two mutually coupled oscillators (evening and morning) differing in their entrainability may participate in measuring of the dark-phase duration. The levels of dopamine (DA) and serotonin (5-HT) in the larval CNS transiently increased (by up to 20%) after the dusk, while no apparent change was observed during the dawn. The dusk-related increase was observed also after the asymmetric extension of the dark-phase into evening, while the asymmetric extension into morning had no effect on the levels of the DA and 5-HT.

Journal Article↗
PubMed2000

Differential regulation of exonic regulatory elements for muscle-specific alternative splicing during myogenesis and cardiogenesis.

Muscle-specific isoform of the mitochondrial ATP synthase gamma subunit (F(1)gamma) was generated by alternative splicing, and exon 9 of the gene was found to be lacking particularly in skeletal muscle and heart tissue. Recently, we reported that alternative splicing of exon 9 was induced by low serum or acidic media in mouse myoblasts, and that this splicing required de novo protein synthesis of a negative regulatory factor (Ichida, M., Endo, H., Ikeda, U., Matsuda, C., Ueno, E., Shimada, K., and Kagawa, Y. (1998) J. Biol. Chem. 273, 8492-8501; Hayakawa, M., Endo, H., Hamamoto, T., and Kagawa, Y. (1998) Biochem. Biophys. Res. Commun. 251, 603-608). In the present report, we identified a cis-acting element on the muscle-specific alternatively spliced exon of F(1)gamma gene by an in vivo splicing system using cultured cells and transgenic mice. We constructed a F(1)gamma wild-type minigene, containing the full-length gene from exon 8 to exon 10, and two mutants; one mutant involved a pyrimidine-rich substitution on exon 9, whereas the other was a purine-rich substitution, abbreviated as F(1)gamma Pu-del and F(1)gamma Pu-rich mutants, respectively. Based on an in vivo splicing assay using low serum- or acid-stimulated splicing induction system in mouse myoblasts, Pu-del mutation inhibited exon inclusion, indicating that a Pu-del mutation would disrupt an exonic splicing enhancer. On the other hand, the Pu-rich mutation blocked muscle-specific exon exclusion following both inductions. Next, we produced transgenic mice bearing both mutant minigenes and analyzed their splicing patterns in tissues. Based on an analysis of F(1)gamma Pu-del minigene transgenic mice, the purine nucleotide of this element was shown to be necessary for exon inclusion in non-muscle tissue. In contrast, analysis of F(1)gamma Pu-rich minigene mice revealed that the F(1)gamma Pu-rich mutant exon had been excluded from heart and skeletal muscle of these transgenic mice, despite the fact mutation of the exon inhibited muscle-specific exon exclusion in myotubes of early embryonic stage. These results suggested that the splicing regulatory mechanism underlying F(1)gamma pre-mRNA differed between myotubes and myofibers during myogenesis and cardiogenesis.

ATP Synthetase Complexes↗
PubMed2000

Magnetic circular dichroism properties of reaction center complexes isolated from the zinc-bacteriochlorophyll a-containing purple bacterium Acidiphilium rubrum.

Reaction center (RC) complexes isolated from a Zn-bacteriochlorophyll (BChl) a-containing purple bacterium, Acidiphilium rubrum, were characterized by absorption, circular dichroism, and magnetic circular dichroism (MCD) spectroscopy. The oxidized-minus-reduced difference spectra indicated that, in this RC, the Zn-BChl a is the primary electron donor. The molecular structure of the donor was examined by measuring the ratio of the MCD intensity of the Faraday B-term (B) to the dipole strength (D). In the Q(y) region, B/D for the donor was about half those of bacteriopheophytin a and the accessory Zn-BChl a, indicating that the primary electron donor is a dimer. The magnitude of bleach of the Q(x) band was half that observed in Rhodobacter sphaeroides, suggesting the cation is localized on a single Zn-Bchl a. The absorption intensity of the higher-energy Q(y) exciton band was approximately 28% of that of the lower-energy band, and the exciton splitting was approximately 570 cm(-1), smaller than that in Rb. sphaeroides. These results indicate that, in A. rubrum, the primary electron donor is a Zn-BChl a dimer but that the interaction between the two molecules is rather weak. On the basis of these results, an adaptive strategy for changes in BChl a species is discussed from an evolutionary perspective.

Acetobacteraceae↗