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Biomedical subjects

K Nagata

Publications and source records attributed to K Nagata.

At least 397 records · Page 22Linked to original sources
PubMed1996

Differential effects of hexachlorocyclohexane isomers on the GABA receptor subunits expressed in human embryonic kidney cell line.

We have recently demonstrated by patch clamp experiments that the four isomers of hexachlorocyclohexane (HCH), alpha-, beta-, gamma- and delta-HCH insecticides, modulated the kinetics of the GABAA receptor-chloride channel complex of rat dorsal root ganglion neurons. The present paper reports the differential effects of HCH isomers of the GABA-induced chloride currents in three combinations of alpha, beta and gamma subunits of GABAA receptor expressed in human embryonic kidney cells. When co-applied with GABA, gamma-HCH strongly suppressed the peak amplitude of GABA-induced current, and delta-HCH strongly enhanced it in the alpha 1 beta 2 gamma 2s, alpha 1 beta 2, alpha 6 beta 2 gamma 2s combinations in a dose-dependent manner. There was little or no difference in the dose dependence of the effects between gamma- and delta-HCH in any of the three subunit combinations. However, alpha- and beta-HCH showed differential effects on GABA-induced chloride currents in the three subunit combinations tested. alpha-HCH showed enhancing effects on the peak current in alpha 1 beta 2 gamma 2s, small enhancing effects on alpha 1 beta 2, and biphasic effects on alpha 6 beta 2 gamma 2s subunit combinations. beta-HCH had little or no effect on the peak current in alpha 1 beta 2 gamma 2s and alpha 1 beta 2 combinations, but suppressed currents in the alpha 6 beta2 gamma 2s subunit combination in a dose-dependent manner. The differential actions of HCH isomers may produce variable effects on different regions of the nervous systems and in different species of animals.

Cell Line↗
PubMed1996

Potent modulation of neuronal nicotinic acetylcholine receptor-channel by ethanol.

Controversies remain over which ion channels are the most sensitive to ethanol. We have found that ethanol potently modulates the neuronal nicotinic acetylcholine receptor-channel at micromolar concentrations with an EC50 of 88.5 microM, which is significantly lower than most values previously reported for other ion channels. Prolonged application of ethanol accelerated the decay phase of acetylcholine-induced currents, caused single-channels to open in bursts, and shortened the mean open time, all of which reflect increased receptor desensitization. However, ethanol slowed the decay phase of the current induced by a brief application of acetylcholine, which may indicate that ethanol manifests its action by causing an increase in the affinity of the receptor for acetylcholine. These results suggest that neuronal nicotinic acetylcholine receptors may be important target sites of ethanol, particularly in the early stages of ethanol intoxication.

Alcoholic Intoxication↗
PubMed1996

Cytochrome P450 enzymes involved in the enhancement of propranolol N-desisopropylation after repeated administration of propranolol in rats.

Repeated oral administration of propranolol (PL, 100 mg/kg daily, for 5, 10 and 15 days) to male Wistar rats increased PL N-desisopropylase and decreased PL 4-,5- and 7-hydroxylase activities in liver microsomes. The increase was highest at the 10 day time point whereas the decrease was relatively constant over the 15 day treatment period. There were no significant changes in the total content of cytochromes P450 (P450) or cytochrome b5 or in NADPH-cytochrome c reductase activity during the PI, treatment. The enhanced N-desisopropylase activities were markedly inhibited by alpha-naphthoflavone (a P450-1A1/2 inhibitor), and moderately by triacetyloleandomycin (a P450-3A1/2 inhibitor) and diethyldithiocarbamate (a P450-2E1 inhibitor). Phenacetin O-deethylase activity, an index of P450-1A2, was significantly increased on day 5, 10 and 15 of the treatment, whereas p-nitrophenol hydroxylase activity was elevated on day 10 only. The PL N-desisopropylation showed a strong and significant correlation with phenacetin O-deethylation, and a weaker but significant correlation with p-nitrophenol hydroxylation. Immunoblot analysis revealed that a protein band corresponding to P450-1A2 was increased by PL pretreatment, and protein band corresponding to P450-3A tended to be increased slightly, but other protein band corresponding to the subfamily of P450-2B, -2C, or -2E was not changed. Pretreatment of rats with P450 inducers (beta-naphthoflavone, phenobarbital, acetone and dexamethasone) increased PL N-dealkylase activity in liver microsomes. Furthermore, antibodies raised against P450-1A and -3A enzymes suppressed PL N-desisopropylation in a concentration-dependent manner, but P450-2E antibody did not. Reconstitution studies showed that P450-1A1, -1A2, -2E1 and -3A2 exhibited catalytic activities for PL N-dealkylation. These results suggest that P450-1A2 is a major PL N-desisopropylase in the PL-treated rats, and P450-3A related enzyme(s) and P450-2E1 as a moderate or minor enzyme are also involved in PL N-dealkylation in native and PL-treated rats.

Administration, Oral↗
PubMed1996

Antiviral effect of Sanicula europaea L. leaves extract on influenza virus-infected cells.

We have investigated the influence of Sanicula europaea L. extracts on influenza virus growth in MDCK cells. Fractions I, II, and III separated from Sanicula extract with Sephadex column chromatography were found to be non-toxic against MDCK cells. The growth of influenza A/PR/8/34 was completely inhibited by these fractions, while that of influenza B/Lee/40 was not affected. Fractions II and III have been shown not to have a direct virucidal activity on influenza A/PR/8/34. Influenza A/Vic/1/75 produced microscopic plaques in the presence of the extract. In vitro RNA synthesis with viral RNA-dependent RNA polymerase was also inhibited by a water soluble extract of Sanicula. These observations suggest that the Sanicula extract contains an anti-influenza virus substance.

Animals↗
PubMed1996

Insulin and insulin-like growth factors induce expression of angiotensin type-2 receptor in vascular-smooth-muscle cells.

Angiotensin type-2 receptor (AT2) is abundant in fetal tissues, including aorta, and its expression level declines after birth. In the present study, the regulation of its expression was studied in cultured vascular-smooth-muscle cells (VSMC). The maximum number of binding sites of AT2 increased in VSMC after they were cultured without serum in the presence of insulin, which was essential for its expression. AT2 expression was inhibited by treatment with phorbol ester. Northern blot analyses revealed that insulin-dependent expression is due to elevation of mRNA level of AT2. Similar induction was observed when insulin-like growth factor (IGF)-I or IGF-II was used instead of insulin. The study on the dose dependencies of these factors revealed that the induction of AT2 expression was mediated through the activation of IGF-I receptors. The insulin-induced expression of AT2 was detected in the aorta of genetically obese (fa/fa) Zucker rats, which reportedly have approximately tenfold-higher plasma concentrations of insulin than their lean littermates. The insulin-dependence seems characteristic of VSMC, because it was not observed for pheochromocytoma cells or adrenal glands. These results suggest that the expression of AT2 is regulated by at least two mechanisms, that is, IGF-I receptor dependent and IGF-I receptor independent, and that the former may play an important role in the expression of AT2 in VSMC.

Animals↗
PubMed1996

Subconductance-state currents generated by imidacloprid at the nicotinic acetylcholine receptor in PC 12 cells.

Nitromethylene heterocyclic insecticides are known to act on the nicotinic acetylcholine receptor-channel complex. The effects of the nitromethylene heterocycle imidacloprid on the nicotinic acetylcholine receptor of clonal rat pheochromocytoma (PC 12) cells were studied using the single-channel patch clamp technique. Imidacloprid generated single-channel currents of multiple conductance states in PC12 cells. Both acetylcholine and imidacloprid induced single-channel currents of main conductance and subconductance states with conductances of 33.3 and 9.4 pS by acetylcholine and 30.4 and 9.8 pS by imidacloprid. However, the main conductance currents were generated predominantly by acetylcholine, whereas the subconductance currents were generated predominantly by imidacloprid. Partial block of acetylcholine-induced currents by imidacloprid may be explained on the basis of the single-channel behavior.

Animals↗
PubMed1996

Interactions of tetramethrin, fenvalerate and DDT at the sodium channel in rat dorsal root ganglion neurons.

Type I and type II pyrethroids and dichlorodiphenyltrichloroethane (DDT) are known to modulate the sodium channel to cause the hyperexcitatory symptoms of poisoning in animals. However, since the degrees to which neuronal sodium channel parameters are altered differ, a question is raised as to whether these insecticides bind to the same site in the sodium channel. Competition patch-clamp experiments were performed using rat dorsal root ganglion neurons which are endowed with tetrodotoxin-sensitive and tetrodotoxin-resistant sodium channels. D-trans-Tetramethrin, S,S-fenvalerate and p,p'-DDT caused a slowly rising and slowly falling tail current to be developed in tetrodotoxin-sensitive sodium channels. In tetrodotoxin-resistant sodium channels, these insecticides, particularly tetramethrin and fenvalerate, generated a large and prolonged tail current upon repolarization. The effects of tetramethrin were reversible after washing with drug-free solution, whereas the effects of fenvalerate and DDT were irreversible. When fenvalerate application was followed by tetramethrin application, the characteristic changes in current by fenvalerate disappeared and the characteristic changes by tetramethrin appeared. After washout, the characteristic current pattern of fenvalerate reappeared. These results can be explained by assuming that the tetramethrin molecule displaces the fenvalerate molecule from the same binding site in the sodium channel protein, or that tetramethrin and fenvalerate bind to separate sodium channel sites which interact allosterically with each other. DDT interacted with fenvalerate and tetramethrin in the same manner.

Animals↗
PubMed1996

Anticoagulant effects of retinoic acids on leukemia cells.

We have recently found that all-trans retinoic acid (ATRA) upregulates thrombomodulin (TM) and downregulates tissue factor (TF) expression in acute myelogenous leukemia (AML) M3 cells (NB4) and acute monoblastic leukemia cells (U937) (Koyama et al, Blood 84:3001, 1994). We have further investigated the effects of ATRA on leukemic cells freshly isolated from patients at diagnosis. Increase of TM antigen was documented in all AML cells: M0 (n = 1), M2 (n = 5), M3 (n = 3), M4 (n = 3), M5 (n = 3), and M6 (n = 1). Decrease of TF antigen was observed in 4 M2, 1 M4, and all M3 and M5 patients. However, no TM and TF antigens were detected in all chronic lymphocytic leukemia cells (n = 3) with or without ATRA treatment. Changes of TM and TF antigen levels were associated with those of TM and TF cofactor levels on the cell surface. A stereoisomer of RA, 9-cis RA, is a high-affinity ligand for the RA receptors (RARs) and the retinoid X receptors, although ATRA and another isomer, 13-cis RA, solely bind to RARs. We have also studied the effects of 9-cis RA and 13-cis RA on the expressions of TM and TF in NB4 and U937 cells. A relatively wide range of 9-cis RA concentrations (0.01 to 1 mumol/L) compared with ATRA was optimal for prolongation of normal plasma-based recalcification time (reduction of cell surface TF activity), decrease of TF antigen, and increase of TM antigen on the surface and in the lysates of NB4 and U937 cells. Western blot analysis under nonreducing conditions showed that both ATRA and 9-cis RA markedly induced the prominent band at 75 kD of TM and reduced the band at 45 kD of TF. Northern blot analysis has shown similar changes of mRNA levels, which indicates that RAs regulate TM and TF expression in leukemic cells at transcriptional levels. Anticoagulant effects of ATRA, ie, upregulation of TM expression and downregulation of TF expression, are applied not only to established cell lines of specific subtypes (M3 and M5) but also to more universal AML (most cases of M3 and M5 and a part of the other types of AML) cells freshly isolated from patients. 9-cis RA may be more effective than ATRA as an inducer of differentiation of AML M3 cells and as an anticoagulant agent for patients with certain types of AML as well.

Anticoagulants↗
PubMed1996

Expression of 47 kDa heat shock protein (HSP47) during development of mouse cornea.

The 47-kDa heat shock protein (HSP47) is assumed to be a molecular chaperone specific to collagen. We examined and compared the expression patterns of HSP47 and alpha 1(I) collagen in the developing mouse cornea. Expression of HSP47 and alpha 1(I) collagen mRNAs was assessed by Northern blot analysis and in situ hybridization, and that of HSP47 protein was determined by Western immunoblotting and immunohistochemistry. HSP47 mRNA and protein were expressed strongly in early embryonic ocular tissues and were developmentally down-regulated during ocular development. Both were distributed in the mesenchymal cells (presumptive corneal stroma, sclera and choroid) and hyaloid blood vessels until post-natal day 14. The level of alpha 1(I) collagen mRNA increased during ocular development, was maximally expressed in the three days after birth, and was detected in mesenchymal cells. HSP47 expression is regulated developmentally toward completion and plays an important role during ocular development especially in corneal morphogenesis. We discuss here the relation between expression of HSP47 and alpha 1(I) collagen.

Animals↗
PubMed1996

Regulation of thermotolerance and ischemic tolerance.

Thermotolerance and ischemic tolerance are two major biological aspects where heat shock (stress) proteins exert essential roles for survival in cells as well as in various tissues. Bioflavonoids prevent the cells from acquiring thermotolerance after stresses through specific inhibition in the induction of heat shock proteins. The mechanism of this inhibition is revealed to be due to the prevention of the activation of heat shock factor 1 after heat shock. The induction of stress proteins during the ischemic stress is then described in global as well as focal cerebral ischemic model in rats. The activation of heat shock factor 1 after ischemia is first shown to induce various stress proteins in the central nervous system.

Brain↗
PubMed1996

Cerebral oxygen and glucose metabolism and blood flow in mitochondrial encephalomyopathy: a PET study.

Cerebral blood flow (CBF), oxygen metabolism (CMRO2), and glucose metabolism (CMRGlc) were measured using positron emission tomography in five patients diagnosed as having mitochondrial encephalomyopathy. The molar ratio between the oxygen and glucose consumptions was reduced diffusely, as CMRO2 was markedly decreased and CMRGlc was slightly reduced. The CBF showed less changes. The CBF increase on hypercapnia was smaller than normal, though this was not significant. CBF with hypocapnia demonstrated a significant reduction compared with the normal. These results suggest that oxidative metabolism is impaired and anaerobic glycolysis relatively stimulated, due to a primary defect of mitochondrial function, and that mild lactic acidosis occurs in brain tissue because of impaired utilisation of pyruvate in the TCA cycle. As these findings appear to indicate directly a characteristic of this disease, such measurements may be a useful tool for assessment of the pathophysiology and for diagnosis of mitochondrial encephalomyopathy.

Acidosis, Lactic↗
PubMed1996

Correlation of EEG activities between slow-wave sleep and wakefulness in patients with supra-tentorial stroke.

Using topographic EEG mapping, we studied the relationships between delta activity during slow-wave sleep (SWS) and the background EEG activity during wakefulness, in 11 normal subjects and 35 stroke patients with unilateral supra-tentorial lesions. Delta-1 power during SWS showed a significant positive correlation with alpha-1 power during wakefulness, in both hemispheres. Delta-1 and delta-2 power during SWS correlated positively not only with alpha-2 power, but also with delta-1 and delta-2 power during wakefulness in the affected hemisphere. these figures indicate that the amount of delta activity during SWS can be associated with that of alpha activity during wakefulness. A close negative correlation was observed between delta power during SWS and the age of the subjects in the patient group. The Barthel index showed no significant correlation with delta-1 or delta-2 power in either hemisphere in patient group. Our results suggest that delta activity during SWS may be associated with dysfunction of the cerebral cortex in stroke patients as well as in normal aged subjects.

Adult↗
PubMed1996

Scalp topography of SEP late components in patients with supra-tentorial lesions.

Somatosensory evoked potentials (SEP) to simultaneous bilateral median nerve stimulation were recorded in 7 patients with unilateral brain lesions, 8 patients with degenerative dementia, and 5 normal volunteers. Right-left amplitude difference was compared from serial topographic images and the amplitude was compared at homologous electrodes. In patients with unilateral lesions, the amplitude from the frontopolar, frontal, anterior-temporal, and/or occipital electrodes was smaller on the affected side at 240 and 360 msec, regardless of whether the subjects showed sensory deficit or not. No significant laterality was seen in the primary sensorimotor areas. Distribution obtained from the patients with degenerative dementia was quite symmetrical. A significant right-left amplitude difference was seen at the anterior temporal site at 180 and 240 msec in normal controls, although an almost symmetrical distribution was obtained on the topographic images. The late components of SEP are not associated with the primary somatosensory function, but possibly with other cortical pathways.

Aged↗
PubMed1996

Acute gastric anisakiasis: 28 cases during the last 10 years.

Anisakiasis is a disease which occurs following eating raw fish infected with anisakis larvae. Many cases have been reported from Japan and from other countries with increasing opportunities of eating raw fish such as "sushi" and "sashimi." We have reviewed 28 patients with acute gastric anisakiasis during the last 10 years from November 1984 to October 1994. This disease has rarely been detected in persons over 60 years of age and in patients with gastric surgery. Therefore it is postulated that gastric acid secretion influences the activities of anisakis larvae. An alkaline gastric pH could interfere with the toxicity of anisakis larvae.

Adult↗
PubMed1996

A mutant trypsin-like enzyme from Streptomyces fradiae, created by site-directed mutagenesis, improves affinity chromatography for protein trypsin inhibitors.

The Ser-170 residue of a trypsin-like enzyme from Streptomyces fradiae (SFT), which is considered to be the active-site serine, was replaced with alanine by site-directed mutagenesis to improve the affinity chromatography step for a Kazal-type trypsin inhibitor pancreatic secretory trypsin inhibitor (PSTI). The resulting mutant SFT, designated as [S170A]SFT, was expressed in Streptomyces lividans and purified to homogeneity. [S170A]SFT was catalytically inactive, but still had the ability to bind tightly to PSTI and to soybean trypsin inhibitor with dissociation constants of 3.1 x 10(-7) M and 1.9 x 10(-8) M respectively. We further demonstrated that recombinant human PSTI secreted into Saccharomyces cerevisiae culture broth could be purified to homogeneity with a one-step [S170A]SFT-affinity column. The purified PSTI contained no molecules intramolecularly cleaved by active trypsin, which are found when trypsin-affinity chromatography is used for the purification. This eliminated the need for further separation of intact PSTI from intramolecularly cleaved PSTI by high-performance liquid chromatography, thus simplifying and improving its purification process.

Chromatography, Affinity↗
PubMed1996

Expression of heat-shock protein 47 in mouse liver.

Expression of heat-shock protein 47 in intact and fibrotic liver and in hepatic constituent cells was investigated in mice. Immunohistochemical study of intact liver and Western blot analysis of the protein from isolated liver cells revealed that stellate cells and smooth muscle cells of interlobular vessels, but not hepatocytes, Kupffer cells, or endothelial cells, expressed heat-shock protein 47. The protein was found in both vitamin-A-storing stellate cells and myofibroblast-like cells. The amount of the protein in cultured stellate cells was reduced by dexamethasone but was not regulated by quercetin, transforming growth factor beta, interferon gamma, or retinoic acid. In CCl4-treated or bile-duct-ligated mouse liver, the number of cells positive for heat-shock protein 47 markedly increased in the centrilobular area or around the periportal area, respectively, and the level of heat-shock protein 47 also increased.

Animals↗