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Biomedical subjects

S Ueno

Publications and source records attributed to S Ueno.

At least 163 records · Page 9Linked to original sources
PubMed1997

Enhancement by baclofen of the Gs-coupled receptor-mediated cAMP production in Xenopus oocytes expressing rat brain cortex poly (A)+ RNA: a role of G-protein beta gamma subunits.

We investigated the mechanism by which GABA-B receptors enhance the Gs-coupled receptor-mediated cAMP production in Xenopus oocytes expressing poly (A)+ RNA derived from rat brain cortex. We expressed the cystic fibrosis transmembrane conductance regulator gene (CFTR) as a reporter for cAMP changes in oocytes. The GABA-B agonist (-)baclofen enhanced the adrenergic beta 2 agonist isoproterenol- or vasoactive intestinal peptide (VIP)-induced CFTR currents, whereas (-)baclofen alone did not cause any currents. The (-)baclofen-enhanced currents were inhibited by the GABA-B antagonist 2-OH saclofen. The enhancement by (-)baclofen was further augmented by coexpressing adenylyl cyclase (AC) type II, an isotype activated by G beta gamma and G alpha s, but not by coexpressing AC type III, an isotype insensitive to G beta gamma. Moreover, pretreatment of the oocytes with pertussis toxin (PTX) abolished the enhanced effect of (-)baclofen. These results indicate that upon GABA-B activation, the G beta gamma released from PTX-sensitive G-proteins activates the AC type II (or IV), and this process requires the G alpha s activation by Gs-coupled receptors.

Adrenergic beta-Agonists↗
PubMed1997

Assembly of the chimeric Na+/K+-ATPase and H+/K+-ATPase beta-subunit with the Na+/K+-ATPase alpha-subunit.

Two sets of chimeric beta-subunits were constructed from subunits of Torpedo californica Na+/K+-ATPase and pig gastric H+/K+-ATPase. Five unique restriction sites (SnaBI, EcoRV, MunI, SphI and EcoT22I) were created at equivalent positions of the respective cDNAs and were used as joining points for the construction. One set of chimeras (HxN series) was made by exchanging the 5' portion of the Na+/K+-ATPase beta-subunit cDNA with the corresponding portion of the H+/K+-ATPase beta-subunit cDNA at the respective joining point. Complementary constructs were also prepared (NxH series). In the HxN series, the chimera joined at the SnaBI site formed a stable trypsin resistant complex with the Na+/K+-ATPase alpha-subunit, which was functional with respect to ATP hydrolysis and pump current generation, although the activities were less than those of the complex with the Na+/K+-ATPase beta-subunit. Trypsin resistance decreased for the complex of the chimera joined at the EcoRV site. In the NxH series, the chimeras joined at the SnaBI site and the EcoRV site formed rather trypsin-resistant complexes, but the expressions of the alpha-subunits were below 50% of the control. The chimeras joined at the MunI, SphI and EcoT22I site formed complexes susceptible to tryptic digestion. None of the chimeras in the NxH series were functional. These results suggest that at least two regions of the Na+/K+-ATPase beta-subunit [SnaBI site(Tyr40) to EcoRV site(Ile89) and EcoT22I site(Cys176) to C-terminus)] are involved in stable assembly with the Na+/K+-ATPase alpha-subunit and that the cytoplasmic domain [N-terminus to SnaBI site(Tyr40)] is functionally replaceable with the corresponding domain of the H+/K+-ATPase beta-subunit.

Amino Acid Sequence↗
PubMed1997

Differential splicing of the GTP cyclohydrolase I RNA in dopa-responsive dystonia.

We characterized the GTP cyclohydrolase I (GTP-CH-I) gene in a patient with hereditary progressive dystonia with marked diurnal fluctuation/dopa-responsive dystonia (HPD/DRD). The sequence analysis revealed a C to A transversion, which predicts a novel missense mutation (Thr186Lys). Unexpectedly, this base change, occurring in the middle of exon 5, resulted in a production of the novel transcript lacking exon 5 and a part of exon 6. Three different transcripts of the GTP-CH-I gene, previously reported in the human liver, were also present in the peripheral lymphocytes from the patient and controls. Quantitative comparison of the truncated-subunit mRNA and the wild-type one implied that differential splicing regulates the GTP-CH-I enzyme activity, leading to the clinical variations in HPD/DRD. The patient showed a unique clinical symptom, suggesting that the nigrostriatal dopaminergic system is more affected than previously thought in HPD/DRD.

Adult↗
PubMed1997

Bicuculline and gabazine are allosteric inhibitors of channel opening of the GABAA receptor.

Anesthetic drugs are known to interact with GABAA receptors, both to potentiate the effects of low concentrations of GABA and to directly gate open the ion channel in the absence of GABA; however, the site(s) involved in direct gating by these drugs is not known. We have studied the ability of alphaxalone (an anesthetic steroid) and pentobarbital (an anesthetic barbiturate) to directly activate recombinant GABAA receptors containing the alpha 1, beta 2, and gamma 2L subunits. Steroid gating was not affected when either of two mutated beta 2 subunits [beta 2 (Y157S) and beta 2 (Y205S)] are incorporated into the receptors, although these subunits greatly reduce the affinity of GABA binding. These observations indicate that steroid binding and subsequent channel gating do not require these particular residues, as already shown for barbiturates. Bicuculline or gabazine (two competitive antagonists of GABA binding) reduced the currents elicited by alphaxalone and pentobarbital from wild-type GABAA receptors; however, gabazine produced only a partial block of response pentobarbital or alphaxalone, and bicuculline only partially blocked responses to pentobarbital. These observations indicate that the blockers do not compete with alphaxalone or pentobarbital for a single class of sites on the GABAA receptor. Finally, at receptors containing alpha 1 beta 2 (Y157S) gamma 2L subunits, both bicuculline and gabazine showed weak agonist activity and actually potentiated responses to alphaxalone. These observations indicate that the blocking drugs can produce allosteric changes in GABAA receptors, at least those containing this mutated beta 2 subunit. We conclude that the sites for binding steroids and barbiturates do not overlap with the GABA-binding site. Furthermore, neither gabazine nor bicuculline competes for binding at the steroid or barbiturate sites. The data are consistent with a model in which both gabazine and bicuculline act as allosteric inhibitors of channel opening for the GABAA receptor after binding to the GABA-binding site.

Allosteric Regulation↗
PubMed1997

Potent protective effect of melatonin on chromium(VI)-induced DNA single-strand breaks, cytotoxicity, and lipid peroxidation in primary cultures of rat hepatocytes.

Incubation of primary cultures of rat hepatocytes with K2Cr2O7 plus the pineal hormone melatonin resulted in a marked decrease in cellular levels of DNA single-strand breaks caused by K2Cr2O7. Cellular treatment with melatonin also suppressed both dichromate-induced cytotoxicity, as evaluated by the leakage of lactate dehydrogenase, and lipid peroxidation, as monitored by malondialdehyde formation. In addition, treatment with melatonin attenuated the suppression of the levels of vitamins E and C as well as the inhibition of catalase activity attributed to K2Cr2O7. However, melatonin had no influence on cellular level of glutathione and the activity of glutathione reductase, glutathione peroxidase, superoxide dismutase, and alkaline phosphatase suppressed by dichromate. Under the same experimental conditions, cellular uptake and distribution of Cr were not affected by melatonin. Electron spin resonance (ESR) studies showed that melatonin did not affect the formation of Cr(V) complexes in the reaction of K2Cr2O7 with reduced glutathione; however, melatonin caused a 25% decrease in the levels of Cr(V)-related hydroxyl radicals in vitro. These results indicate that melatonin protects cells from Cr(VI)-induced DNA strand breaks, cytotoxicity, and lipid peroxidation, possibly through its ability to increase cellular levels of vitamins E and C as well as catalase activity and/or to directly scavenge toxic hydroxyl radicals in cells.

Animals↗
PubMed1997

Protective effect of deferoxamine on chromium (VI)-induced DNA single-strand breaks, cytotoxicity, and lipid peroxidation in primary cultures of rat hepatocytes.

Incubation of primary cultures of rat hepatocytes with K2CR2O7 and deferoxamine (DFO), an iron chelator, resulted in a marked decrease in cellular levels of DNA single-strand breaks caused by K2Cr2O7. Cellular treatment with DFO also suppressed both dichromate-induced cytotoxicity--evaluated by the leakage of lactate dehydrogenase, and lipid peroxidation--as monitored by malondialdehyde formation. In addition, treatment with DFO attenuated the suppression of the levels of vitamin E and C as well as the inhibition of alkaline phosphatase and glutathione peroxidase activity attributed to K2Cr2O7. However, DFO had no influence on the cellular level of glutathione or the activity of glutathione reductase and superoxide dismutase suppressed by dichromate. Under the same experimental conditions, cellular uptake and distribution of chromium were not affected by DFO. These results indicate that DFO protects cells from chromium (VI)-induced DNA strand breaks, cytotoxicity, lipid peroxidation, vitamin E and C depression, and glutathione peroxidase inhibition The role of antioxidants in chromium (VI)-induced cytotoxicity, DNA breaks, and lipid peroxidation is discussed.

Alkaline Phosphatase↗
PubMed1997

Effect of SKF-525A on liver metabolism and hepatotoxicity of tri- and dibutyltin compounds in mice.

The effect of pretreatment with SKF-525A, which inhibits hepatic cytochrome P450 enzymes, on metabolism and hepatotoxicity was examined in mice orally administered tributyltin chloride (TBTC) or dibutyltin dichloride (DBTC) at a dose of 180 mumol/kg. Analysis of butyltin compounds showed that the main metabolites in liver of mice treated with TBTC alone were DBTC (40%) and dibutyl(3-carboxylpropyl)tin chloride (TCOOH; 12-26%), with the levels of other butyltin compounds including TBTC comprising < 12% of total butyltin compounds at 3-24 h following treatment. The pretreatment with SKF-525A resulted in four- to tenfold increased TBTC levels and a significant decrease of debutylated metabolites, particularly DBTC (60 and 37% decrease) at both 3 and 6 h in liver of mice treated with TBTC, leading to complete inhibition of hepatotoxicity at 24 h. At 24 h after TBTC treatment, hepatic levels of TBTC and most of the debutylated metabolites in mice pretreated with SKF-525A did not differ significantly when compared to those in unpretreated mice, resulting in the induction of hepatotoxicity at 48 h, although levels of TCOOH decreased even at 24 h. In the case of DBTC treatment, > 95% of the butyltin compounds were detected as DBTC in liver, and the levels of DBTC inside cells as well as the induction of DBTC hepatotoxicity were unaffected by pretreatment with SKF-525A. These results suggest that debutylated metabolites, in particular DBTC, are the main metabolites of butyltin compounds responsible for the induction of hepatotoxicity following in vivo administration of TBTC. The results also indicate that cytochrome P450 enzymes may play a greater role in the metabolism of TBTC to form DBTC or butyltin trichloride (MBTC) than that of DBTC to form MBTC in liver of mice.

Animals↗
PubMed1997

Immunocytochemical localization of nicotinic acetylcholine receptor in the rat cerebellar cortex.

Although nicotinic acetylcholine receptors (nAChRs) have been reported to function in the cerebellar cortex, nicotinic acetylcholine receptor subtypes contributing to nicotinic currents in the cortex have not been well characterized at the subcellular level. This study deals with immunocytochemical localization of the nicotinic acetylcholine receptor alpha4 subunit using a monoclonal antibody against the alpha4 subunit. Alpha4-LI (alpha4-like immunoreactivity) was detected in the cell bodies of molecular, Purkinje cell and granular layers. In particular, the cell bodies of Purkinje cells were extensively immunostained. In Purkinje cells, alpha4-LI was found in perikarya mainly associated with rough endoplasmic reticulum, plasma membrane, and cytoplasmic matrix. At higher magnification, the immunoreaction product was densely localized along with somatic plasma membranes at the axo-somatic synapse and the plasma membranes at extrasynaptic regions of cell bodies. Alpha4-LI was also found in the axon terminals which form synapses with Purkinje cells. In the granular layer, somatic cell membranes of granular cells were immunostained. These morphological observations suggest that alpha4-containing nAChRs contribute nicotinic currents reported in Purkinje cells, and that presynaptic alpha4-containing nAChRs regulate the release of neurotransmitters on the axon terminals found near Purkinje cells.

Animals↗
PubMed1997

Importance of soft tissue inhomogeneity in magnetic peripheral nerve stimulation.

In magnetic peripheral nerve stimulation with a figure-of-eight coil, a 'tangential-edge' coil orientation (the nerve is beneath the coil intersection and perpendicular to the coil wings) is ideal theoretically. However, some experimental results show that strong muscle responses are elicited with a 'symmetrical-tangential' coil orientation (the nerve is beneath the coil intersection and parallel to the coil wings), which is inconsistent with the cable theory. We hypothesized that the 10:1 conductivity difference between muscle and fat would cause inconsistent results during magnetic median nerve stimulation in the elbow, which was verified using an inhomogeneous volume conductor model. The induced electric fields were measured in a model composed of saline solutions of different concentrations divided by a cellophane sheet. A nerve was imagined along the boundary between the two solutions, and the coil was held in a 'symmetrical-tangential' position. Virtual cathodes, which were off the nerve in the homogeneous model, were on the nerve in the inhomogeneous model. The previous inconsistent results were explained by considering soft tissue inhomogeneity without any modification of the assumption in the cable theory that only the induced electric field component parallel to the nerve is responsible for nerve excitation.

Adult↗
PubMed1997

Significance of the beta-subunit in the biogenesis of Na+/K(+)-ATPase.

This review summarizes our experiments on the significance of the beta-subunit in the functional expression of Na+/K(+)-ATPase. The beta-subunit acts like a receptor for the alpha-subunit in the biogenesis of Na+/K(+)-ATPase and facilitates the correct folding of the alpha-subunit in the membrane. The alpha-subunit synthesized in the absence of the beta-subunit is subjected to rapid degradation in the endoplasmic reticulum. Several assembly sites are assigned in the sequence of the beta-subunit from the cytoplasmic NH2-terminal domain to the extracellular COOH-terminus: the NH2-terminal region of the extracellular domain, the conservative proline in the third disulfide loop, the hydrophobic amino acid residues near the COOH-terminus and the cysteine residues forming the second and the third disulfide bridges. Upon assembly, the beta-subunit confers a resistance to trypsin on the alpha-subunit. The conformations induced in the alpha-subunit of Na+/K(+)-ATPase by Na+/K(+)- and H+/K(+)-ATPase beta-subunits are somehow different from each other and are named the NK-type and KH-type, respectively. The extracellular domain of the beta-subunit is involved in the folding of the alpha-subunit leading to trypsin-resistant conformations. The sequences from Cys150 to the COOH-terminus of the Na+/K(+)-ATPase beta-subunit and from Ile89 to the COOH-terminus of the H+/K(+)-ATPase beta-subunit are necessary to form trypsin-resistant conformations of the NK- and HK-type, respectively. The first disulfide loop of the extracellular domain of the beta-subunits is critical in the expression of functional Na+/K(+)-ATPase.

Amino Acid Sequence↗
PubMed1997

Detached retina affects morphologic and biochemical changes in the retina adjacent to bullous retinal detachment in rabbits.

PURPOSE: Long-term results, more than 10 years after successful retinal detachment surgery, have shown gradually decreasing visual acuity in some cases. It is unclear if reduced functional recovery postoperatively is caused by anatomic changes or biochemical disorders. To determine the etiology of the reduced visual acuity, we cytochemically examined the changes in the cellular responses of the edges of retinal detachments. METHODS: We histochemically studied the glucose-6-phosphatase (G6P) and 5'-nucleotidase (5'-Nase) activity in the rabbit retina. Experimental rhegmatogenous retinal detachment was produced in a rabbit model after partial vitrectomy, followed by retinal tear formation. RESULTS: Although 5'-Nase activity gradually decreased during the period of detachment, activity was still detectable after 24 weeks. G6P activity increased in the region of the detached neural retina. Around the border of the detached retina, the decrease in 5'-Nase activity extended approximately 140 micrometers into the adjacent attached retina at 2 weeks after detachment and 270 micrometers at 24 weeks. CONCLUSIONS: These observations suggest that some anatomical and biochemical damages may occur in the retina adjacent to bullous retinal detachment and may explain the reduction in postoperative vision in some clinical cases.

5'-Nucleotidase↗
PubMed1997

Hepatitis C virus-related liver damage is related to cold activation of complement.

A high positivity of cold activation of complement has been reported in Japanese patients having hepatitis B virus-negative chronic hepatitis. Although the cause of cold activation of complement is unknown, the involvement of hepatitis C virus (HCV) has been suspected. We studied the positivity of cold activation of complement in 253 patients, including 93 patients with chronic hepatitis C infection who received 6MU natural interferon-alpha per day for 24 continuous weeks. Cold activation was positive in 38% of patients with chronic hepatitis C and in 46% of patients with liver cirrhosis C. We did not detect cold activation in asymptomatic HCV carriers; patients with chronic hepatitis B, liver cirrhosis B, or alcohol-related liver damage; or in the controls. Cold activation was also negative in HCV-RNA-negative patients who responded completely to interferon-alpha, and in HCV-RNA-positive patients who responded partially whose serum alanine transaminase levels were normalized after interferon treatment. In the patients who had a relapse of hepatitis C after interferon treatment, positivity of cold activation increased sharply. We conclude that HCV-associated liver damage is related to the development of cold activation of complement. Cold activation is useful for monitoring the response to interferon in patients with chronic hepatitis C infection.

Adult↗
PubMed1997

Magnetically levitated motor for rotary blood pumps.

The noncontact rotary pumps under development for use as artificial heart pumps are highly efficient and can prevent thrombus formation. In these pumps magnetic bearings have been widely used to support the rotors to avoid any physical contact. The use of magnetic bearings, however, has led to requirements for the control of a large degree of freedom and for a separate driving motor. This paper introduces 2 types of levitated motors, each of which uses a combination of a rotary motor and a magnetic bearing. These motors are suitable for use in artificial blood pumps because they are small in size and can replace contact components. The radial type levitated motor has the merit of being small in size and capable of controlling the 2 degrees of freedom in the x and y directions. The axial type motor controls only one degree of freedom in the z direction. This paper also introduces the theoretical background of the functions of the motor and magnetic bearing. Experimental results of tests of the proposed motor show a great potential for its application in rotary blood pumps.

Algorithms↗
PubMed1997

alpha-Tocopherol transfer protein gene: exon skipping of all transcripts causes ataxia.

This report concerns the characterization of the alpha-tocopherol transfer protein (alpha-TTP) gene in a Japanese family affected by ataxia with isolated vitamin E deficiency (AVED). The sequence analysis revealed a G-to-A transition at the 3' end of exon 3 in both alleles, which predicts outsplicing of this exon from premessenger RNA and the concomitant frame shift in the ataxic patient. We used reverse transcriptase-polymerase chain reaction to analyze alpha-TTP gene transcripts. All transcripts in peripheral blood lymphocytes of the AVED patient, who was treated with large doses of vitamin E, lacked exon 3. The deduced truncated protein shares only 43% of the normal alpha-TTP. Normal control tissues and cells contained normal transcripts and, unexpectedly, also the same mutant transcripts as those of the patient, although with different transcription levels. Treatment of normal fibroblasts with clinically relevant concentrations of vitamin E increased production of transcripts in a dose-dependent manner. We propose that exon skipping of all transcripts through the complete inactivation of the splice site accounts for the clinical onset of AVED and for the clinical resistance to vitamin E in our patient.

Adult↗
PubMed1997

Twenty-four-hour profile of growth hormone in cyclic nocturnal total parenteral nutrition.

OBJECTIVE: To detect the effect of the loss of alimentation rhythmicity on a circadian rhythm of human growth hormone (HGH) secretion, a 24-hour profile of HGH was studied in a growing child on cyclic nocturnal total parenteral nutrition (TPN). Twenty-four-hour profiles of substrates and metabolic hormones were also studied to evaluate the efficiency of cyclic nocturnal TPN on childhood growth. STUDY DESIGN: Periodic blood samples from a child with megacystis-microcolon-intestinal-hypoperistalsis syndrome were obtained on five occasions, at ages 6, 7, 8, 9, and 11, when she was on cyclic nocturnal TPN. RESULTS: Peak HGH secretion appeared with the onset of deep sleep despite the concomitant hyperglycemia and hyperinsulinemia induced by TPN at night. Smaller peaks of HGH were also observed during the noninfusion period during the day. Twenty-four-hour profiles of substrates and metabolic hormones indicated a switch from glucose use during the infusion phase to an oxidation of lipids during the noninfusion period. CONCLUSION: The fact that the patient's growth curve remains within normal limits suggests that cyclic nocturnal TPN would be an efficient method of nutritional support. During cyclic nocturnal TPN, regular rhythm of HGH secretion occurs, and normal childhood growth is achieved.

Blood Glucose↗
PubMed1997

[Hereditary progressive dystonia with marked diurnal fluctuation--clinical features and GTP cyclohydrolase I gene mutations].

Hereditary progressive dystonia with marked diurnal fluctuation (HPD) is a disorder characterized by childhood-onset dystonia and a dramatic and sustained response to low doses of levodopa. Recently the GTP cyclohydrolase I(GCH-I) gene was isolated as the first causative gene for HPD. We analyzed the GCH-I gene in 8 clinically diagnosed HPD patients and found different point mutations in GCH-I gene in 3 subjects. The clinical features of these patients considerably resembled each other. Our results imply that although clinically diagnosed HPD subjects could present diverse symptoms, patients with a mutant GCH-I gene might share homogeneous clinical manifestations.

Adolescent↗