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Biomedical subjects

A Hirata

Publications and source records attributed to A Hirata.

At least 91 records · Page 5Linked to original sources
PubMed2000

[Statistical insight to improve reliability of pharmacological studies].

When pharmaceutical scientists describe characteristics of a drug or when they decide whether it is appropriate to initiate clinical trials to determine the drug's effects in humans, their inferences are frequently grounded in information drawn from non-clinical studies. Therefore, certain and highly objective information is required. By introducing the concept of design of experiments to control some nuisance factors and performing confirmatory studies based on sample size estimation, trustworthy information can be efficiently obtained. This paper does not demand that researchers conduct an additional confirmatory study in a series of studies conducted so far. This is a reconsideration how a series of studies should be carried forward. Statistics ought to contribute much more not only to estimation or hypothetical tests after data are collected, but also to methodology of preliminary experiments and planning of studies. Cooperation with statisticians from an early stage of the studies is all the more helpful in non-clinical studies, in which, in a sense, "perfect" experiments can be conducted more than in clinical studies.

Animals↗
PubMed2000

A novel mutation of the ceruloplasmin gene in a patient with heteroallelic ceruloplasmin gene mutation (HypoCPGM).

We found a novel missense mutation in the ceruloplasmin (Cp) gene in a patient with the heteroallelic Cp gene mutation (HypoCPGM). The patient was a 72-year-old woman who came to our hospital with a 1-year history of postural tremor of the hands. The diagnosis was made based on serum Cp and copper readings which were about half the normal levels, as well as MRI tests of her brain which showed characteristics for hereditary ceruloplasmin deficiency (HCD), known to be caused by the homoallelic Cp gene mutation. Polymerase chain reaction (PCR)-direct sequencing analysis of the Cp gene of the patient revealed a novel point mutation, A to T, at nucleotide position 82 in Exon 1. This mutation changes the Ile28 codon (ATT) to a Phe codon (TTT) (missense mutation). PCR-restriction analysis with restriction enzyme Tsp EI for the mutation revealed that both the patient and her son were heterozygotes for the mutation.

Aged↗
PubMed2000

[Respiratory muscle weakness after prolonged use of hydrocortisone and pancuronium bromide].

A 30-year-old man was admitted because of status asthmaticus. He required 7 days of artificial ventilation and was treated with hydrocortisone 1.2 g.day-1 and bronchodilaters. Pancuronium bromide 0.08 mg.kg-1.hr-1 was given for 64 hours for the ease of artificial ventilation. On day 3, severely elevated airway pressure resulted in left pneumothorax and isoflurane 1% was given for the following 2 days. Respiratory muscle weakness was evident 24 hours after discontinuation of pancuronium infusion on day 5 while full 4 twitches of TOF on the adductor pollicis muscle were seen at the time. The respiratory muscle weakness continued for another 3 days and he was extubated on day 8. Serum creatine kinase concentration rose to 2178 U.l-1 on day 6 and returned to normal on day 11. Hematurea, hyperpyrexia and metabolic acidosis were never seen during the course. Acute corticosteroid myopathy was suspected to be the cause of the prolonged respiratory muscle weakness.

Acute Disease↗
PubMed2000

Retinal damage by air infusion during vitrectomy in rabbit eyes.

PURPOSE: Visual field defects after vitrectomy can be seen after any surgery involving fluid-air exchange. To elucidate the effect during surgery of the infused air on the retina, the present study investigated the changes in the morphology of the rabbit retina induced by air infusion and the changes resulting from varying amounts of infused air pressure. METHODS: Eighteen eyes of 18 rabbits were used. A standard three-port vitrectomy with artificial posterior vitreous detachment followed by fluid-air exchange was performed in 12 eyes. During the fluid-air exchange, humidified air was infused with an air pressure of 25 or 40 mm Hg for 30 seconds. As a control, vitrectomy without fluid-air exchange was performed in six eyes. The eyes were enucleated and fixed immediately. Specimens were processed and examined by light and scanning electron microscopy (SEM). RESULTS: With SEM, sharply demarcated retinal lesions were observed at the opposite side from the infusion cannula in all eyes in which a fluid-air exchange was performed. At the lesion, the internal limiting membrane was often detached, and the underlying nerve fiber layer was exposed. Light microscopy revealed that the inner retina was most affected, with concomitant swelling of the inner plexiform layer and the inner granular layer. In addition, the retina was often focally detached with adhesion of some retinal pigment epithelial cells to the photoreceptor cells. Increased infused air pressure was accompanied by a significant increase in the area of retinal damage. In contrast, no morphologic change was observed in the control eyes. CONCLUSIONS: Air infusion during vitrectomy can cause mechanical retinal damage in the rabbit retina. The mechanical damage may result in a visual field defect after vitrectomy.

Air↗
PubMed2000

Levels of interleukin 12, a cytokine of type 1 helper T cells, are elevated in sera from patients with systemic sclerosis.

OBJECTIVE: To determine serum levels and spontaneous production by peripheral blood mononuclear cells (PBMC) of interleukin 12 (IL-12), a potent inducer of type 1 helper (Th1) T cells, in patients with systemic sclerosis (SSc). METHODS: Serum IL-12 levels and spontaneous production levels of IL-12 in culture supernatants from PBMC were examined by ELISA. Serum levels of IL-4, IL-6. IL-10, and IL-13 and production levels by PBMC of IL-6 and IL-10 were also examined by ELISA. Renal vascular damage was determined as a pulsatility index (PI) by color flow Doppler ultrasonography of kidneys. RESULTS: Serum IL-12 levels were significantly elevated in patients with SSc (n = 62) compared with healthy controls (n = 20). Similarly, spontaneous production levels of IL-12 by PBMC in patients with SSc (n = 47) were higher than those in controls (n = 20). Serum IL-12 levels did not correlate with serum levels of any Th2-type cytokines such as IL-4, IL-6, IL-10, and IL-13. However, spontaneous production levels of IL-10 by PBMC significantly correlated with serum IL-12 levels in patients. Patients with elevated serum IL-12 levels had the increased PI values more frequently than those with normal IL-12 levels. Further, serum levels of IL-12 and production levels of IL-12 by PBMC correlated significantly with the PI values in patients with SSc. CONCLUSION: These results suggest that the increased levels of IL-12 may relate to the activation of Th1 cells in SSc and that IL- 12 overproduction may be associated with renal vascular damage.

Adolescent↗
PubMed2000

[Reduced signal intensity of T2 weighted MR imaging of thalamus and putamen in multiple sclerosis in Japan].

Although studies using magnetic resonance imaging (MRI) in multiple sclerosis (MS) patients have focused on findings in the white matter because of its demyelination pathogenesis, Drayer et al. have reported a high incidence of low signal intensity on T2 weighted MR imaging (MRI) in gray matter such as the thalamus and putamen. In Japan there has been no investigation of MRI findings of the basal ganglia in MS patients. Therefore, we attempted to examine the incidence and clinical significance of the imaging phenomenon in 34 Japanese patients with MS (12 male, 22 female, ages 18-54 years). As it is well known that the spinal cord and optic nerves are more frequently involved in MS than the brain in Japanese patients, we divided the patients into two subgroups based on their clinical features and the major sites of demyelination on MRI. One group included the 17 patients whose demyelinations occurred in the brain (brain-type), and the other group included the 17 patients whose abnormalities were found in the spinal cord with or without optic nerve involvement (non-brain type). As a control, MRI studies were also performed in age-matched patients with headache without any neurological signs. On T2 weighted MRI, decreased signal intensity in the thalamus was found in only four patients with MS, 11.8% of the total number examined, and in the putamen in three patients with MS, 8.8% of the total examined. All of the patients who showed abnormal decreased signal intensity in the thalamus and/or putamen belonged to the brain-type group, and these incidences were 23.5% in the thalamus and 17.6% in the putamen among the brain-type patients. No patient belonging to the non-brain type showed this imaging sign. This imaging sign was well correlated with the degree of white matter abnormalities in the brain estimated as a score according to modified Callanan et al.'s method. In addition, this sign was also correlated with the expanded disability status scales (EDSS) in the brain-type patients. These observations suggest that the axonal damages due to severe demyelination may induce the impaired transport of iron resulting in an accumulation of ferritin in the thalamus and putamen, and would cause decreased signal intensity on T2 weighted MRI. The relatively low incidence of decreased signal intensity in the thalamus and putamen in this study may be associated with differences in the clinical phenotype of MS between Japan and the USA. In brain-type patients the evaluation of basal ganglia on T2 weighted MRI may be a useful tool for estimating patients' disabilities.

Adolescent↗
PubMed2000

Microbial ecology of nitrifying bacteria in wastewater treatment process examined by fluorescence in situ hybridization.

The microbial ecology of nitrifying bacteria in various types of wastewater treatment processes and the dynamic response of the microbial ecology in biofilms were investigated using fluorescence in situ hybridization (FISH) with 16S rRNA-targeted oligonucleotide probes. Nitrifying bacteria were found to exhibit various organizational forms under different conditions of substrate composition and concentration. Ammonia-oxidizing bacteria were dominant in ammonia-rich inorganic wastewater, while heterotrophic bacteria and ammonia-oxidizing bacteria were localized at different positions in the biofilm in organic wastewater. The dynamics of the microbial ecology in the biofilm with regard to the spatial distribution of ammonia-oxidizing bacteria and heterotrophic bacteria caused by a gradual change in substrate composition was successfully monitored by FISH analysis.

Journal Article↗
PubMed1999

Expression of E16/CD98LC/hLAT1 is responsive to 2,3,7,8-tetrachlorodibenzo-p-dioxin.

We employed cDNA representational difference analysis to identify new genes that are upregulated by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in a human hepatoblastoma cell line HepG2. We isolated several TCDD-responsive cDNAs. Sequence analysis revealed that one of them encodes E16/CD98LC/hLAT1, an integral membrane protein involved in multiple cellular functions including cellular transport of L-type amino acids. Northern blot analysis confirmed the TCDD-dependent upregulation of the mRNA. Induction of E16/CD98LC/hLAT1 mRNA by TCDD did not require de novo protein synthesis as revealed by the experiment using cycloheximide. Consistent with the changes at mRNA level, the transport of 3H-leucine into HepG2 cells was significantly increased by TCDD treatment. These findings provide a novel aspect of biological effects of TCDD on human hepatocytes.

Amino Acid Transport Systems↗
PubMed1999

Patch clamp studies on V-type ATPase of vacuolar membrane of haploid Saccharomyces cerevisiae. Preparation and utilization of a giant cell containing a giant vacuole.

A method for obtaining giant protoplasts of Escherichia coli (the spheroplast incubation (SI) method: Kuroda et al. (Kuroda, T., Okuda, N., Saitoh, N., Hiyama, T., Terasaki, Y., Anazawa, H., Hirata, A., Mogi, T., Kusaka, I., Tsuchiya, T., and Yabe, I. (1998) J. Biol. Chem. 273, 16897-16904) was adapted to haploid cells of Saccharomyces cerevisiae. The yeast cell grew to become as large as 20 micrometer in diameter and to contain an oversized vacuole inside. A patch clamp technique in the whole cell/vacuole recording mode was applied for the vacuole isolated by osmotic shock. At zero membrane potential, ATP induced a strong current (as high as 100 pA; specific activity, 0.1 pA/micrometer(2)) toward the inside of the vacuole. Bafilomycin A(1,) a specific inhibitor of the V-type ATPase, strongly inhibited the activity (K(i) = 10 nM). Complete inhibition at higher concentrations indicated that any other ATP-driven transport systems were not expressed under the present incubation conditions. This current was not observed in the vacuoles prepared from a mutant that disrupted a catalytic subunit of the V-type ATPase (RH105(Deltavma1::TRP)). The K(m) value for the ATP dose response of the current was 159 microM and the H(+)/ATP ratio estimated from the reversible potential of the V-I curve was 3.5 +/- 0.3. These values agreed well with those previously estimated by measuring the V-type ATPase activity biochemically. This method can potentially be applied to any type of ion channel, ion pump, and ion transporter in S. cerevisiae, and can also be used to investigate gene functions in various organisms by using yeast cells as hosts for homologous and heterogeneous expression systems.

Anti-Bacterial Agents↗
PubMed1999

Yeast tom1 mutant exhibits pleiotropic defects in nuclear division, maintenance of nuclear structure and nucleocytoplasmic transport at high temperatures.

A tom1-1 mutant was isolated from Saccharomyces cerevisiae. At high temperatures, 60% of the cells were arrested as dumbbell forms with a single large nucleus containing duplicated DNA and a short spindle. Electron-microscopy showed electron-dense structures scattered within the nucleus. Indirect immunofluorescent microscopy revealed these structures to be fragmented nucleoli since the dotted structures were stained with anti-Nop1(fibrillarin) antibody in large regions of the nuclei. Fluorescent in situ hybridization analysis using oligo(dT) revealed nuclear accumulation of poly(A)+RNA. We cloned TOM1 which encodes a large protein (380kDa) with a hect (homologous to E6-AP C terminus)-domain at its C terminus. Deletions of either this hect-region or the entire gene made cellular growth temperature-sensitive. Site-directed mutagenesis of the conserved cysteine residue (tom1C3235A) in the hect-domain, supposed to be necessary for thioester-bond formation with ubiquitin, abolished the gene function. When a functional glutathione S-transferase (GST)-tagged hect protein was overproduced, it facilitated the protein conjugation with a myc-tagged ubiquitinRA, while this was not seen when GST-hectC3235A was overproduced. The protein conjugation with a hemagglutinin-tagged Smt3 was not affected by the overproduction of GST-hect. Taken together, we suggest that Tom1 is a ubiquitin ligase. As a multi-copy suppressor of tom1, we isolated STM3/NPI46/FPR3 which encodes a nucleolar nucleolin-like protein. We discuss possible functions of Tom1 with respect to the pleiotropic defects of nuclear division, maintenance of nuclear structure, and nucleocytoplasmic transport.

Amino Acid Sequence↗
PubMed1999

A novel mutation in the gene for the adult skeletal muscle sodium channel alpha-subunit (SCN4A) that causes paramyotonia congenita of von Eulenburg.

BACKGROUND: Paramyotonia congenita (PMC) of von Eulenburg is an autosomal dominant muscular disease characterized by exercise- and cold-induced myotonia and weakness. To date, 18 missense mutations in the adult skeletal muscle sodium channel alpha-subunit (SCN4A) gene have been identified to cause a spectrum of muscular diseases, including PMC of von Eulenburg, PMC without cold paralysis, potassium-aggravating myotonia, and hyperkalemic periodic paralysis. However, no obvious correlations can be made between the location or nature of amino acid substitutions in SCN4A and its clinical phenotypes. OBJECTIVE: To describe clinical and genetic features of a family with PMC of von Eulenburg. RESULTS: A Japanese family with cold-induced myotonia and weakness was diagnosed as having PMC of von Eulenburg. This phenotype was identified to be caused by a novel mutation that substituted a glutamic acid residue for a highly conserved glycine residue in the fourth transmembrane segment (S4) of domain IV. This predicted a decrease in positive charge specific for the S4. CONCLUSION: In addition to the G1456E identified in this study, 4 mutations that cause a decrease in positive charge in the S4/D4 are associated with the phenotype of PMC of von Eulenburg. This provides an important genotype-phenotype correlation in sodium channelopathies.

Adult↗
PubMed1999

Familial paroxysmal dystonic choreoathetosis: clinical findings in a large Japanese family and genetic linkage to 2q.

BACKGROUND: Paroxysmal dystonic choreoathetosis (PDC) is a rare familial movement disorder that has been mapped to chromosome 2q31-36. OBJECTIVE: To study the first Japanese family with PDC clinically and genetically. PATIENTS AND METHODS: We studied a large Japanese family in which at least 17 members in 6 generations have been affected by PDC. We interviewed and examined 26 family members, 8 of whom revealed choreoathetosis-like and dystonialike involuntary movement and 1 of whom revealed no involuntary movement but only muscle stiffness such as the aura of paroxysmal dystonic choreoathetosis (PDC). Genetic linkage studies of this family were carried out with polymorphic DNA markers. RESULTS: The attacks of involuntary movement or muscle stiffness were precipitated by ovulation, menstruation, emotional stress, or caffeine or alcohol ingestion. Magnetic resonance imaging of the brain revealed no abnormalities. Clonazepam therapy was effective for reducing the attacks, and ingestion of garlic was believed by patients to be effective for softening the attacks. An affected woman with only muscle stiffness showed remission after hysterectomy for hysteromyoma. This woman also had the disease haplotype and transferred it to her typical PDC-affected daughter. Maximal pairwise logarithm of odds scores exceeding 2.00 were obtained at D2S2250, D2S1242, D2S377, D2S2148, and D2S126. The PDC gene was demonstrated by linkage analyses to be located in a 15.3-centimorgan interval lying between D2S371 and D2S339 based on pairwise and multipoint logarithm of odds scores and obligate recombination events in affected individuals. CONCLUSIONS: Linkage of PDC to chromosome 2q32-36 was confirmed in a Japanese family. The clinical characterizations of this family with PDC include that ovulation seems also to be a precipitating factor of the attacks and that hysterectomy seems to be effective for softening the attacks. Although low-dose clonazepam treatment was most effective, garlic use was believed by affected members to be effective for softening the attacks. Furthermore, based on the results of clinical and genetic analyses, we suggest that muscle stiffness without involuntary movement may represent a forme fruste of PDC.

Athetosis↗
PubMed1999

Lipocortin (Annexin) I heterotetramer binds to purine RNA and pyrimidine DNA.

Lipocortin I-like protein with a molecular weight of 94,000 Da as judged by Western analysis was found to bind to ssDNA rather than to dsDNA in a Ca(2+)-dependent manner. This protein was also bound to [(32)P]poly(rA) and [(32)P]poly(rG) as measured by EMSA. Poly(rG), poly(rA), poly(dC), and poly(dT) were competitive against binding of either [(32)P]poly(rA) or [(32)P]poly(rG), while poly(rC), poly(rU), and poly(dA) were less effective binding competitors. The binding of this protein to poly(rA) or poly(rG) was inhibited by immunoprecipitable anti-lipocortin I (calpactin II) and anti-S100 protein antibodies, but not by an anti-Ig antibody. Phospholipids such as phosphatidylserine and phosphatidylinositol enhanced the binding of lipocortin I to poly(rA). Taken together, our present observations suggest that the lipocortin I-S100 protein heterotetramer binds to either purine RNAs or pyrimidine ssDNAs in a Ca(2+)- and phospholipid-dependent manner.

Animals↗
PubMed1999

Acute intermittent porphyria with central pontine myelinolysis and cortical laminar necrosis.

Acute intermittent porphyria (AIP) is an autosomal-dominant disease caused by a deficiency of porphobilinogen (PBG) deaminase. Patients with AIP present with neurological syndromes such as autonomic neuropathy, peripheral axonal neuropathy or central nervous system dysfunction. We report serial MRI of a patient with AIP who had cortical and subcortical cerebral changes. A 29-year-old woman with a 6-month history of AIP had an attack with severe hyponatraemia and generalised convulsions, treated with haem arginate and supportive therapy. MRI showed central pontine and extrapontine myelinolysis and cortical laminar necrosis. These are not common in AIP, but are likely to have been caused by rapid correction of hyponatraemia and by vasospasm, which could be induced by AIP.

Adult↗
PubMed1999

Unfolded protein response-induced BiP/Kar2p production protects cell growth against accumulation of misfolded protein aggregates in the yeast endoplasmic reticulum.

Overproduction of delta(pro), a mutated secretory proteinase derived from a filamentous fungus Rhizopus niveus, results in formation of gross aggregates (delta(pro) aggregates) in the yeast endoplasmic reticulum (ER) lumen, activation of the unfolded protein response (UPR) and ER membrane proliferation. To investigate the roles of the UPR against the delta(pro) aggregates, we constructed an IRE1-deleted ((delta)ire1) strain. In contrast to wild-type cells, (delta)ire1 cells ceased to grow several hours after the overproduction of (delta)pro. Two lines of evidence argued against the possibility that the growth defect was due to the inability to make extra ER membrane which accommodates the (delta)pro aggregates. First, by electron microscopy, ER membrane proliferation was observed in (delta)ire1 cells overproducing (delta)pro. Second, disruption of the OPI1 gene in the (delta)ire1 mutant, which is considered to derepress the activities of phospholipid-synthesizing enzymes, did not restore the growth upon the overproduction of (delta)pro. Instead, the growth was restored when an extra copy of the KAR2 gene, which encodes yeast BiP, was introduced, indicating that an increase in the amount of BiP is essential for cell growth when the (delta)pro aggregates accumulate in the ER. Since BiP is included in the insoluble (delta)pro aggregates, it is likely that the amount of free BiP in the ER lumen is insufficient without the UPR to fully exert its functions. Consistently, overproduction of (delta)pro impaired protein translocation and folding in (delta)ire1 cells but not in wild-type cells. The tunicamycin sensitivity of (delta)ire1 cells was also suppressed by extra expression of KAR2, suggesting that BiP plays a principal role in protecting cell growth against misfolded proteins accumulated in the ER.

Base Sequence↗
PubMed1999

Poly(ethylene glycol)-alpha-chymotrypsin complex catalytically active in anhydrous isooctane.

Enzymatic catalysis in predominantly organic media has undergone rapid expansions, particularly over the past decade. There are numerous potential advantages in employing enzymes in organic media. However, there are some crucial defects in the native enzyme-catalyzed biocatalyses, such as a decrease in their reaction rate due to diffusional limitations of substrates. To overcome these defects, enzymes modified chemically with polymers and physically with surfactants, which were soluble in organic solvents, were often used. However, they had inherent drawbacks. On the other hand, the enzymes modified physically with polymers were found to be soluble and catalytically active in organic media. To date, however, the effect of the amount of added polymers on the enzyme activity has not been clarified, even though this may be an important factor governing the activity of polymer-enzyme complexes in organic media. In this study, we obtained a complex of an enzyme and a hydrophilic polymer by lyophilizing an aqueous solution containing a polymer and an enzyme. We found that the polymer-enzyme complex was catalytically active in organic media even when the molar ratio of polymer/enzyme in its preparation stage was unity, and that the activity of the polymer-enzyme complex increased with an increase in the molar ratio of polymer/enzyme, reached the maximum activity (ca. 7200-fold higher than that of the native enzyme suspended in organic media) and then gradually decreased.

Journal Article↗
PubMed1999

Null mutation in IRE1 gene inhibits overproduction of microsomal cytochrome P450Alk1 (CYP 52A3) and proliferation of the endoplasmic reticulum in Saccharomyces cerevisiae.

Overproduction of microsomal cytochrome P450Alk1 (P450Alk1) of Candida maltosa in Saccharomyces cerevisiae resulted in an extensive proliferation of endoplasmic reticulum (ER) and induction of Kar2p and Pdi1p. The ire1 null mutation severely suppressed ER proliferation, reduced the level of functional P450Alk1, and showed no induction of these ER chaperones, suggesting that the function of Ire1p is required for ER proliferation upon the overproduction of P450Alk1. Cerulenin, a potent inhibitor of lipid biosynthesis, also induced these chaperones in an Ire1p-dependent manner and limited the production of functional P450Alk1. These results imply that Ire1p may function to restore the balance between membrane proteins and lipids of the ER when the ER is relatively overcrowded by membrane proteins.

Cytochrome P-450 Enzyme System↗