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Biomedical subjects

J Fujii

Publications and source records attributed to J Fujii.

At least 55 records · Page 3Linked to original sources
PubMed1999

Functional expression of rat thioredoxin reductase: selenocysteine insertion sequence element is essential for the active enzyme.

Mammalian thioredoxin reductase (TR) is a flavoprotein catalysing reduction of oxidized thioredoxin in an NADPH-dependent manner, and contains a selenocysteine (Sec) residue near the C-terminus. We observed that TR activity was decreased in A549 cells by the lowering of the fetal bovine serum content in the culture medium and was recovered by the addition of selenium. To study the role of Sec in TR activity, we have isolated a full-length clone of the rat TR cDNA (3.3 kb) and have expressed it in COS-1 cells in a transient-expression system. TR activities in COS-1 cells expressing rat TR were increased in accordance with supplemented sodium selenite concentrations, whereas levels of TR protein, examined by Western blotting, were not affected by sodium selenite concentrations. We introduced various deletions into the 3'-untranslated region of the TR cDNA to localize and examine the role of a Sec insertion-sequence (SECIS) element in the functional expression of TR. TR activities were observed only in COS-1 cells transfected with the TR cDNAs containing the putative SECIS element located between 1856 and 1915 bp in the correct orientation. We also carried out radiolabelling of proteins by incubation of the cDNA-transfected cells with sodium [75Se]selenite. 75Se was incorporated into the expressed TR protein of the cells transfected with the SECIS element-containing cDNAs, but not into those without the SECIS element or with an inverted SECIS element. These data clearly showed a requirement of selenium for the formation of functional TR protein.

3' Untranslated Regions↗
PubMed1999

Cloning of the peroxiredoxin gene family in rats and characterization of the fourth member.

Peroxiredoxin (PRx) exhibits thioredoxin-dependent peroxidase activity and constitutes a family of proteins. Four members of genes from rat tissues were isolated by PCR using degenerated primers based on the sequences which encode a pair of highly conserved Cys-containing domains, and were then cloned to full-length cDNAs. These included two genes which have previously been isolated in rats, PRx I and PRx II, and two rat homologues of PRx III and PRx IV. We showed, for the first time, the simultaneous expression of all four genes in various rat tissues by Northern blotting. Since a discrepancy exists regarding cellular distribution, we further characterized PRx IV by expressing it in COS-1 cells. This clearly demonstrates that PRx IV is a secretory form and functions within the extracellular space.

Amino Acid Sequence↗
PubMed1999

Expression and characterization of the extracellular domain of guanylyl cyclase C from a baculovirus and Sf21 insect cells.

Guanylyl cyclase (GC)-C, a single-transmembrane receptor protein for heat-stable enterotoxin, guanylin, and uroguanylin, and its N-terminal extracellular domain were prepared at a high level of expression from a system constructed of Sf21 insect cells and recombinant baculovirus. The recombinant GC-C, containing the complete sequence, retained its binding affinity to heat-stable enterotoxin with a KD value (6.2 x 10(-10) M) and cyclase catalytic activity at a level similar to those of GC-C expressed in mammalian cell lines, such as COS-7. The N-terminal extracellular domain was prepared in a form which contained the hexahistidine tail at its C-terminus and was purified as a homogenous protein by Con A and Ni-chelating affinity chromatography from the culture medium of the insect cells. The purified N-terminal extracellular domain of GC-C exhibited the high (KD = 4 x 10(-10) M) and low (KD = 7 x 10(-8) M) affinity sites in binding to heat-stable enterotoxin. These results clearly indicate that the N-terminal extracellular domain of GC-C possesses the same biochemical characteristics as the complete GC-C protein even in the membrane-free form. Moreover, the extracellular domain is able to form an oligomer in a ligand-dependent manner, suggesting that the N-terminal extracellular domains interact with one another in binding to ligands.

Animals↗
PubMed1999

Different expression patterns of nitric oxide synthase isozymes in various gynecological cancers.

The expression of nitric oxide synthase (NOS) in human gynecological cancers, including ovarian cancers, uterocervical cancers, and endometrial cancers for example, was examined by the reverse transcriptase/polymerase chain reaction, coupled with Southern hybridization and by immunohistochemistry. Nitric oxide synthase II (NOS II), an inducible form, was expressed in more than 90% of the cancers. Nitric oxide synthase I (NOS I), a neuronal form, was expressed in 58% of all the ovarian cancers, in which the serous type is found more frequently (5 out of 7) than the mucinous type (2 out of 6), and in all clear-cell cancers. The frequency of NOS I expression in uterocervical cancers and endometrial cancers was relatively low. Nitric oxide synthase III (NOS III), an endothelial form, was detected in 25% of ovarian and 33% of endometrial cancers, while no expression was detected in uterocervical cancers. In terms of cancer types, all clear-cell adenocarcinomas and most of the serous-type adenocarcinomas expressed both NOS I and NOS II, while most uterine squamous carcinomas and endometrial adenocarcinomas expressed only NOS II. However, there was no correlation between the frequency of NOS expression and patients' age or the clinical stage of the disease. Since NO increases vascular permeability and blood flow, the high frequency of NOS expression in gynecological cancers may serve to stimulate and promote tumor growth.

Blotting, Southern↗
PubMed1999

Down regulation of superoxide dismutases and glutathione peroxidase by reactive oxygen and nitrogen species.

The levels of antioxidative enzymes are regulated by gene expressions as well as by post-translational modifications. Although their functions are to scavenge reactive oxygen (ROS) and nitrogen species (RNS), they may also be targets of various oxidants. When ROS and RNS modify the functions of antioxidative enzymes, especially glutathione peroxidase, they may induce apoptotic cell death in susceptible cells. It is conceivable, therefore, that at least a part of the apoptotic pathways mediated by ROS and RNS may be associated with modification of the redox regulation of cellular functions due to elevations of such substances. In this article we review recent findings about the effects of various oxidative conditions associated with alteration of these antioxidative enzymes and the concomitant cellular damage induced.

Animals↗
PubMed1999

Overexpression of the aldose reductase gene induces apoptosis in pancreatic beta-cells by causing a redox imbalance.

To determine the role of the polyol metabolizing pathway under hyperglycemic conditions, the effects of aldose reductase (AR) on the cellular functions of pancreatic beta-cells were examined. Stable transfectants of rat AR cDNA were obtained with a pancreatic beta-cell line, HIT, in which a negligible amount of AR was originally expressed. Overproduction of AR triggered DNA fragmentation, as judged with the TUNEL method and agarose gel electrophoresis. Morphological analysis by electron microscopy also clearly showed apoptosis of the AR-overexpressing HIT cells. Induction by interleukin-1beta of gene expression such as those of an inducible form of nitric oxide synthase (NOS-II) and Mn-superoxide dismutase (Mn-SOD), was much lower in the transfectants than in the control cells, while the expression of constitutively expressed genes such as those for Cu,Zn-superoxide dismutase and insulin was not changed. The susceptibility to interleukin-1beta stimulation of the expression of the NOS II and Mn-SOD genes was due to suppressed NF-kappaB activity, which is essential for the expression of these genes. In addition, the intracellular NADPH/NADP+ ratio was considerably lower in the AR-transfected cells than in control cells. Thus, the overexpression of AR in pancreatic beta-cells induced apoptosis that may be caused by a redox imbalance.

Aldehyde Reductase↗
PubMed1999

Angiography for assessment of preoperative chemotherapy in musculoskeletal sarcomas.

Eleven abnormal findings of digital subtraction angiography were analyzed in 25 patients with bone sarcoma and in 23 patients with soft tissue sarcoma. The relation between digital subtraction angiographic findings and the histologic effect of chemotherapy was evaluated. Digital subtraction angiography was performed with the patient under local anesthesia with the Seldinger technique, using an ADVANT X unit. Contrast medium was injected at a rate of 4 to 6 ml per second, with the usual single bolus dose of approximately 6 to 8 ml. Eleven abnormal findings included tumor stain, hypervascularity, arterial distortion, vascular stretch, arterial dilatation, arteriovenous shunt, arterial encasement, occlusion, blood pool, caliber with irregular wall, and dilatation of draining vein. Each finding after preoperative chemotherapy was compared with that before chemotherapy and divided into three grades; Grade 1, not effective; Grade 2, effective; and Grade 3, very effective. Angiographic Grades 2 and 3 were defined as responders. The histologic effect was examined and scored according to the modified classification proposed by Huvos. Histologic Grades 1 and 2 were classified as nonresponders and Grades 3 and 4 as good responders. In bone sarcomas, hypervascularity and tumor stain were seen in all patients. In soft tissue sarcomas, tumor stain was shown in all 23 patients and hypervascularity was seen in 21 patients. Tumor stain, hypervascularity, vascular stretch, and arterial encasement were correlated closely with histologic findings and showed an accuracy equal to or greater than 70%. When these four findings changed to angiographic Grade 3 after preoperative chemotherapy, 90% of patients had good histologic outcome.

Adolescent↗
PubMed1999

Chemotherapy for enterohemorrhagic Escherichia coli O157:H infection in a mouse model.

The aim of this study is to evaluate the therapeutic effect of the antimicrobial agents, fosfomycin (FOM), minocycline (MINO), kanamycin (KM) and norfloxacin (NFLX) in the enterohemorrhagic Escherichia coli (EHEC) infected mouse model which we established previously (Infect. Immun. 62 (1994) 3447-3453). Each of the antimicrobial agents, 1/16 LD(50), was given to the mice per os (p.o. ) or intraperitoneally (i.p.) for 3 days after bacterial inoculation and then we observed their mortality rate for 2 weeks. The mortality rates of mice administered with MINO (p.o./i.p.), KM (p.o.), NFLX (p. o./i.p.) were significantly lower than those of the control group. Both the bacterial number and VT2c level in the feces of the FOM group were lower than those of the NFLX group on day 1, but reversed on day 3. In an in vitro experiment, each of the four drugs in combination with mitomycin C (MMC) caused a more significant decrease in the bacterial number than sole MMC, and they consequently indicated the suppressive effect on the release of VT2c.

Administration, Oral↗
PubMed1999

Mapping of the alpha-1,6-fucosyltransferase gene, FUT8, to human chromosome 14q24.3.

Alpha-1,6-Fucosyltransferase (alpha1,6FucT) is involved in the biosynthesis of asparagine-linked glycoprotein oligosaccharides. In this study, we isolated a genomic clone for the human alpha1,6FucT gene (FUT8) and mapped it by fluorescence in situ hybridization to chromosome 14q24.3. This study suggests a distinct localization of FUT8 from genes for other human fucosyltransferases reported to date.

Chromosome Mapping↗
PubMed1999

The structural organization of the human aldehyde reductase gene, AKR1A1, and mapping to chromosome 1p33-->p32.

Genomic DNA encoding for human aldehyde reductase (AKR1A1), a member of the aldo-keto reductase superfamily, was isolated and characterized. The genomic DNA is approximately 16 kb in length and contains eight exons which encode the entire coding region and the 3'-untranslated sequences. AKR1A1 was localized on chromosome 1p33-->p32 by fluorescence in situ hybridization.

3' Untranslated Regions↗
PubMed1999

Loculated pericardial effusion due to congestive heart failure: an unusual case of vanishing tumor--a case report.

Loculation of a pleural effusion within an interlobar fissure as a result of congestive heart failure is a well-known entity. It has been termed "vanishing" or "phantom" tumor because its roentgenographic appearance simulates a pulmonary tumor and resolves with treatment of the congestive heart failure. The authors describe an 89-year-old man with a loculated pericardial effusion on the left cardiac border on chest roentgenogram. This was initially thought to represent an occult metastatic malignancy; however, its etiology became obvious when it disappeared with therapy of heart failure. Loculated pericardial effusion should be included in the differential diagnosis of roentgenographic densities in the chest when seen on the left cardiac border.

Aged↗
PubMed1999

Effect of clarithromycin and other macrolides on the sulfoxidation and 5-hydroxylation of lansoprazole in dogs.

The purpose of this study was to evaluate a possible interaction between lansoprazole and clarithromycin as well as other macrolides in dogs. Lansoprazole (30 mg) was orally administered to male beagle dogs, with or without oral pretreatment with 200-mg clarithromycin twice a day for 5 d. The experiments had a randomized cross-over design with a two-week washout period between dosing regimens. Clarithromycin pretreatment for 5 d resulted in a significant increase in the area under the serum lansoprazole concentration-time curve (AUC), whereas the area for a lansoprazole metabolite, lansoprazole sulfone, was significantly reduced, as was the maximum serum concentration (Cmax) of lansoprazole sulfone. When the effects of clarithromycin on the metabolism of lansoprazole were studied using dog liver microsomes, it was found that clarithromycin significantly inhibited the formation of lansoprazole sulfone but not another metabolite, 5-hydroxylansoprazole. These results suggest that co-medication of lansoprazole with clarithromycin may produce a synergistic effect caused by the increased serum levels of lansoprazole of benefit in Helicobacter pylori eradication.

2-Pyridinylmethylsulfinylbenzimidazoles↗
PubMed1999

Dietary salt loading increases nitric oxide synthesis in transgenic mice overexpressing sodium-proton exchanger.

We studied the role of nitric oxide (NO) synthesis in amelioration of blood pressure elevation during dietary salt loading in transgenic mice overexpressing sodium proton exchanger. Systolic blood pressure rose after starting salt loading only in the high-salt group of transgenic mice. However, this elevation of blood pressure was not continued. Urinary excretion of inorganic nitrite and nitrate in the high-salt group of transgenic mice was significantly higher than in the high-salt group of control mice. These results suggest that increased NO synthesis in response to salt loading is one of the anti-hypertensive mechanisms in transgenic mice overexpressing sodium proton exchanger.

Animals↗
PubMed1999

Factors related to impairment of activities of daily living.

OBJECTIVE: We examined the factors related to the impairment of activities of daily living (ADL). METHODS: ADL was evaluated by using ADL-20, which consists of 20 items from 4 major categories of activities; mobility, self-care, instrumental, and communication. The patients' gender, birth date, clinical diagnosis, past history, life styles, physical findings, laboratory data, and details of therapy were also recorded. Patients A total of 1,163 outpatients aged 50 years or older were included. Data from 1,093 patients were analyzed. RESULTS: We divided the subjects into two groups; Group I having full marks of ADL-20 (n=582) and group II exhibiting an impairment of ADL (n=511). Multiple logistic analysis revealed that in both sexes age and stroke were common independent factors related to the impairment of ADL. Other factors associated with impairment of ADL were smoking in men and presence of proteinuria in women. The presence of hyperlipidemia was associated with preservation of the ADL in women. CONCLUSION: The results demonstrated significant associations of smoking in men and the presence of proteinuria in women with the impairment of ADL in elderly Japanese outpatients. There appears to be a sex difference in the risk factors of impairment of ADL.

Activities of Daily Living↗
PubMed1999

[An 88-year-old woman with symptoms of intoxication due to a small dose of digoxin].

An 88-year-old woman was admitted to our hospital because of palpitations, dyspnea, orthopnea and appetite loss. On admission, small crackles were heard on her lower back, and her liver was swollen. Chest rentogenogram showed cardiomegaly (cardio-thoracic ratio 65.5%) and bilateral pleural effusion. Electrocardiograms showed atrial fibrillation with an average heart rate of 95 per minute. Echocardiography revealed mitral stenosis. Because the patient was considered to be suffered from heart failure due to mitral stenosis with atrial fibrillation, furosemide (20 mg per day) and digoxin (0.25 mg per day) was started. After digoxin had been raised to a dose of 0.50 mg per day because of sustained rapid ventricular response on the fourth hospital day, she complained of nausea and vomiting. Serum digoxin concentration was 2.55 ng/ml on the next day, and 1.08 ng/ml 96 hours after discontinuing digoxin. There was no complaint after digoxin was restarted with a dose of 0.05 mg per day. She complained of nausea again on the third day when the digoxin was raised to a dose of 0.083 mg in a blinded study. This observation indicates that digoxin intoxication could occur even in the smaller dose of digoxin than usual in the elderly.

Aged↗
PubMed1999

[The deterioration of regimen comprehension in self-attending elderly patients and the effect of tutorials by pharmacists].

The purpose of this study was to assess whether regimen comprehension deteriorates in the elderly without obvious mental disability. Eligible patients were elderly who could visit hospitals by themselves. We recruited 138 patients (age: 43-89, 75 males and 63 females, underlying diseases: hypertension, hyperlipidemia, arrhythmia etc.) from our outpatient clinic. The participants were tested with a regimen comprehension scale (RCS: Jpn J Geriat 1997; 34:209-214). The differences in scores among individuals increased with age. Scores of 5 or less were recorded in 10 of 69 patients aged 65 or more, but in non of 69 patients aged less than 65 (p < 0.01). The 60 patients who could not get full marks were classified into 2 groups: the tutored group (T) who were tutored by pharmacists about taking drugs, and the non-tutored group (N). Both group were tested again with RCS to evaluate the effect of tutorial. In T-group (n = 28), the second scores increased significantly (from 7.2 +/- 0.9 to 8.6 +/- 2.0 (m +/- SD); p < 0.01). Although the second scores showed a tendency to increase in N-group (n = 29), there was no statistical significance. In 7 patients who obtained less than a score of 5 on the RCS and age- and gender-matched controls who received full marks on the RCS, the HDS-R test failed to show any differences between the two groups. Thus, we concluded that even in the self-attending elderly patients, the regimen comprehension deteriorated with age, and tutorials were considered to be effective.

Adult↗