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Biomedical subjects

S Nagata

Publications and source records attributed to S Nagata.

At least 109 records · Page 6Linked to original sources
PubMed2000

Identification of 187 single nucleotide polymorphisms (SNPs) among 41 candidate genes for ischemic heart disease in the Japanese population.

To investigate whether common variants in the human genetic background are associated with pathogenesis of ischemic heart diseases, we systematically surveyed 41 possible candidate genes for single-nucleotide polymorphisms (SNPs) by directly sequencing 96 independent alleles at each locus, derived from 48 unrelated Japanese patients with myocardial infarction, including 25.8 kb 5' flanking regions, 56.8 kb exonic and 35.4 kb intronic sequences, and 1.8 kb 3' flanking regions. In this genomic DNA of nearly 120 kb, we identified 187 SNPs: 55 in 5' flanking regions, seven in 5' untranslated regions (UTRs), 52 in coding elements, 64 in introns, eight in 3' UTRs, and one in a 3' flanking region. Among the 52 coding SNPs, 26 were non-synonymous changes. Allelic frequencies of some of the polymorphisms were significantly different from those reported in European populations. For example, the Q506R substitution in the coagulation factor V gene, the so-called "Leiden mutation", has a reported frequency of 2.3% in Europeans, but we detected the Leiden mutation in none of the Japanese genomes that we investigated. The allelic frequencies of the -33A>G SNP in the thrombomodulin gene were also very different; this allele occurred at a 12% frequency in the Japanese patients that we examined, although it had been detected in none of 82 Caucasians reported previously. These data support the hypothesis that some SNPs are specific to particular ethnic groups.

3' Untranslated Regions↗
PubMed2000

Enzyme-linked immunosorbent assay for anti-tropomyosin antibodies and its clinical application to various heart diseases.

Tropomyosin is one of the key proteins for muscle contraction. We developed an enzyme-linked immunosorbent assay for antibodies to porcine muscle tropomyosin and measured serum anti-tropomyosin antibodies in patients with heart diseases and in normal controls. The mean values of absorbance in the ELISA assay of patients with ischemic heart disease (n=36, P<0.001), dilated cardiomyopathy (n=28, P<0.005), valvular heart disease (n=27, P<0.05), and collagen disease (n=38, P<0.05) were significantly higher than those of normal controls (n=53), but the value in patients with hypertrophic cardiomyopathy (n=19) was not significantly different from that of normal controls. When the cut-off value was fixed at the mean+2 SD of absorbance in normal controls, positive reactions were found in 19.4%, 7.1%, 18.5% and 15.8% of patients with ischemic heart disease, dilated cardiomyopathy, valvular heart disease, and collagen disease, respectively. An inhibition study revealed that anti-tropomyosin antibodies were different from anti-myosin antibodies, but there was a partial cross-reactivity between the two. Thus, there was a weak correlation of the titers of the two types of antibody within the group of heart diseases. These data indicate that measurement of anti-tropomyosin antibodies by ELISA is helpful for detecting autoimmune abnormalities in various heart diseases.

Adult↗
PubMed2000

Immunoaffinity column as clean-up tool for determination of trace amounts of microcystins in tap water.

Trace amounts of microcystins (MCs) in drinking water should be monitored because of their potential hazard for human health as an environmental tumor promoter. We describe here a new clean-up tool with immunoaffinity column (IAC) for determination of trace amounts of MCs (from pg to microg/litre) in tap water. The water samples were concentrated with IAC clean-up and MCs levels were determined by HPLC with UV detection or enzyme-linked immunosorbent assay (ELISA). In the combination with HPLC analysis, mean recovery of microcystin-LR (MCLR),-RR and-YR spiked to tap water were 91.8%, 77.3% and 86.4%, respectively, in the range 2.5-100 microg/litre. The chromatogram of MCs-spiked tap water sample cleaned up with IAC showed effective elimination of the impurities compared to that with octadecyl silanized cartridge, which had been cleaned up with a conventional method. Also, in the combination with highly sensitive ELISA, mean recovery of MCLR spiked to tap water was 80% in the range 0.1-1000 ng/litre. The combined methods developed here can detect pg to microg/litre of MCs in tap water. The overall results indicated that IAC will be suitable as a clean-up tool for trace amounts of MCs in tap water.

Bacterial Toxins↗
PubMed2000

Dynamic regulation of prothoracic gland ecdysteroidogenesis: Manduca sexta recombinant prothoracicotropic hormone and brain extracts have identical effects.

Multiple assays were conducted in order to determine if the recently available recombinant prothoracicotropic hormone (rPTTH) from Manduca sexta is identical, or similar, to the natural hormone and if results from its use in a variety of assays confirm, or are inconsistent with, previous studies over the past 20years on PTTH action using brain extract. Brain extracts and rPTTH showed similar, if not identical, effects on the cell biology of Manduca prothoracic gland cells with the following results: increased levels of cAMP (adenosine 3':5' cyclic monophosphate) synthesis; requirement for extracellular Ca(2+) in in vitro studies; ecdysteroidogenesis stimulation in vitro; stimulation of general and specific protein synthesis; immunocytochemical identification of the two lateral cells in each brain hemisphere as the source of PTTH (the prothoracicotropes); the ability of antibodies to rPTTH to inhibit ecdysteroidogenesis stimulation in vitro; and the multiple phosphorylation of the ribosomal protein S6. The data revealed that brain extract and rPTTH show equivalent effects in all of the assays, indicating that this rPTTH is the natural PTTH of Manduca and that the data generated with brain extracts over the past two decades are indeed relevant.

Animals↗
PubMed2000

Relationship between the reduction of tetrazolium salt XTT and DNA strand breakage with aminosugars.

Dihydropyrazine derivatives formed by the self-condensation reaction of D-glucosamine have the DNA breaking activity. To establish the monitoring method of the biological active dihydropyrazines, we investigated the relationship between the XTT (3'-[1-[(phenylamino)-carbonyl]-3, 4-tetrazolium]bis(4-methoxy-6-nitro)benzensulfonic acid hydrate) reducibility and the DNA breaking activity of aminosugars. Aminosugar in 50 mM sodium phosphate buffer (pH 7.4) was incubated at 37 degrees C. At a given time, the XTT reducibility and the DNA breaking activity of the incubated aminosugar were measured. Both XTT reducibility and DNA breaking activity showed a maximum value within 1-4 h after the incubation and then gradually decreased with the incubation time. Superoxide anion was suggested to involve in both of the DNA breaking activity and the XTT reducibility by the addition of the radical scavengers into those assay mixtures. The quantity of remaining covalently closed circular DNA and the XTT reducibility of all aminosugars showed a good correlation (r = 0.825, n = 26). This means that the XTT assay is applicable for the monitoring of those biologically active products derived from aminosugars when the participation of superoxide anion in DNA scission is recognized.

DNA↗
PubMed2000

Modulation of T-cell-mediated immunity in tumor and graft-versus-host disease models through the LIGHT co-stimulatory pathway.

LIGHT was recently described as a member of the tumor necrosis factor (TNF) 'superfamily'. We have isolated a mouse homolog of human LIGHT and investigated its immunoregulatory functions in vitro and in vivo. LIGHT has potent, CD28-independent co-stimulatory activity leading to T-cell growth and secretion of gamma interferon and granulocyte-macrophage colony-stimulating factor. Gene transfer of LIGHT induced an antigen-specific cytolytic T-cell response and therapeutic immunity against established mouse P815 tumor. In contrast, blockade of LIGHT by administration of soluble receptor or antibody led to decreased cell-mediated immunity and ameliorated graft-versus-host disease. Our studies identify a previously unknown T-cell co-stimulatory pathway as a potential therapeutic target.

Amino Acid Sequence↗
PubMed2000

Structure of the heterodimeric complex between CAD domains of CAD and ICAD.

We present here the structure of the complex between the CAD domain of caspase activated deoxyribonuclease (CAD) and the CAD domain of its inhibitor (ICAD), determined by nuclear magnetic resonance spectroscopy. The two domains adopt a very similar fold, which consists of an alpha-helix and a beta-sheet, and are aligned side by side in the complex. Notably, the positive charges on the strand beta2 at one end of the beta-sheet of CAD and negative charges around the opposite end of the beta-sheet of ICAD are paired in the complex. Point mutations of the charged amino acids at this interface, on either CAD or ICAD, prevented formation of the functional CAD-ICAD complex. This implies that the interaction between the CAD domains of CAD and ICAD is an essential step in the correct folding of CAD in the complex.

Amino Acid Sequence↗
PubMed2000

Mice with markedly reduced PACAP (PAC(1)) receptor expression by targeted deletion of the signal peptide.

In an attempt to study the pituitary adenylate cyclase-activating polypeptide (PACAP) type 1 (PAC(1)) receptor (PAC(1)R) function in vivo and to produce a mouse model with altered expression of PAC(1)R, we have used gene targeting in embryonic stem cells to disrupt exon 2 of the PAC(1)R gene, which contains the ATG translation start site and the signal peptide. Un-expectedly, active transcription of PAC(1)R mRNA was detected in the mutant mice; however, exon 1 was spliced to exon 3 (skipping exon 2), and (125)I-PACAP27 binding in brain was greatly reduced. PAC(1)R exon 2(-/-) mice were viable, fertile, and morphologically and histologically indistinguishable from their wild-type counterparts. We next examined the ligand binding and cell surface expression of the mutant receptor lacking the signal peptide in transfected COS-7 cells. (125)I-PACAP27 binding of the mutant receptor was approximately one-tenth of that in the wild-type receptor. Although the wild-type receptor was expressed abundantly in both the plasma membrane and the cytoplasm around the nucleus, the mutant receptor was expressed in the plasma membrane with a markedly reduced level. Digestion of the membranes with endoglycosidase F greatly reduced the size of the wild-type receptor but only slightly reduced that of the mutant receptor. These results demonstrate that the signal peptide is required for efficient cell surface expression and N-linked glycosylation of the PAC(1)R. However, the mutant receptors still functionally coupled to adenylate cyclase in COS-7 cells, suggesting the presence of sufficient spare receptors such that the mutant receptors are capable of activating the second messenger system. We suggest that the mutant mice with markedly reduced PAC(1)R expression can serve as a useful animal model or cell culture system for further studies in PAC(1)R function.

Animals↗
PubMed2000

Fas-mediated cholangiopathy in the murine model of graft versus host disease.

Bile-duct injury observed in hepatic graft versus host disease (GVHD) is regarded as an immune-mediated injury, although its precise mechanism is unclear. However, recent studies have suggested the involvement of Fas-mediated cell death in this immune-mediated cholangiopathy. In this study, we first showed the constitutive expression of Fas receptor by cholangiocytes in situ from normal BALB/c mice, which was upregulated in GVHD mice. Also, we confirmed the Fas protein expression in the isolated cholangiocytes from normal BALB/c mice by immunocytochemistry and immunoblotting. Furthermore, the addition of agonistic Fas antibody-(Jo2)-induced cholangiocyte apoptosis confirmed by DNA-ladder formation and annexin V staining. Cholangiocytes from Fas-deficient mice (MRL lpr/lpr) did not show Jo2-induced apoptosis. Interferon-gamma augmented Fas expression and Fas-mediated cell death, respectively. Following these observations, experimental GVHD was induced by transfer of splenocytes from B10.D2 mice to irradiated (800 rad) BALB/c mice. Liver-infiltrating lymphocytes from the recipient showed dose-dependent cytotoxicity against (51)Cr-labeled cholangiocytes isolated from BALB/c mice. Moreover, the addition of blocking Fas-Fc fusion protein reduced this cytotoxicity to 44.7%. Finally, administration of this Fas-Fc protein to the BALB/c mice, which had been adoptively transferred with splenocytes of B10.D2 mice, prevented the development of hepatic GVHD in vivo. These results showed the involvement of Fas-mediated cell death in cholangiopathy observed in GVHD, and a soluble Fas-Fc protein may have a therapeutic potential for hepatic GVHD.

Animals↗
PubMed2000

Significance of Fas antigen-mediated apoptosis in human fulminant hepatic failure.

OBJECTIVE: The aim of this study was to elucidate the role of apoptosis in human fulminant hepatic failure. We studied the expression of Fas antigen on liver tissues, Fas ligand in lymphocytes, and soluble Fas ligand in patients' serum. METHODS: On finding apoptotic cells in fulminant hepatic failure liver, we first examined them using the TUNEL method. Subsequently, the expression of Fas was studied by immunostaining. Simultaneously, Fas ligand presenting on both liver-infiltrated cells and peripheral lymphocytes was studied by reverse transcription-polymerase chain reaction, and soluble Fas ligand in sera was measured by ELISA. RESULTS: By using the TUNEL method, we first demonstrated that many apoptotic cells existed in fulminant hepatic failure but not in normal ones. Our immunohistochemistry study showed that many hepatocytes in fulminant hepatic failure strongly expressed Fas. In addition, Fas ligand on both liver-infiltrating lymphocytes and peripheral lymphocytes in fulminant hepatic failure patients was detected. The serum level of soluble Fas ligand was significantly increased in fulminant hepatic failure (mean value, 2.91 ng/ml in fulminant hepatic failure [n = 10], 1.62 ng/ml in acute hepatitis [n = 10], and 0.27 ng/ml in healthy controls [n = 10]). Furthermore, this serum level of sFas ligand was significantly associated with prothrombin time both in acute hepatitis and fulminant hepatic failure. CONCLUSIONS: The present results indicate that Fas-mediated apoptosis may be one of the triggers for the induction of fulminant hepatic failure.

Adult↗
PubMed2000

The eosinophil peroxidase gene forms a cluster with the genes for myeloperoxidase and lactoperoxidase on human chromosome 17.

Eosinophil peroxidase (EPX) is one of a family of mammalian peroxidases that includes myeloperoxidase (MPO), lactoperoxidase (LPO), and thyroid peroxidase (TPO). Here we show that the human EPX gene maps to chromosome 17q23.1, which localizes 34 kb from the LPO and MPO genes. Our results demonstrate that the EPX, LPO, and MPO genes form a cluster on human chromosome 17.

Chromosomes, Human, Pair 17↗
PubMed2000

[Stress management in the workplace in the era of industrial and economic change].

The globalization of the economy and the recent economic recession in Japan has accelerated down-sizing or restructuring of corporations and has resulted in the induction of a wage system according to achievement instead of the traditional seniority wage system, break-down of the life-long employment system, excess labor and increased unemployment. These rapid changes in the labor situation have increased job stress. It was reported in the survey conducted by the Ministry of Labor in 1997 that 62.8 percent of 16,000 workers had anxiety, worry and stress regarding their working life. The need for effective stress management at work has been increasing in this situation, but in the survey mentioned above only 26.5 percent of 12,000 companies replied that they had incorporated mental health measures. The characteristic features of the approaches for stress management in Japan are summarized as follows: 1) The most popular approaches are education and consultation for individual workers. 2) Systematic preventive approaches such as work control, working environment control, organizational change in the health management system, and systematic and continuous educational programs for managers are inadequate. 3) Systems to evaluation the effectiveness of these interventional approaches are also inadequate. Considering the current situation in which there is increasing job stress and a need for the occupational mental health promotion, we propose a series of mini-reviews regarding stress management at work and mention the composition of this series.

Health Promotion↗
PubMed2000

Pit pattern diagnosis of early colorectal carcinoma by magnifying colonoscopy: clinical and histological implications.

The relation between subclassification of V type pit pattern and histopathologic features, particularly, in regards to the invasion depth, was examined in 75 cases of early colorectal carcinoma. After we classified two grades (VA, VN) of V type pit pattern, we subclassified VN type pit pattern into three subtypes (Grade A, B, and C) according to the degree of appearance of VN pit pattern. V type pit pattern subclassification was diagnosed with use of a magnifying colonoscope. There were 37 VA type lesions, 8 VN.Grade A type, 16 VN.Grade B type, and 14 VN.Grade C type lesions. The incidence of submucosal massive invasion (sm2, sm3) rates was 5 (14%) of the 37 VA type lesions, 3 (38%) of the 8 VN.Grade A type, 15 (94%) of the 16 VN.Grade B type and 14 (100%) of the 14 VN.Grade C type lesions, respectively. The incidence of desmoplastic reaction (++) levels in VN.Grade B and C type lesions was significantly higher than that in VA type lesions (P<0.01), and that in VN.Grade C type lesions was significantly higher than that in VN.Grade A type lesions (P<0.05). The incidence of pit disorder or destruction (+) and (++) levels in VN.Grade B and C type lesions was significantly higher than that in VA type lesions (P<0.01), and that in VN.Grade C type lesions was significantly higher than that in VN.Grade A type lesions (P<0.01). The incidence of high grade carcinoma in VN.Grade B and C type lesions was significantly higher than that in VA type lesions (P<0.01), and that in VN.Grade C type lesions was significantly higher than that in VN.Grade A type lesions (P<0.05). These findings indicate that V type pit pattern subclassification, including our new subtype, is useful for predicting the detail invasion depth of early colorectal carcinoma. These V pit pattern subtypes are based on the degrees of desmoplastic reaction, pit disorder or destruction, and histologic grade of carcinoma.

Carcinoma↗
PubMed2000

Soluble Fas ligand expression in the ocular fluids of uveitis patients.

PURPOSE: The expression of Fas ligand (FasL) in ocular tissues is thought to play a critical role in maintaining immune privilege in the eye. In this study, to clarify the involvement of the Fas-FasL system in inflammatory processes of the eye,we examined soluble FasL (sFasL) in ocular inflammation. METHODS: Using ELISA systems recently developed, sFasL concentrations in aqueous humor (AH) and/or vitreous fluid (VF) were measured. AH was obtained from 17 eyes of 17 uveitis patients and from 12 eyes of 12 non-uveitis (cataract) patients. VF was obtained from 22 eyes of 22 uveitis patients and 7 eyes of 7 non-uveitis (macular hole) patients. Serum levels of sFasL were also determined. RESULTS: sFasL in AH and VF was below the detection limit of the ELISA systems in all non-uveitis eyes. On the other hand, sFasL was detected in AH from uveitis patients where it measured 367.0 +/- 154.7 pg/ml (mean +/- SEM). sFasL was also detected in VF from uveitis patients where it measured 1132.2 +/- 281.7 pg/ml. None of the sera from patients with or without uveitis contained a detectable level of sFas L. CONCLUSIONS: sFasL levels in AH and VF are elevated in the eye during ocular inflammation. Fas-FasL mediated apoptosis may play an important role in the regulation of inflammation during uveitis.

Aged↗
PubMed2000

Detailed colonoscopy for detecting early superficial carcinoma: recent developments.

Early superficial colorectal carcinoma (CRC) has been commonly detected by routine colonoscopic examination in Japan. A series of 769 early CRCs and 4821 adenomas were diagnosed by colonoscopy at Hiroshima University Medical Hospital between 1991 and 1998. Of early CRCs, macroscopically superficial early CRCs accounted for 209 lesions (28%). Among 5590 lesions of adenomas and carcinomas, depressed-type lesions showed a significantly higher malignant potential for cancerous and submucosal invasion than elevated lesions (polypoid, IIa-type lesions, and G-LST). As one of the quantitative examinations for early CRC, pit pattern observed by magnifying video-colonoscopy was useful. We performed magnifying observations for 265 lesions of colorectal neoplasias using Kudo's pit pattern classification for 2 years. Depressed-type lesions characterized the Ills and V pit patterns, and elevated lesions characterized the III(L) pit pattern. The incidence of cancer was significantly higher in lesions with IIIs and V pit patterns. Furthermore, the V(N) pit pattern was considered a significant indicator of submucosal invasion. These results indicated that superficial early CRC could be considered to constitute about one-third of all early CRCs. Of them, the depressed-type lesions showed a significantly higher malignant potential than elevated lesions. Pit pattern observation by magnifying videocolonoscopy is useful for predicting the histology/invasion depth of early CRC.

Adenomatous Polyps↗