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Biomedical subjects

H Ashida

Publications and source records attributed to H Ashida.

At least 37 records · Page 2Linked to original sources
PubMed2000

3-Amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1) induces apoptosis in rat splenocytes and thymocytes by different mechanisms.

3-Amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1) is a potent carcinogen present in cooked meat. Although the target of this carcinogen is mainly in the liver, Trp-P-1 is distributed in the thymus and spleen as well as in the liver after administration. However, the cytotoxic effect of Trp-P-1 on lymphocytes has not been examined in detail. In the present study, we investigated the cytotoxicity of Trp-P-1 against rat splenocytes and thymocytes. Trp-P-1 reduced viability of both types of cells in the same manner, the LD(50) at 6h in culture was 15 microM, and the time for the 50% decrease in cell viability (t(1/2)) at 20 microM was 3h. In both types of cells, Trp-P-1 caused the activation of caspase-3-like proteases and the cleavage of poly(ADP-ribose) polymerase, both of which are biochemical markers of apoptosis. On the other hand, DNA fragmentation occurred in splenocytes, but not in thymocytes although Trp-P-1 activated 32-34kDa nucleases that may not be able to degrade DNA into nucleosomal units. These results indicated that Trp-P-1 induces apoptosis in both splenocytes and thymocytes by different mechanisms in which distinct apoptotic pathways may exist downstream of the caspase cascade.

Animals↗
PubMed2000

Molecular cloning of cDNA encoding alpha-N-acetylgalactosaminidase from Acremonium sp. and its expression in yeast.

Alpha-N-acetylgalactosaminidase (alpha-GalNAc-ase; EC 3.2.1.49) is an exoglycosidase specific for the hydrolysis of terminal alpha-linked N-acetylgalactosamine in various sugar chains. The cDNA, nagA, encoding alpha-GalNAc-ase from Acremonium sp. was cloned, sequenced, and expressed in yeast Saccharomyces cerevisiae. The nagA contains an open reading frame which encodes for 547 amino acid residues including 21 residues of a signal peptide in its N-terminal. The calculated molecular mass of mature protein from the deduced amino acid sequence of nagA is 57260 Da, which corresponds to the value obtained from SDS-PAGE of native and recombinant enzymes treated with endo-beta-N-acetylglucosaminidase H. The amino acid sequence of NagA showed significant similarity to those of eukaryotic alpha-GalNAc-ases and alpha-galactosidases (alpha-Gal-ases), particularly alpha-Gal-ase A (AglA) from Aspergillus niger. Phylogenetic analysis revealed that NagA does not belong to the cluster of vertebrate alpha-GalNAc-ase and alpha-Gal-ase but forms another cluster with AglA and yeast alpha-Gal-ases. Thus, the evolutionary origin of the fungal alpha-GalNAc-ase is suggested to be different from that of vertebrate alpha-GalNAc-ase. This is the first report of a microbial alpha-GalNAc-ase gene.

Acremonium↗
PubMed2000

Flavones and flavonols at dietary levels inhibit a transformation of aryl hydrocarbon receptor induced by dioxin.

Dioxins invade the body mainly through the diet, and produce toxicity through the transformation of aryl hydrocarbon receptor (AhR). An inhibitor of the transformation should therefore protect against the toxicity and ideally be part of the diet. We examined flavonoids ubiquitously expressed in plant foods as one of the best candidates, and found that the subclasses flavones and flavonols suppressed antagonistically the transformation of AhR induced by 1 nM of 2,3,7,8-tetrachlorodibenzo-p-dioxin, without exhibiting agonistic effects that transform AhR. The antagonistic IC(50) values ranged from 0.14 to 10 microM, close to the physiological levels in human.

Animals↗
PubMed2000

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD)-induced changes in activities of nuclear protein kinases and phosphatases affecting DNA binding activity of c-Myc and AP-1 in the livers of guinea pigs.

To study the effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on nuclear protein phosphorylation activities, male guinea pigs were treated in vivo with a single 1 microg/kg i.p. injection of TCDD, and the state of protein kinases and phosphatases in the nuclei of the hepatocytes was examined after 1, 10, and 40 days. TCDD was found to cause a rise in nuclear protein tyrosine kinase on day 1, and to a lesser extent on day 10, but this effect diminished almost completely on day 40. TCDD also caused a reduction in nuclear casein kinase II (CKII) activity at all time points. To study the biochemical events taking place at the early stage of the action of TCDD, a short-term in vitro model system was established using explant liver tissues maintained in tissue culture medium. It was found that TCDD caused a rapid reduction of the activity of nuclear CKII with an accompanying increase in the cytosol. Such changes in protein phosphorylation activities were also accompanied by an increase in the DNA binding activity of activator protein 1 (AP-1). The effect of TCDD on nuclear proteins binding to the c-Myc response element DNA was, on the other hand, biphasic: an initial increase of protein binding to the c-Myc response element was followed by suppression. To test the hypothesis that some of the above changes were caused by TCDD-induced changes in protein kinase activity, nuclear proteins isolated from hepatocytes of in vivo treated guinea pigs were incubated with exogenously added Mg2+ and ATP under cell-free conditions. The results showed that this in vitro phosphorylation treatment exacerbated this tendency of increased AP-1 and decreased c-Myc binding to their respective response element DNAs, indicating that kinases and phosphatases present in the isolated nuclear protein preparation were active and capable of modifying protein binding to DNA. Such effects of Mg2+ and ATP on AP-1 were blocked by heparin, indicating that CKII plays an important role in transducing the signal of TCDD into the nucleus.

Animals↗
PubMed2000

Characterization of endo-alpha-N-acetylgalactosaminidase from Bacillus sp. and syntheses of neo-oligosaccharides using its transglycosylation activity.

Endo-alpha-N-acetylgalactosaminidase was purified to homogeneity from the culture fluid of Bacillus sp. isolated from soil and characterized. The molecular mass of the enzyme was estimated as 110 kDa. The enzyme was stable at pH 4.0-10.0, up to 55 degrees C, and was most active at pH 5.0. The substrate specificity of the enzyme was strict for the disaccharide, galactosyl beta1, 3 N-acetyl-d-galactosamine, bound to aglycone in alpha configuration. On the other hand, the specificity of the enzyme for the aglycone structure was fairly relaxed. The enzyme could transfer the disaccharide from para-nitrophenyl substrate to various acceptors, such as monosaccharides, disaccharides, and sugar alcohols. Using this transglycosylation activity of the endoglycosidase, it may be possible to synthesize neo-oligosaccharides.

Asialoglycoproteins↗
PubMed2000

Suppressive effects of flavonoids on dioxin toxicity.

Dioxin type chemicals such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) cause a variety of toxicity. Most of the toxicity of TCDD has been attributed to a mechanism by which TCDD is bound to aryl hydrocarbon receptor (AhR) and transforms the receptor. Thus, suppression of the AhR transformation by food factors can suppress the dioxin toxicity. In this study, flavonoids at various concentrations were treated to a rat cytosolic fraction containing AhR before adding 1 nM TCDD. The transformed AhR was detected by an electrophoretic mobility shift assay with a DNA oligonucleotide consensus to dioxin response element. As the results, flavones and flavonols at dietary levels act as the antagonists for AhR and suppress the transformation. The antagonistic IC50 values were in a range between 0.14 and 10 microM, which are close to the physiological levels in human. These results suggest that a plant-based diet can prevent the dioxin toxicity.

Animals↗
PubMed2000

A hierarchical structure of motion system revealed by interocular transfer of flicker motion aftereffects.

Interocular transfer of the motion aftereffect (MAE) has been extensively investigated for the purpose of analysing the binocularity of the underlying motion mechanism. Previous studies unanimously reported that the transfer of the classical static MAE is partial, but there is a controversy as to whether the transfer of the flicker MAE (MAE measured using counterphase gratings) is partial or perfect. To gain insight into the discrepancy between studies, we investigated whether the interocular transfer of the flicker MAE is influenced by the MAE measurement method, retinal eccentricity and attention. Our results showed that the transfer was perfect or nearly so when the MAE duration was measured in the central visual field with observers paying attention to the adaptation stimulus, but the transfer was partial when the MAE nulling strength was measured, when the MAE duration was measured in the peripheral visual field, or when the observers' attention was distracted by a secondary task. These results not only resolve discrepancies between previous studies, but also suggest that the flicker MAE reflects adaptation at multiple stages in the hierarchical architecture of motion processing.

Adaptation, Physiological↗
PubMed2000

Fucogalactan isolated from Sarcodon aspratus elicits release of tumor necrosis factor-alpha and nitric oxide from murine macrophages.

Eight species of mushrooms were evaluated for mitogenic activity by the tetrazolium salt 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) method using spleen cells of C3H/HeN female mice. The hot water-soluble (HWS) fraction extracted from Sarcodon aspratus showed the highest activity. The mitogen in Sa. aspratus was isolated by Sepharose 6B and DEAE-Sepharose CL-6B column chromatography. Preliminary structural analyses indicated that the mitogen was a fucogalactan. Fucogalactan elicited the release of tumor necrosis factor-alpha (TNF-alpha) and nitric oxide (NO) in macrophages of mice in vitro. TNF-alpha production induced with 50 microg/ml of fucogalactan was significantly higher than that induced by lentinan (500 microg/ml) by approximately 4.3-fold. Also, fucogalactan showed dose dependence at concentrations from 5 to 500 microg/ml in NO production. Thus, fucogalactan does elicit the release of cytokines such as TNF-alpha and NO.

Adjuvants, Immunologic↗
PubMed2000

Independent aftereffects of attention and motion.

In the motion aftereffect (MAE), a stationary pattern appears to move in the opposite direction to previously viewed motion. Here we report an MAE that is observed for a putatively high level of visual analysis-attentive tracking. These high-level MAEs, visible on dynamic (but not static) tests, suggest that attentive tracking does not simply enhance low-level motion signals but, rather, acts at a subsequent stage. MAEs from tracking (1) can overrule competing MAEs from adaptation to low-level motion, (2) can be established opposite to low-level MAEs seen on static tests at the same location, and (3), most striking, are specific to the overall direction of object motion, even at nonadapted locations. These distinctive properties suggest MAEs from attentive tracking can serve as valuable probes for understanding the mechanisms of high-level vision and attention.

Attention↗
PubMed2000

Left cervical aortic arch with aortic coarctation and saccular aneurysm.

Cervical aortic arch is a very rare malformation and is occasionally accompanied by other cardiovascular anomalies. A 48-year-old male patient had a left cervical aortic arch with aortic coarctation and saccular aneurysm distal to the coarcted segment. The major clinical manifestations were upper body hypertension with a 50-mmHg discrepancy between the upper and lower limbs and a loud continuous murmur in the upper chest and back. Magnetic resonance angiography successfully depicted the anomalous aorta, and the aortic coarctation and aneurysm were surgically resected and the thoracic aorta was reconstructed. The discrepancy in blood pressure diminished after the operation, but antihypertensive medication was continued to satisfactorily control the hypertension.

Aorta, Thoracic↗
PubMed2000

Cytochrome P4501A1-inhibitory action of antimutagenic anthraquinones in medicinal plants and the structure-activity relationship.

We have earlier found that flavones and flavonols in vegetables specifically inhibited one of the carcinogenesis-related enzymes, cytochrome P450 (CYP) 1A1, and subsequently suppressed the mutagenicity of food-derived carcinogens. In this study, we explored other candidates for the enzyme inhibitor in Chinese medicinal plants. Some of them were antimutagenic toward 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2). For example, Rheum officinale contained large amounts of anthraquinones as the active compounds, 3.4 mg of emodin, 2.1 mg of chrysophanol and 1.8 mg of rhein in 10 g of dry matter. Anthraquinones showed similar IC50 values for antimutagenicity against Trp-P-2 to those for inhibition of the N-hydroxylation activity of CYP1A1 toward Trp-P-2, indicating that the antimutagenicity was attributable to CYP inhibition. The structure-activity relationships were then examined with 14 commercial chemicals, and it was found that the interaction with an enzyme required three rings and an oxygen group in the side ring. This characteristic is similar to that of flavones and flavonols.

Anthraquinones↗
PubMed2000

Detection of biomarkers for apoptosis in rat liver after perfusion with 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1).

We previously demonstrated that 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1) induced apoptosis in primary cultured rat hepatocytes. In this study, we investigated apoptotic biomarkers in rat liver after perfusion with 30 microM Trp-P-1 as preliminary experiments for in vivo study. Induction of c-Myc and p53 protein and the activities of caspase-3, -6, and -8 were detected in Trp-P-1-perfused liver. In addition, Trp-P-1 modulated the DNA binding activity of the apoptosis-related transcription factors, NF-kappaB and AP-1. These results imply a possibility that Trp-P-1 would induce apoptosis in vivo.

Animals↗
PubMed2000

Trypsin inhibitory activity of bovine fetuin de-O-glycosylated by endo-alpha-N-acetylgalactosaminidase.

The effects of bovine fetuin O-glycans on its trypsin inhibitory activity were examined. De-sialylated (asialo-) and de-O-glycosylated fetuin were prepared from native fetuin using Arthrobacter neuraminidase and the mixture of it and Bacillus endo-alpha-N-acetylgalactosaminidase, respectively. De-sialylation and de-O-glycosylation from fetuin were confirmed with SDS-PAGE followed by western blotting using anti-human Thomsen-Friedenreich antigen (T antigen) antibody which recognizes O-linked galactosyl beta1,3 N-acetylgalactosamine (Gal beta1-->3GalNAc). Native fetuin completely inhibited the trypsin activity at about a 1:1 molar ratio. In contrast, the trypsin inhibitory activity of asialo- and de-O-glycosylated fetuin decreased about a half and one-third of that of native fetuin, respectively.

Animals↗
PubMed2000

Dietary antioxidants fail in protection against oxidative genetic damage in in vitro evaluation.

Carcinogenesis is believed to be induced through the oxidative damage of DNA, and antioxidants are expected to suppress it. So, the polyphenolic antioxidants in daily foods were investigated to see whether they protect against genetic damage by active oxygen. In the evaluation, we used a bioassay and a chemical determination, a Salmonella mutagenicity test for mutation by a N-hydroxyl radical from one of the dietary carcinogens 3-amino-1-methyl-5H-pyrido[4,3-b]indole and the formation of 8-hydroxyl (8-OHdG) from 2'-deoxyguanosine (2'-dG) in a Fenton OH-radical generating system. Thirty-one antioxidants including flavonoids were compared in terms of radical-trapping activity with bacterial DNA and 2'-dG. Antioxidants inhibited the mutation but the IC50 values were in the mM order. Against 8-OHdG formation, only alpha-tocopherol had a suppressive effect with an IC50 of 1.5 microM. Thus, except alpha-tocopherol, the dietary antioxidants did not scavenge the biological radicals faster than bacterial DNA and intact 2'-dG, indicating that they failed to prevent oxidative gene damage and probably carcinogenesis.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed2000

Malignant schwannoma metastasizing to the parenchyma of the brain--case report.

A 48-year-old male presented with a very rare case of malignant schwannoma metastasizing to the parenchyma of the brain. He had undergone previous radical surgical resection of an abdominal wall tumor that was histologically confirmed as a malignant schwannoma. Five years later, the patient presented with metastases to the parenchyma of the brain and lung. A large mass at the left frontoparietal region was totally resected immediately after intratumoral hemorrhage. He recovered well and the lung metastasis was also removed, but he later developed further multiple metastases in the lung and brain.

Brain Neoplasms↗
PubMed2000

Synergistic effects of food colors on the toxicity of 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1) in primary cultured rat hepatocytes.

The synergistic effect of food additives or food colors on the toxicity of 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1) was investigated using primary cultured rat hepatocytes. When hepatocytes from rats fed a standard diet were treated with a mixture of four major food additives (sorbitol, sodium L(+)-glutamate, benzoic acid, and propylene glycol) or a mixture of six typical artificial food colors (erythrosine, allura red, new coccine, brilliant blue, tertrazine, and fast green), the in vitro treated food-color mixture itself showed cytotoxicity: the reduction of cell viability and decreases in the activities of gluconeogenesis and ureogenesis. The food-color mixture enhanced cytotoxicity of Trp-P-1 obviously. We then investigated the effects of in vivo-dosed food additives or food colors on Trp-P-1-caused toxicity. Hepatocytes were isolated and cultured from rats fed a diet containing a mixture of food additives or a mixture of food colors with half the amount of their respective acceptable daily intake for 4 wk. Trp-P-1 was administered to the hepatocytes at various concentrations for 12 h. Synergistic effects of in vivo-dosed food additives and food colors were not observed on Trp-P-1-caused cytotoxicity as estimated by a loss of cell viability and the reductions of DNA and protein syntheses. On the contrary, we have observed that in vivo administered food colors synergistically facilitated to reduce the activities of gluconeogenesis and ureogenesis in Trp-P-1-treated hepatocytes. These results suggest that the daily intake of artificial food colors may impair hepatic functions such as gluconeogenesis and ureogenesis, when dietary carcinogens are exposed to the liver cells.

Animals↗
PubMed2000

[A case of spontaneous cervical carotid artery dissection managed by using primary stent placement].

The authors report the case of a 35-year-old male who underwent stenting for spontaneous cervical carotid dissection. He presented with sudden onset of hemicrania and left facial palsy followed by left hemiparesis and dysarthria. On admission, carotid angiography revealed postsinus tapering occlusion of the right internal carotid artery. Initially, he was managed with conservative treatment, but his neurological status deteriorated. Findings of brain CT, MRI and IMP-SPECT suggested hypoperfusion of the right cerebrum. A Palmaz stent, 39 mm in length, was successfully placed over the site of the dissection to restore normal patency through the dissected carotid artery. Following stent implantation, his neurological signs improved gradually but completely. Since the procedure, with oral administration of antiplatelet medication, he has suffered no cerebral ischemic events. Follow-up carotid angiography one year after stent implantation showed good patency of the stented segment. The present case emphasizes the usefulness of stenting for spontaneous cervical carotid dissection.

Adult↗