Search PubMed⌕ Search

Biomedical subjects

Y Nishikawa

Publications and source records attributed to Y Nishikawa.

At least 73 records · Page 4Linked to original sources
PubMed1999

Inhibition of hepatoma cell growth in vitro by arylating and non-arylating K vitamin analogs. Significance of protein tyrosine phosphatase inhibition.

We recently found that a thioether analog of K vitamin (Cpd 5) inhibited the activity of protein-tyrosine phosphatases (PTPases) and induced protein-tyrosine phosphorylation in a human hepatoma cell line (Hep3B). We have now examined the structural requirements for induction of protein-tyrosine phosphorylation and PTPase inhibition by several K vitamin analogs. Thioether analogs with sulfhydryl arylation capacity, especially those with a hydroxy (Cpd 5) or a methoxy group at the end of the side chain, induced protein-tyrosine phosphorylation, but non-arylating analogs, such as those with an all-carbon or O-ether side chain, did not. Among the receptor-tyrosine kinases, epidermal growth factor receptors were tyrosine-phosphorylated by treatment with thioether analogs, whereas insulin and hepatocyte growth factor receptors were not. An increase in tyrosine-phosphorylated ERK2 mitogen-activated protein kinase was also observed. The activity of purified T cell PTPase was inhibited only by the thioether analogs, but not by non-arylating analogs. Furthermore, the epidermal growth factor receptor dephosphorylation activity of Hep3B cell lysates was inhibited by Cpd 5 treatment. A similar induction of protein-tyrosine phosphorylation by Cpd 5 was seen in other human hepatoma cell lines together with growth inhibition. However, one cell line (HepG2), which was relatively resistant to growth inhibition by Cpd 5, did not increase its phosphorylation levels upon Cpd 5 treatment. These results suggest that cell growth inhibition by thioether analogs is closely associated with inhibition of PTPases by sulfhydryl arylation and with tyrosine phosphorylation of selected proteins.

Carcinoma, Hepatocellular↗
PubMed1999

Regulation of shear stress in the canine coronary microcirculation.

BACKGROUND: Physical forces, such as pressure and flow, are well known to affect vascular function in the coronary circulation. Increases in shear stress produce vasodilation in coronary arterioles in vitro, and constant-flow preparations suggest a role for shear stress-induced vasodilation during adjustments to metabolic demand in vivo. Hypothetically, the regulation of shear stress can be viewed as a negative feedback control scheme (increased velocity --> increased shear --> vasodilation --> decreased velocity --> shear normalized). Therefore, we hypothesized that shear stress would be at least partially regulated during conditions of elevated flow. METHODS AND RESULTS: We used fluorescence microangiography to measure microvascular diameters and velocities in the coronary circulation in vivo and used these variables to calculate shear stress. Measurements were obtained under basal conditions, during maximal coronary blood flow, and after inhibition of NO synthase. Basal shear stress in the coronary circulation averaged 10 dyn/cm2 in small arteries and 19 dyn/cm2 in arterioles. Regulation of shear stress was observed in small arteries during adenosine-induced increases in coronary blood flow, but arterioles showed minimal regulation. NO synthase blockade had no effect on basal shear stress but completely abolished its regulation in small arteries during vasodilation. CONCLUSIONS: Our data provide the first quantitative estimates of microvascular shear stress in the coronary circulation. Moreover, our results suggest that shear stress in small coronary arteries is regulated by NO release from the endothelium.

Adenosine↗
PubMed1999

Activation of ascending antinociceptive system by vagal afferent input as revealed in the nucleus ventralis posteromedialis.

Thalamic nociceptive neurons receiving afferent input from the tooth pulp (TP) were recorded from the nucleus ventralis posteromedialis proper (VPM) in cats anesthetized with urethane and chloralose. Effects of cervical vagus nerve stimulation on responses of TP neurons in the VPM were investigated. Twenty-one tooth pulp specific (TPS) and eight wide dynamic range (WDR) neurons with TP input were obtained from the periphery (shell region) of the posterior half of the VPM. Of these, many were also excited by electrical stimulation of trigeminothalamic tract (TTT) fibers in the trigeminal medial lemniscus. A conditioning-test paradigm was used to examine effects of vagal stimulation on responses of VPM neurons to electrical stimulation of TP and TTT. Inhibition of the responses was observed in 12 TPS and seven WDR neurons. Local anesthetic block of the mesencephalic periaqueductal gray (PAG) and/or nucleus raphe dorsalis (NRD) eliminated the inhibitory effects of vagal stimulation on the responses of both classes of TP neurons to TTT stimulation. In contrast, the inhibitory effects on responses to TP stimulation were insignificantly affected. These data suggest that vagal afferents can activate the ascending antinociceptive pathway from PAG/NRD onto VPM, in addition to activating the descending antinociceptive system acting upon the lower brain stem.

Afferent Pathways↗
PubMed1999

Application of region-specific immunoassay for human chromogranin A: substantial clue for detection and measurement of chromogranin A in human plasma.

Chromogranin A (CgA), a secretory protein, is co-released with catecholamines from storage vesicles. It is known to be elevated in the circulation of patients with neuroendocrine and endocrine tumors. For further investigation of the protein, especially in humans, it is essential to facilitate quantitative analysis of the protein in human biological materials. In order to introduce novel immunological methodology for this purpose, we purposely selected human CgA(344-374) for the synthetic immunogen to produce region-specific CgA antibodies. The anti-synthetic peptide antibody thus obtained made it possible to develop an immunological method for measurement and characterization of CgA in human plasma. The plasma CgA-immunoreactivity (LI) level measured by the method was 0.31+/-0.01 pmol/ml (mean+/-SEM) in normal subjects and 1.55+/-0.29 pmol/ml in pheochromocytoma. On gel chromatography and HPLC analysis of the plasma of patients with pheochromocytoma, the region-specific assay system enabled us to show the presence of N-terminal truncated CgA, besides CgA itself. By following up changes of plasma CgA-LI in a pheochromocytoma patient using samples that were collected consecutively over a two-year period, the present assay system using the region-specific antibody, anti-human CgA (344-374) serum, was confirmed to be extremely valuable for the measurement of CgA-LI in human plasma. The characteristic features and high sensitivity of the present assay system will give us a substantial clue to the detection and measurement of CgA to develop further investigation of the protein in humans.

Adrenal Gland Neoplasms↗
PubMed1999

The alkaline single cell gel electrophoresis assay with mouse multiple organs: results with 30 aromatic amines evaluated by the IARC and U.S. NTP.

The genotoxicity of 30 aromatic amines selected from IARC (International Agency for Research on Cancer) groups 1, 2A, 2B and 3 and from the U.S. NTP (National Toxicology Program) carcinogenicity database were evaluated using the alkaline single cell gel electrophoresis (SCG) (Comet) assay in mouse organs. We treated groups of four mice once orally at the maximum tolerated dose (MTD) and sampled stomach, colon, liver, kidney, bladder, lung, brain, and bone marrow 3, 8 and 24 h after treatment. For the 20 aromatic amines that are rodent carcinogens, the assay was positive in at least one organ, suggesting a high predictive ability for the assay. For most of the SCG-positive aromatic amines, the organs exhibiting increased levels of DNA damage were not necessarily the target organs for carcinogenicity. It was rare, in contrast, for the target organs not to show DNA damage. Organ-specific genotoxicity, therefore, is necessary but not sufficient for the prediction of organ-specific carcinogenicity. For the 10 non-carcinogenic aromatic amines (eight were Ames test-positive and two were Ames test-negative), the assay was negative in all organs studied. In the safety evaluation of chemicals, it is important to demonstrate that Ames test-positive agents are not genotoxic in vivo. Chemical carcinogens can be classified as genotoxic (Ames test-positive) and putative non-genotoxic (Ames test-negative) carcinogens. The alkaline SCG assay, which detects DNA lesions, is not suitable for identifying non-genotoxic carcinogens. The present SCG study revealed a high positive response ratio for rodent genotoxic carcinogens and a high negative response ratio for rodent genotoxic non-carcinogens. These results suggest that the alkaline SCG assay can be usefully used to evaluate the in vivo genotoxicity of chemicals in multiple organs, providing for a good assessment of potential carcinogenicity.

Amines↗
PubMed1999

Purification and cloning of a thermostable manganese catalase from a thermophilic bacterium.

We have purified a heat-stable catalase from a thermophilic bacterium, Thermus species strain YS 8-13. The enzyme was purified 160-fold from crude cellular extracts and possessed a specific activity of 8000 units/mg at 65 degrees C. The purified enzyme displayed the highest activity at pH 7 to 10 and temperatures around 85 degrees C. The catalase was determined to be a manganese catalase, based on results from atomic absorption spectra and inhibition experiments using sodium azide. The enzyme was composed of six identical subunits of molecular weight 36,000. Amino acid sequences determined from the purified protein were used to design oligonucleotide primers, which were in turn used to clone the coding gene. The nucleotide sequence of a 1.4-kb fragment of Thermus sp. YS 8-13 genomic DNA containing a 909-bp open reading frame was determined. The gene encoded a 302-residue polypeptide of deduced molecular weight 33,303. The deduced amino acid sequence displayed a region-specific homology with the sequences of the manganese catalase from a mesophilic organism, Lactobacillus plantarum.

Amino Acid Sequence↗
PubMed1999

Evaluation of endoscopic injection sclerotherapy with and without simultaneous ligation for the treatment of esophageal varices.

For more effective and simple endoscopic injection sclerotherapy (EIS) for esophageal varices, we developed an EIS procedure with ligation (EISL) that is non-invasive, in which EIS and endoscopic variceal ligation (EVL) are performed simultaneously. In this study, we compared EISL and EIS in a randomized sample of patients (n = 14 for each procedure). For EISL, EVL was performed, including the injection site, after the injection of 5% ethanolamine oleate with iopamidol (EOI) into a varix. The mean number of treatment sessions required for eradication of esophageal varices was 2.3+/-0.5 for EISL and 3.9+/-0.8 for EIS (P < 0.001); the mean number of treatment sites was 6.2+/-2.2 for EISL and 14.0+/-5.0 for EIS (P < 0.001); the mean total amount of EOI used was 13.8+/-5.2ml for EISL and 26.3+/-9.8ml for EIS (P < 0.001). There were no significant differences in rates of recurrence of varices or in bleeding between the two groups. For EISL, fewer treatment sessions and less sclerosant were sufficient, probably because the sclerosants were more effective due to the blockage of variceal blood flow by the ligation. This method should provide a novel modification of EIS.

Endoscopy, Digestive System↗
PubMed1999

Biosynthesis and interaction of glycoproteins E and I of canine herpesvirus.

In cells infected with canine herpesvirus (CHV), the mature form of glycoprotein E (gE) had a molecular weight of 94 kDa, and that of glycoprotein I (gI) had a broad range of molecular weights of 55-62 kDa. gE and gI formed a complex like gE and gI of other alphaherpesviruses. When cells were infected with the gI minus mutant of CHV (gI/Z), the mature form of the 94 kDa gE was not formed, but a 76 kDa gE polypeptide was found. Similarly, no mature gI was formed in cells infected with the gE minus mutant of CHV (gE/Z), but a 40 kDa gI polypeptide was formed. When cells were coinfected with gE/Z and gI/Z, the molecular masses of gE and gI were increased from 76 to 94 kDa and from 40 to 55-62 kDa, respectively. We constructed vaccinia virus recombinants which expressed CHV gE or CHV gI. Only when cells were coinfected with both the vaccinia recombinant which expressed gE and the vaccinia recombinant which expressed gI, gE and gI were processed into their mature forms. Our results suggest that the presence of both gE and gI is necessary for efficient processing of the precursors of gE and gI to their mature forms.

Animals↗
PubMed1999

Characterization of an alg2 mutant of the zygomycete fungus Rhizomucor pusillus.

The zygomycete fungus Rhizomucor pusillus secretes an aspartic proteinase (MPP) that contains asparagine ( N )-linked oligosaccharides at two sites. Mutant strain 1116 defective in N -glycosylation secretes MPP with truncated oligo-saccharide chains. Lipid-linked oligosaccharides in mutant 1116 were labeled with [6-(3)H]glucosamine and [2-(3)H]mannose, prepared by cycles of solvent extraction, and analyzed by gel filtration chromatography on a Bio-Gel P-4 column after mild acid-hydrolysis. Mutant 1116 accumulated an intermediate, Man(1)GlcNAc(2)-dolichol pyrophosphate (PP-Dol), whereas wild-type strain F27 synthesized the fully assembled oligosaccharide precursor Glc(3)Man(9)GlcNAc(2)-PP-Dol. Consistent with this, alg2 encoding a mannosyltransferase in the lipid-linked oligosaccharide biosynthetic pathway in mutant 1116 had a 5 bp insertion that generated a stop codon in the middle of the coding sequence. Transformation of mutant 1116 with the intact alg2 gene on a pUC19-derived plasmid generated transformants that contained multicopies of alg2 at the alg2 locus. Glycosylation of the total proteins in the transformants was recovered to the same level as in strain F27, as determined with peroxidase-concanavalin A. These transformants produced MPP mainly with the same N -linked oligosaccharides as that produced by strain F27, but still with truncated oligosaccharides in small amounts. All of these data show that Alg2 is an alpha-1,3 or alpha-1,6 mannosyltransferase that elongates Man(1)GlcNAc(2)-PP-Dol to Man(2)GlcNAc(2)-PP-Dol. The slower growth of mutant 1116 was significantly recovered on introduction of alg2. The viability of the alg2 mutants of the zygomycete R.pusillus makes a contrast with the lethal effect of ALG2 mutations in the yeast Saccharomyces cerevisiae.

Amino Acid Sequence↗
PubMed1999

Phase separation with multiple length scales in polymer mixtures induced by autocatalytic reactions.

A ternary polymer blend with two components photo-cross-linked independently in its miscible region undergoes phase separation, exhibiting morphology with multiple length scales. Contrary to the case of thermally induced phase separation, the morphology exhibits a unimodal-->multimodal transition. It is shown that these multiple length scales are caused by the inhomogeneous freezing kinetics of the cross-linking process. This inhomogeneity arises from the autocatalytic feedback driven by the couplings between concentration fluctuations and the photo-cross-linking reactions.

Journal Article↗
PubMed1999

Development and distribution of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP)-induced aberrant crypt foci in the rat large intestine.

Aberrant crypt foci (ACF) are generally considered to be preneoplastic lesions for colon cancer. To assess their induction by 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), a colon carcinogen, we performed a sequential study of ACF morphology and localization. F344 male rats were given PhIP, and methylene blue-stained colon epithelium and isolated crypts were analyzed at weeks 12, 25, 50, and 75. Each crypt was classified into 2 groups, "single" with round bottoms and "bifurcating" displaying V-shaped clefts (indicating proliferation). In combination with the number of crypts in an ACF, this classification was a good indicator for the generation of ACF in line with the fission mechanism of growth. Increasing numbers of crypts in ACF through weeks 12 to 75 and decreased percentages of ACF with bifurcating crypts at the late time points indicated that proliferation of crypts occurs predominantly during the early stages. The distribution pattern showed a significant shift (P < 0.000005) from the distal to the proximal part of the large intestine between weeks 25 and 50. Adenocarcinomas were first found to develop at week 50 in the ascending colon and cecum where bifurcating crypts were generally lacking at weeks 12 and 25. These data suggest the existence of (1) proliferating ACF which contains bifurcating crypt(s) and (2) quiescent or senescent ACF which consists of only single crypts.

Animals↗
PubMed1999

Endothelin antagonists block alpha1-adrenergic constriction of coronary arterioles.

We have previously observed that intracoronary administration of the alpha1-adrenergic agonist phenylephrine (PE) over a period of minutes induced both an immediate and long-lasting (2 h) vasoconstriction of epicardial coronary arterioles. Because it is unlikely that alpha1-adrenergic constriction would persist for hours after removal of the agonist, this observation supports the view that another constrictor(s) is released during alpha1-adrenergic activation and induces the prolonged vasoconstriction. Therefore, we hypothesized that the prolonged microvascular constriction after PE is due to the production of endothelin (ET). We focused on ET not only because this peptide produces potent vasoconstriction but also because its vasoconstrictor action is characterized by a long duration. To test this hypothesis, the diameters of coronary arterioles (<222 micrometers) in the beating heart of pentobarbital-anesthetized dogs with stroboscopic intravital microscopy were measured during a 15-min intracoronary infusion of PE (1 microgram. kg-1 . min-1) and at 15-min intervals for a total of 120 min. All experiments were performed in the presence of beta-adrenergic blockade with propranolol. At 120 min, arterioles in the PE group were constricted (-23 +/- 9% change in diameter vs. baseline). Pretreatment with the ET-converting enzyme inhibitor phosphoramidon or the ETA-receptor antagonist FR-139317 prevented the PE-induced constriction at 120 min (-1 +/- 3 and -6 +/- 3%, respectively, P < 0.01 vs. PE). Pretreatment with the selective alpha1-adrenergic antagonist prazosin (Prz) also prevented the sustained constriction (0 +/- 2%, P < 0.01 vs. PE) but Prz given 60 min after PE infusion did not (-13 +/- 3%). In the aggregate, these results show that vasoconstriction of epicardial coronary arterioles via alpha1-adrenergic activation is blocked by an ET antagonist and an inhibitor of its production. From these data, we conclude that alpha1-adrenergic activation promotes the production and/or release of ET, which produces or facilitates microvascular constriction of epicardial canine coronary arterioles.

Adrenergic alpha-Agonists↗
PubMed1999

In vivo location and mechanism of EDHF-mediated vasodilation in canine coronary microcirculation.

Responses of epicardial coronary arterioles to ACh were measured using stroboscopic fluorescence microangiography in dogs (n = 38). ACh (0.1 and 0.5 microg. kg(-1). min(-1) ic) dilated small (<100 micron, 11 +/- 2 and 19 +/- 2%, respectively) and large (>100 micron, , 6 +/- 3 and 13 +/- 3%, respectively) arterioles at baseline. Combined administration of N(omega)-monomethyl-L-arginine (L-NMMA; 1. 0 micromol/min ic) and indomethacin (10 mg/kg iv) eliminated ACh-induced dilation in large coronary arterioles but only partially attenuated that in small arterioles. Suffusion of a buffer containing 60 mM KCl (high KCl) completely abolished cromakalim-induced dilation in arterioles and in combination with L-NMMA plus indomethacin completely blocked ACh-induced dilation in small arterioles. This indicated that the vasodilation to ACh that persists in small arterioles after administration of L-NMMA and indomethacin is mediated via a hyperpolarizing factor. The ACh-induced vasodilation remaining after L-NMMA and indomethacin was completely blocked by the large-conductance potassium-channel antagonist iberiotoxin or by epicardial suffusion of miconazole or metyrapone, inhibitors of cytochrome P-450 enzymes. These observations are consistent with the view that endothelium-derived hyperpolarizing factor (EDHF) is a product of cytochrome P-450 enzymes and produces vasodilation by the opening of large-conductance potassium channels. We conclude that ACh-induced dilation in large coronary arterioles is mediated mainly by nitric oxide (NO), whereas, in small arterioles both NO and EDHF mediate dilation to ACh. These data provide the first direct evidence for an in vivo role of EDHF in small coronary arterioles.

Acetylcholine↗
PubMed1999

Roles of two basic nitrogen atoms in 1-substituted 4-(1,2-diphenylethyl)piperazine derivatives in production of opioid agonist and antagonist activities.

To ascertain roles of the two basic nitrogen atoms in 1-substituted 4-[2,(3-hydroxyphenyl)-1-phenylethyl]-piperazine derivatives (1) in the expression of opioid agonist and antagonist activities, a methine group (CH) was isosterically substituted for nitrogen atom at the 1-position (N-1) in compound 1 to obtain 4-substituted 1-[2-(3-hydroxyphenyl)-1-phenylethyl]piperidine derivatives (2). Their analgesic action and ability to produce physical dependence (jump-producing activity) as the mu-opioid receptor specific in vivo actions, and narcotic antagonist action in mice were compared with those of compound 1. Results of this study showed that, in cases of the racemate and the (S)-(+) enantiomer, opioid agonist activities (analgesic and jump-producing activities) were not greatly affected by the methine-substitution for N-1 in compound 1, but that the narcotic antagonist activity of the (R)-(-) enatiomer was abolished by this substitution. It thus appears that N-1 in compound 1 contributes to the expression of narcotic antagonist activity, whereas the nitrogen atom at the 4-position corresponds to the tyramine moiety necessary for the expression of mu-opioid agonist activity.

Animals↗
PubMed1999

Characterization of pseudorabies virus glycoprotein B expressed by canine herpesvirus.

A recombinant canine herpesvirus (CHV) which expressed glycoprotein B (gB) of pseudorabies virus (PrV) was constructed. The antigenicity of the PrV gB expressed by the recombinant CHV is similar to that of the native PrV. The expressed PrV gB was shown to be transported to the surface of infected cells as judged by an indirected immunofluorescence test. Antibodies raised in mice immunized with the recombinant CHV neutralized the infectivity of PrV in vitro. It is known that the authentic PrV gB exists as a glycoprotein complex, which consists of gBa, gBb and gBc. In MDCK cells, PrV gB expressed by the recombinant CHV was processed like authentic PrV gB, suggesting that the cleavage mechanism of PrV gB depends on a functional cleavage domain from PrV gB gene and protease from infected cells.

Animals↗
PubMed1999

Salivary secretion of highly concentrated chromogranin a in response to noradrenaline and acetylcholine in isolated and perfused rat submandibular glands.

Chromogranin A (CgA) is a member of a family of highly acidic proteins, chromogranins, which are co-stored in the adrenergic neurons and paraneurons and co-released with adrenaline and noradrenaline (NAd) in response to adequate stimulation. The present study provides novel evidence that CgA-like immunoreactivity (IR) is stored in the exocrine cells in the granular convoluted tubule, and is secreted into saliva by stimulation with NAd and acetylcholine (ACh) in the isolated and perfused rat submandibular gland. NAd at 1 microM produced maximum secretion of CgA-like IR (<< 0.9 mM) and a marked increase in salivary flow. Further increases in NAd concentration (10 or 100 microM) yielded concentration-dependent decreases in both responses. ACh at 1 microM produced maximum salivary flow and a slight elevation of CgA-like IR secretion (6 microM); 100 microM ACh decreased the salivary flow but increased the CgA-like IR secretion (0.6 mM). Electron microscopic examination showed vigorous compound exocytosis of secretory granules in the cells of the granular convoluted tubule when the submandibular gland was stimulated with 1 microM NAd. These results provide an experimental basis for the view that the salivary CgA-like IR secretion may be a sensitive and quantitative index of the activity of the sympathetic nervous system innervating the gland.

Acetylcholine↗
PubMed1999

Serum soluble Fas antigen in gastric MALT (mucosa-associated lymphoid tissue) lymphoma patients.

We measured the serum level of soluble Fas (sFas) antigen in gastric MALT (mucosa-associated lymphoid tissue) lymphoma patients to investigate immune response of B lymphoma cells in gastric MALT lymphoma. The serum sFas antigen levels in 21 patients with Helicobacter pylori (HP)-positive MALT lymphoma were 4.51+/-0.25 (mean+/-SD) in the high grade malignant group (n=9) that demonstrated diffuse large cell (DL) components and 2.53+/-0.46 (mean+/-SD) in the low grade malignant group (n=12) that did not demonstrate DL components. Among 12 patients who underwent HP eradication therapy, the blood sFas antigen level lowered after eradication in 7 out of 10 patients who showed complete response or partial response, while sFas antigen level did not decrease in the 2 patients who showed no change or progression of the disease. In the high grade malignant group, 2 of 3 patients who underwent HP eradication treatment showed a mild decrease in the sFas antigen level as well as an improvement on endoscopic and histological examinations, but these sFas level were not normalized. The blood sFas antigen level tended to coincide with pathological activity and malignancy, suggesting that the sFas antigen level may be a prognostic indicator and useful index for the response of HP eradication treatment in gastric MALT lymphoma.

Apoptosis↗