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T Akaike

Publications and source records attributed to T Akaike.

At least 145 records · Page 8Linked to original sources
PubMed1997

Activation of human neutrophil procollagenase by nitrogen dioxide and peroxynitrite: a novel mechanism for procollagenase activation involving nitric oxide.

The involvement of nitric oxide (NO) and its reactive intermediates such as nitrogen dioxide (NO2) and peroxynitrite (ONOO-) in the activation of matrix metallo-proteinase was investigated. The human neutrophil procollagenase (matrix metalloproteinase-8) (M(r), 85 kDa) was purified to homogeneity from human neutrophils by using column chromatography. After incubation of human neutrophil procollagenase with various nitrogen oxide-generating systems, collagenolytic activity in each reaction system was measured. In addition, neutrophil collagenase activity was determined by assessment of proteolysis of human alpha 1-protease inhibitor. NO was formed by the propylamine NONOate, and NO2 was generated by oxidation of NO with 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxide (carboxy-PTIO). NO2, formed by NONOate and carboxy-PTIO, and the synthetic ONOO- exhibited strong activation of the procollagenase at 1-20 microM. Significant activation of the procollagenase was observed with use of authentic NO2 gas as well. Constant flux infusion of ONOO- into the procollagenase solution resulted in stronger procollagenase activation than did a bolus addition of ONOO- to the reaction mixture. However, NO showed only weak activating potential under the aerobic (ambient) condition; an NO concentration of more than 10 mM was needed for appreciable activation of the procollagenase. Of considerable importance was the fact that NO participates in activation of the neutrophil collagenase through its conversion to NO2 or ONOO- in human neutrophils. These results suggest that NO2 and ONOO- may be potent activators of human neutrophil procollagenase.

Benzoates↗
PubMed1997

Prevention of experimental allergic encephalomyelitis by targeting nitric oxide and peroxynitrite: implications for the treatment of multiple sclerosis.

In this study we provide further evidence associating activated cells of the monocyte lineage with the lesions of multiple sclerosis (MS). Using a combination of immunohistochemistry and reverse transcriptase-dependent in situ polymerase chain reaction analysis, we have identified monocytes expressing inducible nitric oxide synthase (iNOS) to be prevalent in the plaque areas of post mortem brain tissue from patients with MS. In addition, we have obtained evidence of the nitration of tyrosine residues in brain areas local to accumulations of iNOS-positive cells. In parallel studies we have assessed the effects of inhibitors of iNOS induction, as well as scavengers of nitric oxide and peroxynitrite in the experimental allergic encephalomyelitis model. Significant therapeutic effects were seen with the inhibitor of iNOS induction, tricyclodecan-9-xyl-xanthogenate, a nitric oxide scavenger, 2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide, and a peroxynitrite scavenger, uric acid. In particular, treatment with high doses of uric acid virtually prevented clinical symptoms of the disease. Together with our demonstration of the presence of activated macrophages expressing high levels of iNOS and evidence of peroxynitrite formation in brain tissue from patients with MS, these findings are of importance in the development of approaches to treat this disease.

Animals↗
PubMed1997

Superoxide scavenging activity of erythromycin-iron complex.

We evaluated superoxide (O2-.) scavenging activity of erythromycin (EM) and of EM-iron complex by means of electron spin resonance spectroscopy, luminol-dependent chemiluminescence assay, and cytochrome c reduction assay. The EM-iron complex was produced by mixing EM with equal molar iron chloride and was stable in neutral buffer. The EM-iron complex reduced the amount of O2-. produced by xanthine oxidase/hypoxanthine without inhibiting the enzyme activity. It also reduced the amount of O2-. release from phorbor ester-stimulated human neutrophils and alveolar macrophages. EM alone showed few such effects. The scavenging activity of the complex was equal to that of L-ascorbic acid. These results in vitro suggest a possibility that the O2-.-scavenging effect of EM-iron complex contributes to the anti-inflammatory action of EM used in treating chronic inflammatory lung disease independent of its antimicrobial activity.

Erythromycin↗
PubMed1997

Activation of human matrix metalloproteinases by various bacterial proteinases.

Matrix metalloproteinases (MMPs) are zinc-containing proteinases that participate in tissue remodeling under physiological and pathological conditions. To test the involvement of bacterial proteinases in tissue injury during bacterial infections, we investigated the activation potential of various bacterial proteinases against precursors of MMPs (proMMPs) purified from human neutrophils (proMMP-8 and -9) and from human fibrosarcoma cells (proMMP-1). Each proMMP was subjected to treatment with a series of bacterial proteinases at molar ratios of 0.01-0.1 (bacterial proteinase to proMMP), and activities of MMPs generated were determined. Among six different bacterial proteinases, thermolysin family enzymes (family M4) such as Pseudomonas aeruginosa elastase, Vibrio cholerae proteinase, and thermolysin strongly activated all three proMMPs via limited proteolysis to generate active forms of the MMPs. N-terminal sequence analysis of the active MMPs revealed that cleavage occurred at the Val82-Leu83 and Thr90-Phe91 bonds of proMMP-1 and proMMP-9, respectively, which are located near the N terminus of the catalytic domain of MMPs. In contrast, Serratia 56-kDa proteinase and Pseudomonas alkaline proteinase, both of which are classified as members of the serralysin subfamily of zinc metalloproteinases (family M10), and Serratia 73-kDa thiol proteinase did not evidence proteolytic processing or activation of proMMP-1, -8, and -9 under these experimental conditions. These results indicate that bacterial proteinases may play an important role in tissue destruction and disintegration of extracellular matrix at the site of infections.

Amino Acid Sequence↗
PubMed1997

Generation of anaphylatoxins through proteolytic processing of C3 and C5 by house dust mite protease.

BACKGROUND: The group 3 allergen of Dermatophagoides species (Der p 3 and Der f 3) has been identified as a 30 kd trypsin-like protease of the house dust mite. We previously showed that the 30 kd protease from Dermatophagoides farinae (Df-protease) could activate the bradykinin-generating cascade and exacerbate inflammatory reactions. OBJECTIVE: The purpose of this study was to determine whether Df-protease could enzymatically generate anaphylatoxins from complement components C3 and C5. METHODS: Df-protease was incubated with human serum C3 or C5 in a purified system, and the anaphylatoxin activity produced was assayed by measuring enhancement of vascular permeability and release of histamine from mast cells triggered by C3a and by assessing chemotaxis of polymorphonuclear cells caused by C5a. We also attempted to determine whether protease isolated from house dust could cause release of C5a from C5. RESULTS: Df-protease showed strong activation of C3 and C5, producing C3a and C5a by proteolytic cleavage of the complements. An appreciable amount of Df-protease was recovered in the house dust extract, and the house dust protease caused C5a release from C5. CONCLUSION: Df-protease activated the complement system to produce anaphylatoxins. Thus it is suggested that house dust mite proteases may contribute to the pathogenesis of allergic and inflammatory diseases caused by house dust allergens.

Air Pollution, Indoor↗
PubMed1997

Simple preparation of nanoparticles coated with carbohydrate-carrying polymers.

Nanoparticles bearing carbohydrate chains on the surface can be prepared by the simple diafiltration method. The nanoparticles prepared by the present method displayed high yield, no-aggregation formation, small size, narrow size distribution, and one-step procedure. Also, the high density carbohydrate chains on the particles can be recognized by liver cells.

Animals↗
PubMed1997

Analysis of IFN-gamma-induced cell cycle arrest and cell death in hepatocytes.

The mechanism by which IFN-gamma induces cell cycle arrest and cell death in primary cultured hepatocytes was examined. The cell death exhibits apoptotic characters such as the appearance of apoptotic bodies and DNA fragmentation. IFN-gamma induced cell cycle arrest at the initial stage, followed by cell death. A protein synthesis inhibitor, cycloheximide, significantly inhibited cell death, implying that IFN-gamma induces de novo proteins involved in the death of hepatocytes. One of the most important apoptosis-related proteins, p53, was induced by IFN-gamma in hepatocytes in a dose- and time-dependent manner. Northern blot analysis demonstrated that IFN-gamma enhanced p53 mRNA expression as well as p21(WAF1/Cip1/Sdi1) mRNA expression, which is mediated by the increased expression of the p53 protein. Interestingly, IFN-gamma also induced cell death in p53-deficient hepatocytes. The cell death occurred rather earlier in p53-deficient cells than in normal hepatocytes. However, the cell death was not accompanied by apoptotic bodies. Therefore, IFN-gamma-induced hepatocyte cell death is p53-independent, and p53 may contribute to the apoptotic characters. In conclusion, IFN-gamma is supposed to cause cell cycle arrest by inducing p53 and p21(WAF1/Cip1/Sdi1), and it was demonstrated that IFN-gamma induces p53-independent cell death in primary cultured hepatocytes.

Animals↗
PubMed1997

Specific interaction between erythrocytes and a glucose-carrying polymer mediated by the type-1 glucose transporter (GLUT-1) on the cell membrane.

A reducing glucose-carrying polymer, called poly [3-O-(4'-vinylbenzyl)-D-glucose] (PVG), interacted with erythrocytes carrying the type-1 glucose transporter (GLUT-1) on the cell membrane. The cooperative interaction between a number of GLUT-1s and a number of reducing 3-O-methyl-D-glucose moieties on a PVG polymer chain is responsible for the increase in the interaction with erythrocytes. In contrast to the PVG homopolymer, other sugar-carrying polymers showed lower interaction with erythrocytes. The affinity of erythrocytes and PVG was studied using FITC-labeled glycopolymers. The fluorescence intensity significantly changed, whereas a small change in fluorescence intensity was observed for other homopolymers. The specific interaction between GLUT-1 on erythrocytes and the PVG polymer carrying reducing glucose was suppressed by the inhibitors, phloretin, phloridzin, and cytochalasin B, and a monoclonal antibody to GLUT-1. Direct observation by confocal laser microscopy with the use of FITC-labeled PVG demonstrated that erythrocytes interacted with the soluble form of the PVG polymer via GLUT-1, while fluorescence labeling of the cell surface was prevented on pretreatment with the monoclonal antibody to GLUT-1.

Animals↗
PubMed1997

Nanomolar quantification and identification of various nitrosothiols by high performance liquid chromatography coupled with flow reactors of metals and Griess reagent.

Nitrosothiols (RS-NOs) appear to be critically involved in various signal transduction mechanisms. We describe here a specific and highly sensitive quantification method for RS-NOs by using high performance liquid chromatography (HPLC) combined with a flow reactor system. RS-NOs were applied to an HPLC system of C18-reverse phase or a gel filtration column and eluted with 10 mM sodium acetate buffer (pH 5.5) plus 0.5 mM diethylenetriamine pentaacetic acid with or without either 0-7% methanol or 0.15 M NaCl. The eluate from the HPLC column was mixed with a solution containing 1.75 mM HgCl2 or 1.75 mM CuSO4 for RS-NO decomposition in a reaction coil via a three-way connector. NO2- generated via the metal-induced RS-NO decomposition was then reacted with Griess reagent, which was infused through a second three-way connector, yielding a diazo-compound detected at 540 nm. In a separate experiment, a copper particle-loaded column was used for RS-NO degradation instead of the metal-ion flow reactor. In all RS-NOs tested, i.e., nitrosoglutathione (GS-NO), nitroso-L-cysteine, and nitrosoalbumin, the nitroso- group was converted to NO2- by the Hg2+-reaction system as well as copper-loaded column, and the recovery was almost 100%. The Cu2+-solution flow reaction system, however, yielded only 30% recovery of RS-NOs as NO2-. Also, the RS-NOs could be identified at nanomolar concentrations: detection limit, 3.0 nM in a 150-microl aliquot. These RS-NOs showed well-resolved elution profiles even in the presence of NO2- and NO3-. More importantly, biological generation of GS-NO was quantitatively demonstrated with RAW264 cells in culture incorporating free GSH in the medium. In conclusion, our novel RS-NO assay will be useful to examine the formation and functions of RS-NOs in biological systems.

Animals↗
PubMed1997

Nitric oxide generation from hydroxyurea via copper-catalyzed peroxidation and implications for pharmacological actions of hydroxyurea.

We investigated the generation of nitric oxide (NO) by H2O2-dependent peroxidation of hydroxyurea in the presence of copper-containing proteins such as Cu,Zn-superoxide dismutase (Cu,Zn-SOD) or ceruloplasmin as a catalyst. In the reaction mixture of hydroxyurea, CuZn-SOD, and H2O2, NO generation was identified by measuring the specific electron spin resonance (ESR) signal of 2-phenyl-4, 4,5,5-tetramethylimidazoline-1-oxyl 3-oxide (PTIO). The ESR signal of the NO-hemoglobin adduct was also detected in human red blood cells during copper-catalyzed peroxidation of hydroxyurea. The NO production during peroxidation of hydroxyurea was quantified as NO2- formation, measured by using the Griess assay, the amount of NO2- was dependent on the concentrating of hydroxyurea of the reaction mixture. ESR spin trapping with 5,5-dimethyl-1-pyrroline N-oxide (DMPO) showed hydroxy radical (OH) generation in the reaction of H2O2 with either Cu,Zn-SOD or ceruloplasmin. Several OH scavengers, such as ethanol, thiourea, DMPO, and dimethylsulfoxide, and the metalchelating agent diethylenetriaminepentaacetic acid significantly inhibited NO generation from hydroxyurea. This indicates that NO release from hydroxyurea may be mediated by OH derived from the copper-catalyzed Fenton-like reaction. Incubation of hydroxyurea and Cu,Zn-SOD with xanthine oxidase and hypoxanthine in a system forming O2- -->H2O2 also resulted in appreciable NO production. These results suggest that NO production from hydroxyurea catalyzed by copper-containing proteins may be the molecular basis of the pharmacological and antitumor action of hydroxyurea.

Ceruloplasmin↗
PubMed1997

Induction of nitric oxide synthesis and xanthine oxidase and their roles in the antimicrobial mechanism against Salmonella typhimurium infection in mice.

The role of superoxide anion (O2-) and nitric oxide (NO) in the host defense mechanism against Salmonella typhimurium (LT-2) was examined by focusing on xanthine oxidase (XO) as an O2(-)-generating system and on inducible NO synthase (iNOS). When ICR mice were infected with a 0.1 50% lethal dose (2 x 10(5) CFU) of S. typhimurium, bacterial growth in the liver reached a peak value 3 days after infection (10(4.32) CFU/g of liver) and decreased thereafter. XO activity in the liver became maximum at 7 days after infection; the value was 34.6 +/- 1.4 mU/g of liver at 7 days (compared with 11.0 +/- 1.3 mU/g of liver before infection). The time profile of NO production in the liver as determined by electron spin resonance spectroscopy was consistent with that of XO activity. Histological examination of infected liver showed the formation of multiple microabscesses with granulomatous lesions consisting of polymorphonuclear cells and mononuclear cells, and iNOS-expressing cells were localized in the confined areas of the microabscesses. When XO inhibitors such as allopurinol and 4-amino-6-hydroxypyrazolo[3,4-d]pyrimidine (AHPP) were administered to the infected mice, the mortality of the mice was significantly increased (10 of 21 and 11 of 20 for the allopurinol- and AHPP-treated groups, respectively, versus 2 of 20 for control mice), and bacterial growth was significantly enhanced. A similar exacerbation of the infection was obtained with N(omega)-monomethyl-L-arginine (L-NMMA) treatment of the mice. Of considerable importance is that granuloma formation in the liver was poorly developed by treatment with either XO inhibitors or L-NMMA. These results suggest that XO and NO play an important role in the antimicrobial mechanism against S. typhimurium in mice.

Allopurinol↗
PubMed1997

Development of a fluorescent indicator for the bioimaging of nitric oxide.

Nitric oxide (NO) has been reported to play various roles as a signal transmitter. However, detailed functions of NO have yet to be clarified. We have developed a fluorescent indicator for NO imaging in living cells. The N-nitrosation of newly designed and synthesized 4-((3-amino-2-naphthyl)aminomethyl)benzoic acid (DAN-1) by NO yielded the highly fluorescent triazole-form. The membrane permeable ester derivative of DAN-1 (DAN-1 EE) was applied to the imaging of NO produced in activated rat aortic smooth muscle cells. After DAN-1 EE has been loaded into cells, the ester bond is hydrolyzed by intracellular esterase, yielding original DAN-1 with less permeability. The fluorescence intensity of the cells loaded with DAN-1 EE increased according to NO production. The imaging method with fluorescent indicators will be significant for the functional clarification of NO in vivo.

2-Naphthylamine↗
PubMed1997

Synthesis of monoesters of pyrroloquinoline quinone and imidazopyrroloquinoline, and radical scavenging activities using electron spin resonance in vitro and pharmacological activity in vivo.

Monoesters with the ester groups at C-2 of pyrroloquinoline quinone (PQQ) and C-9 of imidazopyrroloquinoline (IPQ) were synthesized, and radical scavenging activities of coenzyme PQQ, IPQ compounds synthesized from PQQ and various amino acids, and monoesters of PQQ and IPQ were studied in vitro and in vivo. PQQ and PQQ monoesters had strong radical scavenging activity using ESR in in vitro experiments. The IC50 value for superoxide (O2-) was from 1 to 6 x 10(-8) M and that for the hydroxy radical (.OH) was from 4 to 6 x 10(-5) M. IPQ compounds and IPQ monoesters also showed radical scavenging activity. These compounds prevented injury during in vivo experiments, such as hydrocortisone-induced cataracts, endotoxin shock and CCl4-induced liver injury (isolated hepatocytes and rats). Especially, the monoesters of PQQ and IPQ prevented liver injury in rats equally by oral or intraperitoneal administration. These results suggest that PQQ functions as a radical scavenging factor in addition to being a cofactor of quinoprotein enzymes, and monoesters with the ester groups at C-2 of PQQ and C-9 of IPQ are developed as treatment or preventive medicine for disease caused by radical compounds on the basis of strong radical scavenging activities, absorbability into cells, toxicity, safety and chemical stability.

Administration, Oral↗
PubMed1997

Poly(L-lysine)-graft-dextran copolymer is a novel stabilizer of triplex DNA (I): stabilization of poly(dA).2poly(dT) triplex.

Comb-type polylysine copolymer having grafted hydrophilic side chains was newly designed as a novel stabilizer of triplex DNAs. The comb-type copolymer elevated melting temperature of poly(dA).2poly(dT) triplex by 50 degrees C without affecting reversibility, melting and reassociation, of the triplex in buffer with physiological salt concentrations. The stabilizing effect of the copolymer was greater than spermine. Our results indicate that the molecular designing of polycation with comb-type structure is a successful strategy for creating an effective triplex stabilizer.

Biocompatible Materials↗
PubMed1997

Poly(L-lysine)-graft-dextran copolymer is a novel stabilizer of triplex DNA(II): potassium-insensitive triplex formation.

Triple helix formation involving guanine-rich oligonucleotides is inhibited by physiological concentrations of potassium (K+). Using in vitro electrophoretic mobility shift assay (EMSA), we show that PLL-g-dex copolymer significantly stabilizes triplex structure at physiological levels of K+ that greatly inhibited triplex formation with a unmodified, guanine-rich oligonucleotide.

Base Sequence↗
PubMed1996

Excessive production of nitric oxide in rat solid tumor and its implication in rapid tumor growth.

BACKGROUND: Rapid tumor growth is caused by angiogenesis factors, growth factors, etc. We previously reported a possible connection between nitric oxide (NO) and enhanced vascular permeability in solid tumor. In the present experiment, the role of NO in solid tumor pathology was further investigated in animal tumor. METHODS: To identify NO formed in solid tumor (AH136B) implanted in the feet of rats, electron paramagnetic resonance (EPR) spectroscopy was performed directly on the frozen tumor tissue at 110K by measuring endogenous nitrosyl iron-sulfur complexes, and by using exogenously added NO capturing agents, i.e., diethyldithiocarbamate (DETC)-Fe2+ and N-(dithiocarboxy) sarcosine (DTCS)-Fe2+ complexes. Induction of inducible isoform of nosymthase iNOS mRNA was examined with reverse transcriptase-polymerase chain reaction (RT-PCR) combined with Southern blot analysis. In addition, vascular permeability was assessed by measuring extravasation of 51Cr-labeled bovine serum albumin in solid tumor. RESULTS: Strong EPR signals from NO adducts of DETC-Fe2+ and DTCS-Fe2+ as well as strong signals from NO-hemoglobin and dinitrosyl iron sulfur complex were generated by tumor. The signal height of NO-(DTCS)2-Fe2+ observed in AH136B solid tumor was increased as the tumor gained up to 1.75 g. Induction of iNOS mRNA expression was confirmed by the above methods. Enhanced vascular permeability was suppressed by NOS inhibitors N omega- monomethyl-L-arginine or S-methylisothiourea sulfate and augmented with administration of L-arginine. CONCLUSIONS: Excessive NO production by iNOS in solid tumor was identified unequivocally by EPR spectroscopy. NO formed in solid tumor can be involved in enhanced vascular permeability and increased blood flow, and hence sustain tumor growth.

Animals↗
PubMed1996

Pathogenesis of influenza virus-induced pneumonia: involvement of both nitric oxide and oxygen radicals.

The role of nitric oxide (NO) in the pathogenesis of influenza virus-induced pneumonia in mice was investigated. Experimental influenza virus pneumonia was produced with influenza virus A/Kumamoto/Y5/67(H2N2). Both the enzyme activity of NO synthase (NOS) and mRNA expression of the inducible NOS were greatly increased in the mouse lungs; increases were mediated by interferon gamma. Excessive production of NO in the virus-infected lung was studied further by using electron spin resonance (ESR) spectroscopy. In vivo spin trapping with dithiocarbamate-iron complexes indicated that a significant amount of NO was generated in the virus-infected lung. Furthermore, an NO-hemoglobin ESR signal appeared in the virus-infected lung, and formation of NO-hemoglobin was significantly increased by treatment with superoxide dismutase and was inhibited by N(omega)-monomethyl-L-arginine (L-NMMA) administration. Immunohistochemistry with a specific anti-nitrotyrosine antibody showed intense staining of alveolar phagocytic cells such as macrophages and neutrophils and of intraalveolar exudate in the virus-infected lung. These results strongly suggest formation of peroxynitrite in the lung through the reaction of NO with O2-, which is generated by alveolar phagocytic cells and xanthine oxidase. In addition, administration of L-NMMA resulted in significant improvement in the survival rate of virus-infected mice without appreciable suppression of their antiviral defenses. On the basis of these data, we conclude that NO together with O2- which forms more reactive peroxynitrite may be the most important pathogenic factors in influenza virus-induced pneumonia in mice.

Animals↗