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Biomedical subjects

N Muto

Publications and source records attributed to N Muto.

At least 73 records · Page 4Linked to original sources
PubMed1990

Enzymatic formation of a nonreducing L-ascorbic acid alpha-glucoside: purification and properties of alpha-glucosidases catalyzing site-specific transglucosylation from rat small intestine.

We have previously found that some mammalian tissue homogenates can catalyze a unique transglucosylation from maltose to L-ascorbic acid (AA), resulting in a chemically stable AA derivative, L-ascorbic acid alpha-glucoside (AAG). In the present study, the enzyme responsible for this transglucosylation was isolated from rat intestinal membrane. The formation of AAG was determined by HPLC with an ODS column. The specific activity of AAG-forming enzyme was increased in parallel with that of alpha-glucosidase (maltose hydrolase) during the purification, and two neutral alpha-glucosidases, termed alpha-glucosidases I and II, were purified to apparent homogeneity. Their enzymological properties showed that they corresponded to maltase [EC 3.2.1.20] and sucrase-isomaltase complex [EC 3.2.1.48/10], respectively. Both enzymes could form AAG by splitting only maltose among the disaccharides examined, although alpha-glucosidase I possessed a considerably higher activity than the other enzyme. Both AAG formation and maltose hydrolysis were dependent on incubation temperature with the maximal activity at 60 degrees C, but there was an apparent difference between their pH optima. AAG thus formed could also be hydrolyzed by the purified enzymes. From these results, it is concluded that membrane-bound neutral alpha-glucosidases from rat intestine have site-specific transglucosylase activity to form nonreducing AAG which is distinct from L-ascorbic acid-6-O-alpha-D-glucoside.

Animals↗
PubMed1990

Prognosticating study for cervical myelopathy using evoked spinal cord potentials.

One hundred twenty-three cases of cervical spondylotic and ossification of the posterior longitudinal ligament (OPLL) myelopathy cases with long tract signs subjected to surgical treatment were studied to identify the most important factors having an influence on postoperative outcome using evoked spinal cord potentials (ESCP). Disappearance and positive wave changes of these potentials at the level of responsible lesions and slow conduction velocity under 40 m indicated an unsatisfactory outcome. Localized-lesion cases diagnosed by ESCPs had excellent results, significantly more so than extensive-lesion cases, regardless of operative methods. In 123 cases, 76% were found to have localized lesions, while the other 24% showed extensive lesions. Concerning the difference between CSM and OPLL, 54% of OPLL and only 14% of CSM demonstrated extensive lesions.

Aged↗
PubMed1990

Antiscorbutic activity of L-ascorbic acid 2-glucoside and its availability as a vitamin C supplement in normal rats and guinea pigs.

Bioavailability of a newly-synthesized and chemically-stable 2-O-alpha-D-glucopyranosyl-L-ascorbic acid (AA-2G) as a vitamin C supplement was investigated in rats and guinea pigs. Oral administration of AA-2G to the animals resulted in an increase of serum ascorbic acid (AA) levels. However, in these sera the intact form was not detectable by the high performance liquid chromatography (HPLC) method, indicating its hydrolysis through the process of absorption. After an intravenous injection of AA-2G, the intact form diminished rapidly from the serum, followed by prolonged and marked elevation of serum AA levels. Various tissue homogenates from rats and guinea pigs were examined for their releasing activity of AA from AA-2G. High activity was observed in kidney, small intestine and serum of rats and in small intestine and kidney of guinea pigs. These hydrolytic activities were completely inhibited by castanospermine, a specific alpha-glucosidase inhibitor, suggesting the participation of alpha-glucosidase in the in vivo hydrolysis of AA-2G. AA-2G was found to exhibit obvious therapeutic effect in scorbutic guinea pigs by its repeated oral administrations. These results indicate that AA-2G is a readily available source of vitamin C activity in vivo.

Administration, Oral↗
PubMed1990

L-ascorbic acid alpha-glucoside formed by regioselective transglucosylation with rat intestinal and rice seed alpha-glucosidases: its improved stability and structure determination.

The definite structure and chemical stability of a new glucoside of L-ascorbic acid (AA) which was enzymatically glucosylated with rat intestinal and rice seed alpha-glucosidases were reported. The stability of this AA derivative in water under aerobic conditions was proved by its remarkable resistance against enhanced oxidative degradation by heat, Cu2+ ion or ascorbate oxidase, and it was found to have no reducing activity toward radicals. These properties were obviously distinguishable from those of AA. This glucoside was effectively hydrolyzed by alpha-glucosidases which possessed the ability to synthesize itself, resulting in the liberation of AA activity. The conjugate was composed of equimoles of AA and glucose. Nuclear magnetic resonance spectra, mass spectra, pH profiles of ultraviolet spectra and pK(a) value of 3.10 supported the coupling of alpha-glucose to the 2-position of AA. From these results, its structure was assigned 2-O-alpha-D-glucopyranosyl-L-ascorbic acid, being distinct from 6-O-alpha-D-glucopyranosyl-L-ascorbic acid formed with Aspergillus niger alpha-glucosidase. These findings indicate that the 2-O-glucoside formed by regioselective transglucosylation withstands oxidative degradation even in aqueous solutions and it can be used as an available active AA source for multicomponent liquid products.

Animals↗
PubMed1989

Interaction of rat brain cytidylate cyclase with phospholipids.

The interaction of rat brain cytidylate cyclase with some phospholipids such as L-alpha-phosphatidylcholine (PC), L-alpha-phosphatidylserine (PS), L-alpha-phosphatidylethanolamine (PE) and L-alpha-phosphatidic acid (PA) was studied. Cytidylate cyclase activity of Triton X-100 - solubilized fraction was inhibited by PS, PE and PA, but not with PC. The addition of PC to the incubation mixture containing PS, PE or PA dose - dependently reversed the inhibition of enzyme activity by these phospholipids. Phospholipids showed similar effect on the intact membrane - bound enzyme. PC could reactivate the enzyme which was inactivated by deoxycholate treatment, suggesting that PC may be an important factor to reconstitute an active conformation of the enzyme. These findings indicate that cytidylate cyclase could be regulated by phospholipids constituting its microenvironment of the membrane.

Animals↗
PubMed1989

An enzyme immunoassay for K-76 monocarboxylic acid, a novel anticomplementary compound.

A competitive enzyme immunoassay for K-76 monocarboxylic acid (K-76COOH), a novel anticomplementary compound, was developed. K-76COOH was directly coupled with bovine serum albumin through a formation of Schiff base and successive reduction. The spectral data of the conjugate showed no evidence of a Schiff base form. Using the specific antiserum, the proposed homologous assay made it possible to detect K-76COOH at the lowest value of 1 ng/ml of plasma. The immunoassay was validated by the correlation with HPLC analyses. The time courses of plasma levels of K-76COOH after a single oral administration to beagle dogs were precisely determined with a very low absorption efficiency. From these results, it is suggested that the plasma values obtained are insufficient for K-76COOH to exert its anticomplementary action in vivo; thus K-76COOH may have another immunopharmacological function.

Animals↗
PubMed1989

Acid secretory effect of extracted guinea-pig gastrin on isolated guinea-pig parietal cells.

It was confirmed that histamine, cholinergic drugs and gastrin separately produce acid secretory effects of isolated parietal cells. However, the acid secretory response of isolated parietal cells to gastrin stimulation is known to be weaker than that to histamine or cholinergic drugs, which is different from the acid secretory response found in vivo studies. Tetragastrin and pentagastrin were less potent than histamine and carbachol in stimulating acid secretion in our previous study performed using parietal cells isolated from guinea-pigs. In this study, the effect of gastrin extracted from the pyloric antrum of the guinea-pig on parietal cells isolated from the guinea-pig was investigated based on the assumption that the acid secretory response of isolated parietal cells to gastrin stimulation will be especially good if gastrin isolated from the same species is used.

Animals↗
PubMed1989

[Changes in cytosolic free Ca++ by histamine stimulation in isolated guinea-pig parietal cells (the second report)].

It was demonstrated in our previous report that cytosolic free Ca++ concentration showed increase following histamine stimulation in the dose dependent manner, by double wave length excitation method using fura-2 as an intracellular Ca++ probe. In this paper, the investigation about the source of cytosolic free Ca++ coming out following histamine stimulation was performed. Both Ca++ channel blocker (Verapamil and Diltiazem) and an intracellular Ca++ antagonist (TMB-8) suppressed the increase of cytosolic Ca++ concentration following histamine stimulation. 10(-4)M verapamil, 10(-4)M diltiazem and 10(-4)M TMB-8 suppressed cytosolic Ca++ concentration from 355.9 +/- 5.7 nM to 213.8 +/- 4.9 nM, from 330.1 +/- 7.3 nM to 149.8 +/- 1.7 nM and from 324.9 +/- 6.1 nM to 151.3 +/- 3.8 nM respectively in the values at the point of 20 min after 10(-4)M histamine stimulation. The time course study revealed that the course of increase consisted of two components: the "initial rise" which was observed in the first 1 min. followed by the "continuous rise". The initial rise may reflect the Ca++ coming out from the intracellular Ca++ pool and the continuous rise may reflect the Ca++ flowed into cell from outside. Verapamil and diltiazem did not suppress the initial rise, while TMB-8 suppressed it. It is concluded that increase of cytosolic Ca++ concentration following histamine stimulation may be concerned with both extra- and intracellular Ca++ and that at first, intracellular Ca++ may be mobilized.

Animals↗
PubMed1988

Further studies on rat cathepsin E: subcellular localization and existence of the active subunit form.

The subcellular localization of rat neutrophil cathepsin E was examined by a modification of the method of N. Borregaard et al. [(1983) J. Cell Biol. 97, 52-61]. When the postnuclear cavitate of rat neutrophils was subjected to density centrifugation on discontinuous Percoll gradients, three particulate bands, P1 (lowest; azurophil granule rich), P2 (middle; specific granule rich), and P3 (highest; plasma membrane rich), were segregated. A combined application of immunochemical and electrophoretic methods revealed a striking difference in subcellular localization between cathepsin E and cathepsin D: Cathepsin E was associated with P3 and soluble fractions, and cathepsin D was chiefly associated with P1 and P2 fractions. The results thus indicate that cathepsin E is a nonlysosomal acid proteinase in rat neutrophils. It was found that cathepsin E existed in two enzymatically active molecular forms, referred to as CE-I and CE-II, in rat neutrophil extracts. To examine the relationships between the two forms, cathepsin E was purified to homogeneity from rat gastric mucosae. The purified enzyme exhibited a single protein band of 43 kDa on sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis, but electrophoresis without SDS, followed by visualization of activity in the gel, revealed two activity bands corresponding to CE-II and CE-I in neutrophil extracts. Pretreatment of the enzyme with beta-mercaptoethanol or dithiothreitol resulted in an increase in CE-I activity with a concomitant decrease in CE-II activity on gels. Upon gel filtration, the molecular weights of CE-II and CE-I were estimated to be 98,000 and 51,000, respectively, strongly suggesting that they are the dimeric and monomeric forms of the cathepsin E subunit.

Animals↗
PubMed1988

Characteristic distribution of cathepsin E which immunologically cross-reacts with the 86-kDa acid proteinase from rat gastric mucosa.

The antiserum raised against the high-molecular-weight acid proteinase from rat gastric mucosa, termed 86-kDa acid proteinase, has been shown to recognize rat cathepsin E, but not cathepsin D (Muto, N. et al. (1987) J. Biochem. 101, 1069-1075). Using this specific antiserum, characteristic distribution of cathepsin E in rats was demonstrated. The enzyme was detected in a limited number of tissues, such as stomach, thymus, spleen, bladder, and erythrocyte membranes. Among them, the highest activity was observed in the stomach. In contrast, cathepsin D immunoreactive with the antiserum specific to rat gastric cathepsin D was demonstrated in all the tissues examined. Cathepsin E-type enzymes partially purified from these five tissues were precipitated in the same manner by the specific antiserum, and they had the same molecular weight, electrophoretic mobility, and resistance against denaturation by 4 M urea. These results indicate that they could be exactly classified as cathepsin E. This type of enzyme was also detectable in mice and guinea pigs, but they showed relatively weak immunoreactivities with the antiserum. Thus, it is concluded that the distribution of cathepsin E is intrinsically different from ordinary cathepsin D, suggesting that it has a different physiological role from cathepsin D.

Animals↗
PubMed1987

Inhibition of cytochrome P-450-linked monooxygenase systems by naphthoquinones.

Several naphthoquinones, except 2-hydroxy-1,4-naphthoquinone, were found to inhibit microsomal cytochrome P-450-linked monooxygenase activities in rabbit liver and human placenta. In particular, 5-hydroxy-1,4-naphthoquinone inhibited placental estrogen biosynthesis more effectively than it did hepatic drug oxidation reactions. There was little contribution by superoxide radicals to these enzyme inhibitions by naphthoquinones. Spectrophotometric studies revealed that naphthoquinones bind to the cytochrome P-450 component of the monooxygenase complex in both microsomal systems, suggesting that the inhibition is caused by direct interaction of these compounds with the heme.

Androstenedione↗
PubMed1987

Immunochemical similarity between a gastric mucosa non-pepsin acid proteinase and neutrophil cathepsin E of the rat.

Antiserum against a rat gastric mucosa non-pepsin acid proteinase precipitates rat neutrophil cathepsin E, with a precipitation curve essentially similar to that of the gastric enzyme. Taken together that the antiserum precipitates a cathepsin E-like acid proteinase from rat spleen (Muto, N., Yamamoto, M. and Tani, S. (1987) J. Biochem. (Tokyo) in press), the data indicate that the non-cathepsin D acid proteinases in rat neutrophils, gastric mucosa and spleen are immunochemically closely related. In contrast with the earlier data, cathepsin E from rabbit neutrophils exhibited a maximal activity at around pH 3.0-3.2 and preferred hemoglobin to albumin as substrate, which supports that the non-cathepsin D acid proteinases in the rat tissues are relevantly classified as cathepsin E.

Animals↗
PubMed1987

Comparative studies of two types of acid proteases from rat gastric mucosa and spleen.

Two types of acid proteases, cathepsin D and cathepsin E-like enzyme, from rat gastric mucosa and spleen were compared in their biochemical and immunochemical properties. The enzymes were partially purified by employing the same chromatographic procedures and they showed a single proteolytically active band in polyacrylamide gel electrophoresis. Two low molecular weight enzymes, cathepsins D, from both tissues showed the same molecular weight and the same sensitivities to various inhibitors, but slightly different electrophoretic mobilities. The rabbit antiserum raised against gastric mucosa cathepsin D precipitated both enzymes. On the other hand, high molecular weight enzymes, gastric mucosa cathepsin D-like acid proteinase and spleen cathepsin E-like acid proteinase, were similar to each other as judged by their chromatographic profiles, electrophoretic mobilities, and high stabilities in urea solution. Furthermore, the antiserum specific to gastric mucosa cathepsin D-like acid proteinase inhibited both enzyme activities in a similar manner. However, the antiserum specific to one type of enzyme did not react with the other type. These results indicate that: gastric mucosa cathepsin D is immunologically identical with spleen cathepsin D; gastric mucosa cathepsin D-like acid proteinase has biochemical and immunological properties quite similar to spleen cathepsin E-like enzyme; these two types of acid proteases are quite different proteins existing in the individual tissues.

Animals↗
PubMed1987

Alteration in gastric mucosal acid protease activity induced by necrotizing agents and prevention by prostaglandin E2.

Tissue levels of two gastric mucosal acid proteases, pepsinogen and cathepsin D-like acid proteinase, were determined in rat gastric mucosa damaged by various necrotizing agents and the protective effects of prostaglandins against these biological alterations were investigated. Gastric mucosal damage by each necrotizing agent used was associated with a marked decrease in tissue level of cathepsin D-like acid proteinase. Particularly, ethanol ingestion caused its significant reduction parallel to the production of gastric lesions in a time-dependent manner. On the other hand, mucosal pepsinogen level increased markedly only in ethanol-damaged gastric mucosa, indicating that this change was mediated by a different mechanism from that for cathepsin D-like enzyme. In rats pretreated with prostaglandin E2 and prostaglandin inducers before ethanol administration, these biological alterations of two enzymes were effectively prevented as were gastric lesions. However, ethanol ingestion caused these changes to occur to the same degree in both the necrotic and non-necrotic areas of glandular mucosa. It was considered that cathepsin D-like acid proteinase was released from damaged gastric mucosa through a direct action on cellular membrane different from vasoconstrictor and platelet aggregating actions mediated by arachidonic acid metabolites.

Animals↗