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Biomedical subjects

K Miyazaki

Publications and source records attributed to K Miyazaki.

At least 541 records · Page 30Linked to original sources
PubMed1990

Purification and characterization of extracellular matrix-degrading metalloproteinase, matrin (pump-1), secreted from human rectal carcinoma cell line.

A metalloproteinase with Mr 29,000 was purified to homogeneity as a latent proenzyme from the conditioned medium of a human rectal carcinoma cell line CaR-1. This enzyme hydrolyzed casein more potently than gelatin embedded in polyacrylamide gels in zymography assay. Calcium ion was essential for the activity. It exerted the maximum activity at pH 7-9. Its activity was stimulated by organomercurials, such as p-amino-phenyl mercuric acetate and p-chloromercuric benzoic acid, and was inhibited by 1,10-phenanthroline but was hardly affected by diisopropyl fluorophosphate and pepstatin. When the purified proenzyme was activated by the organomercurials, it effectively hydrolyzed fibronectin, laminin, type IV basement membrane collagen, and several types of gelatins but not interstitial type I and III collagens. The treatment of the purified proenzyme with p-aminophenyl mercuric acetate or trypsin formed an active peptide with Mr 20,000. The structural analysis indicated that it was most likely identical to putative metalloproteinase-1, the complementary DNA of which had been cloned from human tumor mRNAs capable of hybridizing to a rat transin complementary DNA. Based on the fact that this enzyme is secreted extracellularly and degrades the matrix proteins, we propose the name "matrin" for this newly identified enzyme.

Amino Acid Sequence↗
PubMed1990

Potent growth inhibition of human tumor cells in culture by arginine deiminase purified from a culture medium of a Mycoplasma-infected cell line.

Two kinds of growth-inhibitory substances were found in culture of a Rous sarcoma virus-transformed rat liver cell line, RSV-BRL. The two substances were purified from the serum-free culture medium and identified as transforming growth factor beta 1 and Mycoplasma-derived arginine deiminase (EC 3.5.3.6), respectively. The arginine deiminase was an acid-labile but dithiothreitol-resistant protein with a molecular weight of 45,000 and pI 4.7. Its Km value for L-arginine was 0.3 mM, which is about 30 times lower than that of bovine liver arginase. It was stable and active under culture conditions. When added into cultures, the arginine deiminase inhibited the growth of various human cancer cell lines at a dose of 5 ng/ml or higher by depleting L-arginine in the culture media. This effective dose was about 1000 times lower than that of bovine liver arginase. These results suggested the possibility of chemotherapeutic use of arginine deiminase for human cancers.

Amino Acid Sequence↗
PubMed1990

Calcified ligamenta flava in a patient with Coffin-Lowry syndrome: biochemical analysis of glycosaminoglycans.

Glycosaminoglycans in calcified cervical ligamenta flava from a male patient with Coffin-Lowry syndrome were analyzed biochemically. Hexuronate content based on the tissue dry weight was 8-fold and 5-fold concentrated in the calcified center as compared with the peripheral area and with age-matched, normal ligamenta flava, respectively. Two-dimensional electrophoresis of glycosaminoglycan preparations on cellulose acetate film revealed that, although dermatan sulfate was the major glycosaminoglycan in the normal ligamenta flava, chondroitin sulfate was the predominant glycosaminoglycan in the pathological tissue. Relative amounts of chondroitin sulfate disaccharide units were determined after digestion of glycosaminoglycans with chondroitinase, and the ratio of chondroitin 6-sulfate to chondroitin 4-sulfate tended to decrease with the distance from the calcified center to the peripheral area. These observations suggest that an alteration in glycosaminoglycan metabolism may be a contributing factor in calcification of ligamenta flava in Coffin-Lowry syndrome.

Abnormalities, Multiple↗
PubMed1990

Serial measurement of CA 125 in patients with primary carcinoma of the fallopian tube.

Serial assay of serum CA 125 was carried out in five patients with advanced primary carcinoma of the fallopian tube. Pretreatmental levels of these patients were elevated, ranging from 145 to 535 U/ml (305 +/- 140.1, mean +/- SD). All patients showed rapid decreases in response to treatment; however, reelevations were noted in two patients concomitant with recurrence. In the remaining three patients, who are alive with no evidence of recurrence at 52, 29, and 21 months, serum CA 125 levels remained below 5 U/ml after remission. These results suggest that CA 125 is useful in monitoring patients with primary carcinoma of the fallopian tube.

Antigens, Tumor-Associated, Carbohydrate↗
PubMed1990

Unscheduled DNA synthesis after treatment with 20-methylcholanthrene and N-nitrosodimethylamine in primary culture of human gallbladder epithelial cells.

Unscheduled DNA synthesis (UDS) induced by treatment with 20-methylcholanthrene (MCA) and N-nitrosodimethylamine (DMN) was measured in cultured human gallbladder epithelial cells. MCA induced UDS very efficiently, while DMN was far less effective than MCA. Addition of rat S9 mixture did not affect the amount of UDS by the chemicals. Differences between the present results in human cells and our previous findings in bovine cells could be due to species and tissue specificity.

Cells, Cultured↗
PubMed1990

The effect of sodium hyaluronate on the migration of rabbit corneal epithelium. I. An in vitro study.

The effect of sodium hyaluronate (Na-HA) on the migration of corneal epithelium was investigated in vitro using corneo-scleral sections of rabbits. The effect on migration was evaluated by the length of migrating epithelium on the cut surface of corneal stroma after incubation in media containing Na-HA of various molecular weights or concentrations. The medium concentration of Na-HA was 400 micrograms/ml, and the molecular weights were 0.4 x 10(4), 2 x 10(4), 10 x 10(4), 68 x 10(4), 102 x 10(4) and 216 x 10(4). Regarding Na-HA with a molecular weight of 102 x 10(4); media containing Na-HA at concentrations of 50, 100, 200, 400, 800, 2000 and 4000 micrograms/ml were also investigated. When the concentration of Na-HA was 400 micrograms/ml, Na-HA with molecular weights of over 68 x 10(4) accelerated significantly the epithelial migration. Regarding Na-HA with a molecular weight of 102 x 10(4), acceleration was observed at concentrations of 200, 400 and 800 micrograms/ml. Furthermore, the concentration of fibronectin (FN) in medium containing the effective Na-HA was significantly lower than that in the control medium after the incubation. These results suggest a possibility that the accelerative effect of Na-HA depends on the reduction of FN release from the cornea or the facilitation of FN deposition on the cornea.

Animals↗
PubMed1990

Purification, catalytic properties, and thermal stability of threo-Ds-3-isopropylmalate dehydrogenase coded by leuB gene from an extreme thermophile, Thermus thermophilus strain HB8.

Threo-Ds-3-isopropylmalate dehydrogenase coded by the leuB gene from an extreme thermophile, Thermus thermophilus strain HB8, was expressed in Escherichia coli carrying a recombinant plasmid. The thermostable enzyme thus produced was extracted from the E. coli cells, purified, and crystallized. The enzyme was shown to be a dimer of identical subunits of molecular weight (4.0 +/- 0.5) x 10(4). The Km for threo-Ds-3-isopropylmalate was estimated to be 8.0 x 10(-5) M and that for NAD 6.3 x 10(-4) M. The optimum pH at 75 degrees C in the presence of 1.2 M KCl was around 7.2. The presence of Mg2+ or Mn2+ was essential for the enzyme action. The enzyme was activated about 30-fold by the addition of 1 M KCl or RbCl. The high salt concentration decelerated the thermal unfolding of the enzyme, and accelerated the aggregation of the unfolded protein. Based on these effects, the molecular mechanism of the unusual stability of the enzyme is discussed.

3-Isopropylmalate Dehydrogenase↗
PubMed1990

Purification and properties of extracellular matrix-degrading metallo-proteinase overproduced by Rous sarcoma virus-transformed rat liver cell line, and its identification as transin.

Rous sarcoma virus-transformed rat liver cell line RSV-BRL secreted a neutral proteinase in a latent precursor form with a molecular weight (Mr) of 57,000 (57k) as a major secreted protein. This enzyme was a calcium-dependent metallo-proteinase. The proenzyme was purified from the serum-free conditioned medium of the transformed cells by affinity chromatographies on a zinc chelate Sepharose column and a reactive red agarose column. When activated by treatment with trypsin or p-aminophenylmercuric acetate (APMA) in the presence of Ca2+, the purified enzyme effectively hydrolyzed casein, fibronectin, and laminin. Type IV collagen was hydrolyzed at 37 degrees C but not at 30 degrees C by the enzyme, whereas type I and type III collagens were hardly hydrolyzed even at 37 degrees C. The treatment with trypsin or AMPA in the presence of Ca2+ converted this 57k proenzyme to an active and stable enzyme with Mr 42k. In the absence of Ca2+, however, APMA converted the proenzyme to an intermediate form with Mr 45k, while trypsin digested it to an inactive peptide with Mr 30k. These results demonstrate that calcium ion is essential for the activation, activity expression, and stabilization of this metallo-proteinase. Analysis of its partial amino acid sequence and amino acid composition showed that the 57k proenzyme was identical or closely related to the putative protein transin, a rat homologue of stromelysin.

Amino Acid Sequence↗
PubMed1990

Tiopronin-induced lichenoid eruption in a patient with liver disease and positive patch test reaction to drugs with sulfhydryl group.

A 62-year-old woman suffering from liver cirrhosis developed lichenoid eruptions after 2 years of treatment with tiopronin. The lesions healed spontaneously within a month after withdrawal of the drug. In patch testing, the patient reacted positively, not only to tiopronin, but also to captopril and D-penicillamine, neither of which she had ever taken before. The provocation test was positive only to tiopronin, and its histological findings revealed lichenoid reaction. It is suggested that the sulfhydryl group may play a role in the etiology of tiopronin-induced lichenoid drug eruption.

Drug Eruptions↗
PubMed1990

Correlation between structures of organic cations and their binding behaviours to brush border membrane isolated from rat small intestine.

We have investigated the correlation between the molecular structures of various organic cations and their binding to rat small intestinal brush border membrane. The binding of small quaternary ammonium compounds such as tetramethylammonium and choline to brush border membrane was not sufficient to inhibit methylchlorpromazine binding. However, lauryltrimethylammonium and cetyltrimethylammonium, both quaternary amines with a long carbon chain, inhibited binding significantly. The inhibition was competitive. When the unbranched hydrocarbon chain of the quaternary amines was extended in steps from C1 (methyl) to C16 (cetyl), the inhibitory effect increased sharply with length from C7 (heptyl) to C16. These results suggest that the size of the hydrophobic part of the molecule is an important factor in binding of quaternary ammonium compounds to the brush border membrane. The structure of the hydrophilic part was another factor. In imipramine-related compounds, the order of binding was N-didesmethylimipramine (primary amine) greater than desipramine (secondary) greater than imipramine (tertiary) greater than methylimipramine (quaternary). However, with the small molecular ethylamine-related compounds, binding properties did not reflect differences in the hydrophilic component. Therefore, the effect of the hydrophilic part may be secondary and may depend on the size of the hydrophobic part. We suggest that organic cations which are amphiphilic can bind to a common binding site on brush border membrane through hydrophobic and/or hydrophilic interactions.

Animals↗
PubMed1990

Contribution of passive transport mechanisms to the intestinal absorption of beta-lactam antibiotics.

The transport characteristics of aminopenicillins (ampicillin and amoxicillin), aminocephalosporins (cephalexin, cephradine and cefadroxil) and cefazolin have been compared with those of an actively transported substance (D-glucose) and a passively transported substance (L-glucose). Although the initial uptake of the aminocephalosporins was stimulated in the presence of an inward H+ gradient, there was no overshoot in the uptake of any of the drugs tested, even in the presence of an H+ gradient. Also, the time course and the degree of uptake of these drugs were similar to those of L-glucose, especially in the absence of an H+ gradient. These results suggest that the beta-lactam antibiotics tested, like L-glucose, pass through the rat intestinal brush border membrane mainly by passive diffusion. However, the differences in absorption between these drugs, like the differences in their disappearance from a proximal loop of rat intestine, cannot be explained by a simple permeation process alone.

Animals↗
PubMed1990

Determination of triethylenetetramine in plasma of patients by high-performance liquid chromatography.

A sensitive and simple fluorometric method for the determination of N,N'-bis(2-aminoethyl)-1,2-ethanediamine dihydrochloride (triethylenetetramine) in human plasma by high-performance liquid chromatography is described. Free triethylenetetramine (TETA) obtained by passing the TETA-copper chelate compound through a solid-phase cation exchange resin was converted to its fluorescamine derivative in the presence of ethylenediaminetetraacetic acid to mask the interfering metal ions in the reaction solution, and the derivatives were separated on a nitrile high-performance liquid chromatograph column (Nucleosil 5-CN) using isocratic elution. The plasma levels of TETA were measured in eight patients receiving treatment for excess copper. Absorption rates of TETA were relatively slow and the peak levels were significantly different among patients. The bioavailability of TETA in the rat was also examined and the ratio of intestinal absorption was extremely low.

Animals↗
PubMed1990

[Intestinal absorption and urinary excretion of triethylenetetramine for Wilson's disease in rat].

Triethylenetetramine.2.HCl (trientine, TE) is an orphan drug for the treatment of Wilson's disease. There has been no reports regarding the absorption, distribution, metabolism, and excretion in the body. In the current study, the absorption and excretion of TE in rats were examined. The observed mean percentage amount of TE absorbed at the jejunum and ileum with the loop method for 1 h was 42.0% and 22.5%, respectively. Tight junction blocking agent inhibited the absorption of TE from the jejunum loop with 27%, but the absorption of TE from the ileum loop was not affected by this blocking agent. Therefore, the main absorption route for TE might be permeation across the plasma membrane of intestinal epithelial cells. TE and amikacin, a polycationic compound like TE, bound to the brush border membrane (BBM) of rat small intestine in the absence of inorganic ions such as Na+, K+, Ca2+, Mg2+ and Cu2+. But the binding of TE to BBM was inhibited markedly under the physiological concentration of these ions. The bioavailability of TE was below 10% and the plasma levels of TE in non-fasted rats were significantly lower than that observed in fasted rats. The urinary excretion of unchanged TE during 24 h was only 3.5% of the orally administered dose. However, the urinary excretion of total TE including metabolites, though they have not been identified, was 35.7%. These results suggest that low bioavailability of TE might be due to the rapid metabolism in the body after absorption from the gastrointestinal tract.

Administration, Oral↗
PubMed1990

[The male reproductive function in patients with spinal cord injury].

Seminal findings and blood hormone levels were studied for evaluating the male reproductive function in patients with spinal cord injury. The patients were divided into 3 groups, namely, 18 patients with complete injury, 5 patients with incomplete injury and 3 patients with dyspermatism. The number of sperms, the rate of movement and rate of deformation were measured for semen obtained by forced ejaculation. The number of sperms was kept at a relatively high level in the three groups, while the rate of movement fell off in all of the three groups. The rate of deformation was highest in the patients with complete injury and lowest in the patients with dyspermatism. As for blood hormone levels, LH, FSH and Testosterone (hereinafter referred to as TES) were determined by the RIA. The cases were classified into those in the acute stage and those in the chronic stage 3 months after sustaining injury for a comparative study. The subjects consisted of 27 cases in the acute stage and 47 cases in the chronic stage. For 8 patients in the acute stage, the blood hormone levels were determined even in the chronic stage and follow-up observations were made on the changes in the levels. The FSH level was low in both stages, while LH and TES tended to increase in the chronic stage. Particularly, the TES level was elevated in all the cases in the follow-up observations made in 8 patients. From the results mentioned above, transient disturbance of the interstitial function is suggested as the mechanism of male gonadal disturbance due to spinal cord injury.

Adolescent↗
PubMed1990

[Clinico-pathological investigation on frequency and prognosis in patients with incidental prostatic carcinoma: preliminary report].

To investigate the relationship between some clinico-pathological features of incidental prostatic carcinoma and its prognosis, totally 96 consecutive cases with simple prostatectomy from 1968 to 1973 under the diagnosis of prostatic hypertrophy were examined by the 3 mm step-section technique. Twenty-one cases (21.9%) of incidental carcinoma were detected histologically. The age of the patients with or without prostatic carcinoma ranged from 59 to 81 years old, 72.0 years old on the average, and from 54 to 87 years old, 69.9 years old on the average, respectively. The weight of the specimen varied from 7 to 84 grams, 32.4 g on the average, for the patients with carcinoma, and 10 to 64 grams, 29.7 g on the average, for the patients without carcinoma. Fifteen of 21 cases showed well differentiated carcinoma. None of the Japanese histological differentiation, Gleason's classification and volume of carcinoma presumably affected the survival of the patients.

Aged↗
PubMed1990

[Characterization of tumor cell growth inhibitor present in rabbit serum].

Growth inhibitors present in various kinds of sera were surveyed using the rat liver epithelial cell line BRL and its tumorigenic transformant RSV-BRL as indicator cells. This survey revealed that normal rabbit serum contained two types of growth inhibitors: one (GI-A) was more growth-inhibitory on RSV-BRL than BRL, whereas the other (GI-B) vice versa. GI-A was purified 3,000-fold to show a major protein band with Mr 70k on SDS-PAGE. It was an acid-and heat-labile protein and potently inhibited the growth of three kinds of transformed cell lines and two human carcinoma cell lines, but hardly that of non-transformed cell lines, at a dose of 0.5-1.0 micrograms/ml. On the other hand, GI-B was an acid- and heat-stable protein with Mr 25k and was considered to belong to the TGF-beta family.

Animals↗