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T Kusaka

Publications and source records attributed to T Kusaka.

At least 55 records · Page 3Linked to original sources
PubMed1992

[A case of intrathoracic chest wall type lipoma].

Intrathoracic lipoma is a comparatively rare disease. We have recently experienced a case of intrathoracic chest wall type lipoma associated with Charcot-Marie-Tooth disease. The case was a 16-year-old man, in whom preoperative examination of his foot-drop due to Charcot-Marie-Tooth disease pointed out an abnormal chest shadow. Image diagnosis led to intrathoracic chest wall type lipoma. The tumor was easily resectable and was lipoma pathohistologically as well. The literature presented us with 22 cases of such lipoma, which appeared to be predominant for age not more than 10 and age 41-60 and to occur favorably in postero-superior chest wall. Preoperative diagnosis of the present disease became possible to a certain extent by means of CT, MRI. However, in view of imperfect differentiation between lipoma and liposarcoma, surgical resection should be considered, if possibly. No case of intrathoracic lipoma associated with Charcot-Marie-Tooth disease had been reported in the literature, and the causal relationship there between also remains unknown, but to be further examined.

Adolescent↗
PubMed1991

[Analysis of dysanapsis in healthy twins and sons of patients with chronic obstructive lung disease].

Pulmonary function tests and tracheal cross-sectional area (X-SA) measured on chest roentgenograms were analyzed in 72 healthy pairs of twins and in 34 healthy sons of patients with chronic obstructive lung disease. In the twin study, genetic influence was positive on most pulmonary function tests, including VC, FEV1/FVC, V25, but not so on X-SA. The ratio of X-SA to VC, which represents dysanapsis, had no relation to sex nor height. However, the ratio of X-SA/VC showed marked sex-associated differences, when expressed as a function of VC, and had a negative correlation with VC both in men and women. The ratio of X-SA/VC was significantly lower in sons of patients with COPD than in normal controls, which was caused by significantly larger VC in the former group. The relatively low ratio of tracheal size to lung size may partly explain the susceptibility to the possible future development of COPD in those subjects.

Adolescent↗
PubMed1989

Covalent immobilization of the estrogen receptor to an electrostatically neutral N-hydroxysuccinimide ester derivative of agarose.

Immobilization of the estrogen receptor to the N-hydroxysuccinimide ester of succinylethylenediaminocarboxymethyl agarose (Reagent B) is described and compared with that to the charged N-hydroxysuccinimide ester derivative (Reagent A), previously described. The time course for immobilization was examined. Thirty-six percent of the input receptor was immobilized within 1 h. The optimum pH in immobilization is 7.0-7.4. The dissociation rate of [3H]estradiol(3,17 beta-1,3,5(10)-estratriene) from the [3H]estradiol-receptor complex immobilized to Reagent B was similar to that in Reagent A. The receptor immobilized to Reagent B was saturated with estradiol at 5 h. The [3H]estradiol concentration necessary for saturation was 10 nM. The dissociation constant (KD) for the receptor immobilized to Reagent B was 0.95 X 10(-9) M.

Binding, Competitive↗
PubMed1988

Covalent immobilization of the estrogen receptor to a cationic N-hydroxysuccinimide ester derivative of agarose.

Covalent immobilization of the soluble estrogen receptor from a rabbit uterus to N-hydroxysuccinimide ester derivative of agarose is shown. At first, the condition for the immobilization reaction was examined. The non-immobilized receptor was extracted with 0.4 M NaCl-containing medium. Sixty seven to 80% of the input receptor were immobilized within 30 min at 0 degrees C in 0.1 M HEPES (N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid, pH 7.4). The immobilized [3H]estradiol(3,17 beta-dihydroxy-1,3,5(10)-estratriene)-receptor complex was stable for at least 24 h. The optimum pH for immobilization was 7.4. Ca2+ or Na+ ions in the reaction media decreased the yields in immobilization of the receptor to the reagent with an electrostatically positive spacer arm. Next, influences of immobilization on the receptor were examined. The dissociation rate of [3H]estradiol from the immobilized receptor was a little slower than that from the native receptor. The estrogen-free immobilized receptor was saturated by incubating with 10 nM [3H]estradiol for 10 h at 0 degrees C in 0.1 M HEPES (pH 7.4). From Scatchard plot analysis, it was found that the hormone binding affinity in the immobilized receptor decreased to approximately one-fourth of that in the native receptor.

Animals↗
PubMed1988

Liquid chromatography/mass spectrometry of fatty acids as their anilides.

The mass spectra of a series of saturated (C16:0-C30:0) and unsaturated (C16:1, C18:1, C18:2, C18:3) fatty acids have been recorded as their anilides using liquid chromatography/mass spectrometry with the atmospheric-pressure-ionization interface system. The spectra show an intense peak for (molecule + H)+ ion in each case. The liquid chromatographic/mass spectrometric separation was performed on a reverse phase column using a solvent system of methanol alone or methanol + 2-propanol. This method seemed promising for application to both qualitative and quantitative micro-analysis of fatty acids including very long chain fatty acids.

Anilides↗
PubMed1988

Hexacosanoate contents in Japanese common foods.

Recently, a diet enriched in oleate and moderately restricted in hexacosanoate (C26:0) was found effective to reduce the plasma very long chain fatty acid (VLCFA) levels in patients with adrenoleukodystrophy (ALD), an X-linked disorder characterized by demyelination of the adrenal cortex and cerebral white matter, and accumulation of saturated VLCFA, particularly C26:0, in tissues of the demyelination. The information about the C26:0 content in Japanese food was, however, almost nil except for one report about foods in the USA, but this did not include some Japanese common foods. With the hope of treating an ALD patient in our hospital, C26:0 contents in Japanese common foods (42 items) were measured. In our case, a one-hour direct transesterification method was used to obtain methylesters of total fatty acids in foods and they were applied directly to a selected ion monitoring gas chromatography-mass spectrometry for the quantitative C26:0 analysis. The C26:0 content in nuts and seeds as well as in fats and oils was found to be significantly higher than in other foods; the content was highest in peanuts. The content in almost all kinds of examined fishes, the common protein foods in Japan, was relatively low. From these data and that in the national nutrition survey in 1986, the daily intake of C26:0 from the average Japanese diet could be estimated to be 12-36 mg. It can be recommended, therefore, that nuts and seeds as well as fats and oils should be restricted as severely as possible from the diet of ALD patients in Japan in order to keep daily C26:0 intake below 10 mg as recommended in the USA.

Adrenoleukodystrophy↗
PubMed1986

Requirement of glucose for mycolic acid biosynthetic activity localized in the cell wall of Bacterionema matruchotii.

When the localization of mycolic acid biosynthetic activity was examined with Bacterionema matruchotii cells disrupted by the ultrasonic vibration method, activity was detected only in the cell wall fraction, not in the inner membrane nor in the 78,000g supernatant. Either the supernatant or sugar was absolutely required for the incorporation of [14C]palmitate into mycolic acids. Among sugars examined, glucose was most effective, with maltose being second. Unexpectedly, trehalose was inert. As to substrate, the present system utilized free palmitic acid rather than palmitoyl-CoA. The reaction products from palmitate and glucose were glucose mycolate and trehalose monomycolate, in which the label from [14C]palmitate or [14C]glucose was incorporated. Glucose palmitate was also formed. Addition of trehalose resulted in a shift from glucose mycolate to trehalose monomycolate. These data clearly indicate that sugars play an important role in the synthesis of mycolic acids from free fatty acids.

Actinomycetaceae↗
PubMed1986

Isolation and partial characterization of a very long-chain fatty acid desaturation system from the cytosol of Mycobacterium smegmatis.

The supernatant fraction after 105,000 X g centrifugation of an extract of sonically disrupted cells of Mycobacterium smegmatis catalyzed the desaturation of lignoceroyl-CoA to a delta15-monounsaturated derivative in the presence of molecular oxygen and NADPH. This desaturation system was separated by ammonium sulfate fractionation, gel filtration, DEAE-cellulose column chromatographies, and affinity column chromatography on immobilized dye, into three components; a NADPH-oxidase, a ferredoxin-containing fraction and a desaturase, all of which were required for the reconstituted desaturation system for lignoceroyl-CoA. This system was inhibited by FMN and ferrous ions but not by KCN. All of these features clearly distinguish this system from the previously known fatty acid desaturation systems of various origins.

Acyl Coenzyme A↗
PubMed1986

Pyrolysis gas chromatography-mass spectrometry of mycobacterial mycolic acid methyl esters and its application to the identification of Mycobacterium leprae.

Pyrolysis gas chromatography-mass spectrometry of methyl mycolates from 32 species of mycobacteria, including Mycobacterium leprae, was carried out. The mycobacteria could be classified into four groups in respect of the fatty acid ester patterns detected within the range C20 to C26. The applicability of this pyrolysis-gas chromatographic method for identifying M. leprae is discussed.

Chromatography, Gas↗
PubMed1985

Mass-spectrometric identification of trehalose 6-monomycolate synthesized by the cell-free system of Bacterionema matruchotii.

The fluffy layer fraction prepared from Bacterionema matruchotii was found to possess high activity for the biosynthesis of mycolic acids which were bound to an unknown compound by an alkali-labile linkage [T. Shimakata, M. Iwaki, and T. Kusaka (1984) Arch. Biochem. Biophys. 229, 329-339]. To determine the structure of the mycolate-containing compound, it was purified and analyzed by field desorption (FD) and secondary ion mass spectrometry (SI-MS). When non-labelled palmitic acid was used as a precursor in the in vitro biosynthetic system, the underivatized product had a cationized molecular ion, [M + Na]+, at m/z 843 in FD-MS and a protonated ion, [M + H]+, at m/z 821 in SI-MS, corresponding to the quasimolecular ion of trehalose monomycolate (C32:0). In SI-MS, characteristic fragment ions due to cleavage of glycosidic linkages were clearly detected in addition to the molecular ion. If [1-13C]palmitic acid was the precursor, 2 mass unit increases in both the quasimolecular and fragment ions were observed, indicating that two molecules of palmitate were incorporated into the product. alpha-Trehalose was found in the aqueous phase after saponification of the product. By the electron impact mass spectrometry of the trimethylsilylated product, the mycolate was found to be esterified with an hydroxyl group at position 6 of the trehalose molecule. These results clearly demonstrated that the predominant product synthesized by the fluffy layer fraction with palmitate as substrate was 6-monomycolate (C32:0) of alpha-D-trehalose. Because newly synthesized mycolic acid was mainly in the form of trehalose monomycolate instead of free mycolate or trehalose dimycolate, the role of trehalose in the biosynthesis of mycolic acid is discussed.

Actinomycetaceae↗
PubMed1984

Differential growth response of normal human diploid fibroblasts and in vitro transformed human fibroblasts in serum-free defined culture medium.

Two neoplastic human cell lines, WI-38 CT-1 and SUSM-1, which were transformed in vitro with gamma rays and 4-nitroquinoline 1-oxide, respectively, grew continuously in a serum-free defined medium. The defined medium used was a 1:1 mixture by volume of Dulbecco's modified Eagle's medium and Ham's F12 (DF) supplemented with 0.1% bovine serum albumin fraction V, 10 micrograms/ml of transferrin, 1 microgram/ml of insulin, and 5 micrograms/ml of oleic acid. In the case of SUSM-1, 100 micrograms/ml of fetuin were added to cultures when the cells were subcultured. Under these conditions the growth rates of the two transformed human cell lines were almost the same as those in a DF medium containing 10% fetal bovine serum (FBS). In addition, the defined medium permitted the cells to grow indefinitely without a lag period when they were transferred from serum-containing medium into this defined medium, indicating that no selection or adaptation of the cells had occurred. Interestingly, these cell lines did not require for their growth any polypeptide growth factors such as epidermal, platelet-derived or fibroblast growth factors. On the other hand, the control WI-38 cells stopped growing in the defined medium after about 2 divisions. Another control normal cell strain of fibroblasts derived from a human embryo showed a decreased growth rate in the defined medium as compared with that in the DF medium with 10% FBS. These results suggest that the defined medium described here is useful for the selective growth of human cells transformed in vitro after treatment with carcinogens from an untransformed cell population. In addition, the defined medium for transformed human cells should contribute to studies on their growth mechanisms.

Cell Division↗
PubMed1984

In vitro synthesis of mycolic acids by the fluffy layer fraction of Bacterionema matruchotii.

Biosynthetic activity for mycolic acid occurred in the fluffy layer fraction but not in the 5000g supernatant of Bacterionema matruchotii. With [1-14C]palmitic acid as precursor for the in vitro system, the predominant product was identified as C32:0 mycolic acid by radio-gas-liquid chromatographic (radio-GLC) and gas chromatographic/mass spectroscopic analyses; if [1-14C]stearic acid was used, two major radioactive peaks appeared on GLC: one corresponding to the peak of (C34:0 + C34:1) mycolic acids and the other to (C36:0 + C36:1) mycolic acids. By pyrolysis/radio-GLC analysis, C32:0 mycolic acid synthesized by [1-14C]palmitic acid was pyrolyzed at 300 degrees C to form palmitaldehyde (the mero moiety) and methyl palmitate (the branch moiety). The pH optimum for the incorporation of [1-14C]palmitate into bacterionema mycolic acids was 6.4 and the reaction required a divalent cation. The in vitro system utilized myristic, palmitic, stearic and oleic acids (probably via their activated forms) well as precursors, among which myristic and palmitic acids were more effective than the rest. Avidin showed no effect on the biosynthesis of mycolic acid from 14C-palmitate whereas cerulenin, a specific inhibitor of beta-ketoacyl synthetase in de novo fatty acid synthesis, inhibited the reaction at a relatively higher concentration. Thin-layer chromatographic analysis of lipids extracted from the reacting mixture without alkaline hydrolysis showed that both exogenous [1-14C]fatty acid and synthesized mycolic acids were bound to an unknown compound by an alkali-labile linkage and this association seemed to occur prior to the condensation of two molecules of fatty acid.

Actinomycetaceae↗
PubMed1984

Purification of NADPH-dependent enoyl-CoA reductase involved in the malonyl-CoA dependent fatty acid elongation system of Mycobacterium smegmatis.

NADPH-Dependent enoyl-CoA reductase [EC 1.3.1.8] was purified to homogeneity, for the first time, from the crude extract of Mycobacterium smegmatis. The molecular weight of this enzyme was estimated to be around 32,000 using sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The purified enzyme reduced 2-trans-hexadecenoyl-CoA (Km value, 100 microM) and -eicosenoyl-CoA (Km value, 83 microM) almost equally well in the presence of NADPH as a sole electron donor. The Km value for NADPH was 34.5 microM. When NADP3H was incubated with 2-eicosenoyl-CoA and the purified enzyme, the sole tritiated product was arachidate. This enzyme was almost inert to enoyl-CoAs with chains less than 12 carbon atoms long. The purified enzyme still retained FMN, which was detectable by acid ammonium sulfate and was essential for full activity of the enzyme. The enzyme was sensitive to SH-reagents such as N-ethylmaleimide and monoiodoacetamide but was not sensitive to isonicotinamide hydrazide. Anti-NADPH-dependent-enoyl-CoA-reductase rabbit serum was found to inhibit the activities of both the reductase and the malonyl-CoA dependent fatty acid elongation system, supporting the involvement of the reductase in this elongation system.

Acyl Coenzyme A↗
PubMed1983

Purification and characterization of 3-oxoacyl-CoA synthase of Mycobacterium smegmatis.

3-Oxoacyl-CoA synthase, that condenses malonyl-CoA to other acyl-CoAs and takes part in the malonyl-CoA-dependent, acyl carrier protein (ACP)-non-requiring fatty acid elongation system ("fatty acid elongation system II or elongation system II" (Kikuchi, S. & Kusaka, T. (1982) J. Biochem. 92, 839-844)), was purified to homogeneity for the first time from the crude extract of Mycobacterium smegmatis by column-chromatographies. The molecular weight of this enzyme was estimated to be around 64,000 by Sephacryl S-300 gel filtration and 59,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The enzymic product from malonyl-CoA and stearoyl-CoA was identified as 3-oxoeicosanoyl-CoA by mass-spectrometry. Km values of the enzyme for malonyl-CoA and stearoyl-CoA were 41.7 microM and 52.6 microM, respectively. The enzyme was more active toward acyl-CoAs having acyl-carbon-numbers of 18 or more, either saturated or monounsaturated, than those with below 18. Cerulenin, a specific inhibitor of 3-oxoacyl-ACP synthase [EC 2.3.1.41], had no effect on this enzyme but iodoacetamide and N-ethylmaleimide (NEM) showed inhibitory effects.

3-Oxoacyl-(Acyl-Carrier-Protein) Synthase↗
PubMed1983

Gaschromatography of constitutive fatty acids in Mycobacterium leprae.

A constitutive saturated and monounsaturated fatty acid pattern of Mycobacterium leprae, isolated from the liver of a nine-banded armadillo with experimental leprosy, was analyzed gaschromatographically and compared with that of cultured M. lepraemurium, M. avium, M. bovis, strain BCG and M. smegmatis. In comparing the fatty acid pattern thus obtained and the known structure of mycolic acids in these mycobacteria, an experiential rule that each species of mycobacteria has a relatively high content of normal (straight-chained) saturated fatty acid having two more carbons than those of the alpha-branch in this species' mycolic acids, coincided well for all mycobacteria tested. In particular, M. leprae was found to contain a relatively high content of behenic acid (n-C22:0) and the carbon-number of the alpha-branch in this species' mycolic acids is 20 as we previously reported. These data suggested the possibility of simple detection of M. leprae by gaschromatography, and results sustaining this possibility were obtained.

Chromatography, Gas↗
PubMed1982

New malonyl-CoA-dependent fatty acid elongation system in Mycobacterium smegmatis.

Beside de novo fatty acid synthetase, two kinds of fatty acid elongating systems have been found in Mycobacterium smegmatis; they are the malonyl-CoA-dependent, acyl carrier protein (ACP)-requiring system (fatty acid synthetase II, named by K. Bloch's group, 1969) and the acetyl-CoA-dependent ACP-non-requiring one (acetyl-CoA-dependent elongation of fatty acids reported by us, 1977). Now, a third fatty acid elongating system, malonyl-CoA-dependent and ACP-non-requiring, has been isolated from an extract of this microorganisms, separately from each of the previous two elongating systems. Primer specificity and cofactor requirements, especially of pyridine-nucleotide-coenzyme, of the last elongation system also distinguish it from the two previously known systems. All three systems, however, were found in a soluble fraction of M. smegmatis, therefore the regulatory mechanism for these elongation systems should be investigated hereafter.

Acyl Coenzyme A↗