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Y Nagata

Publications and source records attributed to Y Nagata.

At least 289 records · Page 16Linked to original sources
PubMed1995

Substrate specificity of beta 1,4-N-acetylgalactosaminyltransferase in vitro and in cDNA-transfected cells. GM2/GD2 synthase efficiently generates asialo-GM2 in certain cells.

The substrate specificity of beta 1,4-N-acetylgalactosaminyltransferase has been analyzed using a fusion enzyme which consisted of the catalytic domain of the enzyme and the IgG binding domain of protein A, and also by extracts from cDNA transfectants. Both enzyme sources were capable of producing not only GM2 and GD2, but also asialo-GM2, GalNAc-sialylparagloboside, and Gal-NAc-GD1a from appropriate acceptors, although the efficiencies were at most 1-3% of those of GM2/GD2. The biological significance of these low specificities was studied with transient and stable transfectant cells. From the results of transient expression of the cDNA, asialo-GM2 expression appeared to inversely correlate with GM2 synthase levels in those lines. Consequently, GM2 seemed to be preferentially synthesized when both GM3 and lactosylceramide are available, and asialo-GM2 is synthesized in the absence of GM3 synthesis. However, the results of double immunostaining of CHO transfectants with anti-GM2 and anti-asialo-GM2 antibodies indicated that another factor may be involved in asialo-GM2 synthesis. From the in vitro assay using mixed acceptors, it was concluded that the presence of certain levels of GM2 might enhance the asialo-GM2 synthesis. These results suggest that even acceptors showing low efficiencies in vitro might be used in certain cells depending on the availability of precursors, expression levels of other gangliosides, as well as the kinetic properties of the enzyme, and the compartmentation of the glycosylation machineries in the cells.

Animals↗
PubMed1995

N-[2-[N'-pentyl-(6,6-dimethyl-2,4-heptadiynyl)amino]ethyl]- (2-methyl-1-naphthylthio)acetamide (FY-087). A new acyl coenzyme a:cholesterol acyltransferase (ACAT) inhibitor of diet-induced atherosclerosis formation in mice.

FY-087 (N-[2-[N'-pentyl-(6,6-dimethyl-2,4-heptadiynyl)amino]ethyl]- (2-methyl-1-naphthylthio)acetamide) was found to be a competitive inhibitor of human microsomal acyl coenzyme A:cholesterol acyltransferase (ACAT) with an IC50 value of 0.11 microM. Under our assay conditions, other ACAT inhibitors tested, specifically YM-750, E-5324, and melinamide, all of which are now in phase I clinical trials or in clinical use in Japan, inhibited this enzyme with IC50 values of 0.18, 0.14, and 3.2 microM, respectively. FY-087 also inhibited ACAT in acetyl-low density lipoprotein loaded human macrophages (THP-1 cells) with an IC50 of 0.17 microM. Following the oral administration of FY-087 (30 mg/kg) to rats, the plasma concentration of FY-087 reached 0.42 microgram/mL after 2 hr. This concentration of FY-087 was enough to inhibit blood vessel ACAT activity. Cholesterol-lowering and anti-atherogenic effects of FY-087 were examined using C57BL/6J mice fed an atherogenic diet. In this mouse model, treatment with FY-087 (28 mg/kg) inhibited the increase in plasma cholesterol levels by about 20% and decreased the hepatic accumulation of free and esterified cholesterol by 61 and 67%, respectively. FY-087 also significantly inhibited the atherogenic diet-induced increase in the fatty-streak lesion area of the proximal aorta by 57% in C57BL/6J mice. These results indicate that FY-087 is not only a therapeutically bioavailable ACAT inhibitor that lowers plasma cholesterol levels, but also an effective anti-atherogenic agent in mice fed an atherogenic diet.

Acyl Coenzyme A↗
PubMed1995

Phosphorylation of helix-loop-helix proteins ID1, ID2 and ID3.

Id is a helix-loop-helix protein which forms heterodimer with ubiquitous and/or tissue-specific basic helix-loop-helix proteins and inhibits their DNA binding. It has been noted that putative phosphorylation sites for various protein kinases exist in rat Id1, Id2 and Id3. We show here that Id1 and Id2 can be phosphorylated in vitro by cAMP-dependent protein kinase, Id2 and Id3 by cdc2 kinase, and all three Ids by protein kinase C. The phosphorylated Id1 was actually immunoprecipitated in nerve-growth-factor-stimulated PC12 cells. Gel mobility shift assays, however, demonstrated that neither phosphorylation of Id proteins by cAMP-dependent protein kinase nor phosphorylation of E47 by protein kinase C affected the inhibition of E47 homodimer formation and its DNA binding. Taken together with other observations, phosphorylation of Id proteins may play a role in regulation of cell differentiation but not directly in the dimerization and DNA binding.

Amino Acid Sequence↗
PubMed1995

Determination of portal short-chain fatty acids in rats fed various dietary fibers by capillary gas chromatography.

A simple, rapid and sensitive capillary gas chromatographic method was investigated to measure portal short-chain fatty acids (SCFAs). A 20-microliters sample of portal plasma was denatured with sulfosalicylic acid and then extracted with diethyl ether before the removal of protein precipitate. The resultant extract was concentrated by a transfer to 50 microliters of 0.2 M NaOH, thus avoiding tedious further concentration steps. This reduced the sample volume to one-fourth. Since the ratio of acetic acid, a major SCFA, to other acids varies widely, ranging from 10-fold to 100-fold, acrylic and methacrylic acids were used as internal standards to simultaneously measure SCFAs having a carbon number of 2-6. As a result, good recovery (90.38-103.17%) and reproducibility (coefficient of variation 0.83-8.85%) were observed over a wide range. Furthermore, portal SCFAs in rats fed various dietary fibers were determined by the present method. We showed that the amounts not only of the major acids such as acetic acid and propionic acid, but also of the minor fermented products such as n-valeric acid and n-caproic acid, could be significantly changed by dietary manipulation. Thus, the present method is simple and reliable, and requires only a small amount of sample.

Animals↗
PubMed1995

Prodrugs of vitamin E. 1. Preparation and enzymatic hydrolysis of aminoalkanecarboxylic acid esters of d-alpha-tocopherol.

Nine aminoalkanecarboxylic acid esters of d-alpha-tocopherol were synthesized and evaluated as potential water-soluble prodrugs suitable for parenteral administration. The hydrochloric acid salts of the esters were soluble in water. The kinetics of hydrolysis of the esters was studied in isotonic phosphate buffer, rat plasma, human plasma, and rat liver homogenate at 37 degrees C. The hydrolysis of the esters was proved to be catalyzed by liver esterases. The susceptibility of the esters to undergo liver esterase hydrolysis was affected by the structure of the amino functionality and size of the acyl moiety on the promoiety. The N-methylaminoacetyl and N,N-dimethylaminoacetyl esters of d-alpha-tocopherol were more rapidly hydrolyzed than d-alpha-tocopheryl acetate, a commercially available d-alpha-tocopheryl ester. These results suggested that the salts of the N-methylaminoacetyl and N,N-dimethylaminoacetyl esters are promising prodrug candidates of d-alpha-tocopheryl for parenteral use.

Animals↗
PubMed1995

Alteration of transglutaminase activity in rat and human spinal cord after neuronal degeneration.

We measured the activity of transglutaminase (TG), a Ca(2+)-dependent enzyme and a biochemical marker of cell degeneration, in the adult rat spinal cord after unilateral occlusion of a branch of the dorsal spinal artery, and compared it to the enzyme activity in the tissue on the contralateral side without ischemic damage. The affected half of the spinal cord showed a significant rise in intrinsic (endogenous) TG activity one day after ischemic insult while no apparent morphological changes were observed in the tissue. However, the enzymic activity on the affected side gradually decreased to reach the level in the non-affected tissue, accompanying severe degeneration of neuronal cells at 7 days after the surgery, then it declined to nearly half the level in the intact tissue 30 days after the operation. We also determined the TG activity in transverse sections of the human spinal cord obtained at autopsy from 5 amyotrophic lateral sclerosis (ALS) and 9 non-ALS patients. TG activity in thoracic and lumbar cords was markedly low in ALS patients not only in ventral and lateral regions but also in the dorsal portion. These findings imply that the reduced TG activity in the ALS spinal cord is one of the characteristic features of the disease reflecting exhaustion of the enzyme in the tissue resulting from degeneration of the spinal neurons through cross-linkage of soluble intraneuronal cytoplasmic proteins.

Adult↗
PubMed1995

Clinical evaluation of 430 MHz microwave hyperthermia system with lens applicator for cancer therapy.

The clinical efficacy of a microwave (MW) hyperthermia system using an electric-field converging (lens) applicator is evaluated for 42 malignant tumours with a maximum tumour depth of less than 7 cm. The mean of the maximum, average and minimum tumour temperature of the 42 tumours are 44.5, 42.5 and 40.7 C, respectively. The thermal parameters are higher for tumours in the chest, abdominal walls and hip than for those in the neck, groin and extremities. No apparent difference in thermal parameters according to the depth of tumour is shown. Of 40 tumours treated by hyperthermia in combination with radiotherapy, 20 (50%) showed complete regression, 14 (35%) showed partial regression, and six (15%) showed no change. This phase I and II study indicates clinical feasibility of the newly developed MW heating apparatus, and strongly suggests the usefulness of thermoradiotherapy in the treatment of localised superficial and subsurface malignancies.

Adolescent↗
PubMed1995

Body fat distribution in women with polycystic ovary syndrome.

OBJECTIVE: To investigate body fat distribution in women with polycystic ovary syndrome (PCOS). METHODS: Body weight, body mass index (BMI), and six indices of body fat measured by dual-energy x-ray absorptiometry were compared in 40 women with PCOS and 97 age-matched controls. The possible correlations between the body fat characteristics and serum androgen levels were evaluated in the 40 PCOS women. Body fat distribution was classified into upper- (N = 24) and lower-half body type (N = 16), and androgen levels and the incidence of hirsutism were compared in the two types. RESULTS: The BMI, body fat ratio, upper-half body fat ratio, and upper-half/lower-half body fat ratio were significantly higher in PCOS women than in controls. After adjustment for age, height, and body weight, the upper-half/lower-half body fat ratio was still significant (P < .001). The PCOS subjects exhibited a significant positive correlation between the upper-half/lower-half body fat ratio and dehydroepiandrosterone-sulfate (DHEA-S) levels (r = 0.607, P < .01) as well as testosterone levels (r = 0.585, P < .05). Dehydroepiandrosterone-sulfate and testosterone levels were significantly higher in those with the upper-half body type than in those with the lower-half body type (P < .001). After adjustment for confounding variables, only DHEA-S was still significantly higher in this body type (P < .05). CONCLUSION: Serum DHEA-S levels seem to be associated with upper-half body fat distribution in women with PCOS, irrespective of body weight.

Adipose Tissue↗
PubMed1995

Angiographic extravasation of contrast medium in acute "spontaneous" subdural hematoma.

Acute spontaneous subdural hematoma is very rare. We have encountered four such cases and verified the arterial origin of the bleeding at operation. None of the patients had a history of head trauma, and each had developed sudden onset of headache and other neurologic deficits, which simulate other cerebrovascular diseases. CT directly revealed subdural hematoma but gave no indication as to the source of the bleeding. Cerebral angiography was performed in all cases, with three of them showing localized extravasation of the contrast material into the subdural space. The extravasation was noted usually in the late arterial phase. This is a useful finding for diagnosing this disease and localizing the bleeding point. It is expected that with more routine use of cerebral angiography in cases of acute spontaneous subdural hematoma, extravasation of the contrast medium will be seen more frequently.

Acute Disease↗
PubMed1995

Possible involvement of nitric oxide in carbachol-induced activation of transglutaminase in rat superior cervical sympathetic ganglia.

The addition of a muscarinic agonist, carbachol (Carb, 0.1 mM), to a physiological medium markedly increased Ca(2+)-dependent transglutaminase (TG) activity (approximately 10-fold) in isolated rat superior cervical sympathetic ganglia (SCG) following in vitro aerobic incubation for 30 min at 37 degrees C. The Carb-evoked stimulation of ganglionic TG activity was considerably reduced (-51%) in the presence of NG-monomethyl-L-arginine (L-NMMA, 50 microM), a selective inhibitor of nitric oxide (NO) synthase. While the suppressant effect of L-NMMA was completely eliminated by the addition of an excess concentration of L-arginine (0.5 mM), a precursor of NO. These observations imply that Carb-induced TG activation possibly involves NO mediation in SCG tissue. The Carb-induced elevation in ganglionic TG activity was markedly reduced (-84%) at as early as 15 min of incubation in the medium containing hemoglobin (Hb) (20 microM), an agent that scavenges only extracellular NO gas. Thus, it is evident that a large fraction of NO released from inside the neuronal cells to extracellular space could rapidly diffuse back into the same group of cells to induce activation of the tissue TG. Methylene blue (MB), an inhibitor of soluble guanylate cyclase (GC), at 0.5 mM, a concentration which is effective in almost abolishing the Carb-evoked synthesis of cyclic GMP (cGMP), had no effect on ganglionic TG activation induced by Carb. Therefore, an increase in cGMP synthesis mediated by NO might not participate in NO-dependent ganglionic TG activation following the stimulation with Carb.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

Free D-serine concentration in normal and Alzheimer human brain.

We have analyzed both free L- and D-serine in frontal cortex of normal and Alzheimer human brain by high-performance liquid chromatography (HPLC). There was no significant difference between the two brains. In normal brain, L- and D-serine concentrations were 666 +/- 222 and 66 +/- 41 nmol/g of wet tissue, respectively, and the ratio of D-isomer to L-isomer (D/L) was 0.099 +/- 0.031. In Alzheimer brain, the concentrations were 750 +/- 150 and 66 +/- 40 nmol/g, respectively, and the D/L ratio was 0.086 +/- 0.040. Thus, it was shown that the free D-serine concentration in the Alzheimer brain was comparable to that in the normal brain.

Aged↗
PubMed1995

The role of p53 inactivation in human cervical cell carcinoma development.

We investigated the association between human papillomavirus (HPV) infection and p53 gene mutation in 47 primary uterine cervical cancers. HPV DNA sequences were present in 43 cancers (91.5%), and one of these cancers contained a p53 gene mutation. In addition, one of the remaining four HPV-negative cancers also contained a p53 gene mutation. As a result, p53 inactivation corresponded to the development of 44 of the primary uterine cervical cancers studied (93.6%). We obtained both primary and recurrent tumours from four cases. In two of these cases, the HPV genomes that were present in an episomal state in the primary tumours were observed to have disappeared in the recurrent tumours. One of these recurrent tumours also contained a p53 gene mutation, which suggested the possibility that p53 inactivation was required in order to maintain the aggressive behaviour in this cancer either by an HPV infection or by a p53 gene mutation. No MDM2 gene amplification was observed in the tumours that carried neither HPV DNAs nor p53 gene mutations.

Adenocarcinoma↗
PubMed1995

Effects of embryo density and co-culture of unfertilized oocytes on embryonic development of in-vitro fertilized mouse embryos.

We have evaluated the effects of embryo density and the co-culture of unfertilized (degenerating) oocytes on the development of in-vitro fertilized (IVF) mouse embryos. In experiment 1, groups of one, five, 10 or 20 zygotes were cultured in 20 microliter drops of modified human tubal fluid (HTF) medium for 168 h at 38.7 degrees C in 5% CO2 and 95% air. As the embryo density increased, significantly (P < 0.05) higher rates of embryos reached hatched blastocyst stage. In addition, the time required for hatching after IVF was significantly (P < 0.05) shortened by the increase in embryo density. In experiment 2, 10 IVF zygotes were cultured with or without 10 unfertilized (degenerating) oocytes in 20 microliter drops of HTF medium. The rates of IVF embryos that developed to morula, blastocyst, expanded blastocyst and hatched blastocyst stages were decreased significantly (P < 0.01) by culturing embryos with unfertilized oocytes compared with culturing embryos alone. In experiment 3, groups of one or 10 IVF zygotes or 10 IVF zygotes plus 10 unfertilized oocytes were cultured in 20 microliter drops of HTF medium and the number of cells per blastocyst was examined at 120 h after IVF. Increasing embryo density resulted in a significant (P < 0.05) increase in the number of cells per blastocyst. In contrast, the cell number of IVF embryos that developed to blastocyst decreased significantly (P < 0.05) when they were cultured with unfertilized oocytes. The results suggest that in-vitro development of IVF mouse embryos is enhanced by increasing embryo density and is impaired by co-culture with unfertilized (degenerating) oocytes.

Animals↗
PubMed1995

A 6-kb upstream region of the human transthyretin gene can direct developmental, tissue-specific, and quantitatively normal expression in transgenic mouse.

To ascertain whether a 6-kb upstream region of the human transthyretin (TTR) gene contains the cis-element(s) required for proper specificity and level of expression, transgenic mice carrying the human mutant TTR gene containing either 6-kb (6.0-hMet30) or 0.6-kb (0.6-hMet30) of the upstream region were produced and studied. The 6.0-hMet30 gene was expressed in the yolk sac, liver, and choroid plexus, where the mouse endogenous TTR gene is also expressed. In contrast, expression of the 0.6-hMet30 gene was restricted to the yolk sac and liver. The expression levels of the 6.0-hMet30 gene in the liver and serum were similar to those of the mouse TTR gene, and about 10-fold those of the 0.6-hMet30 gene. Before birth, the developmental profiles of the expression of both transgenes in each tissue were similar to those of the mouse TTR gene. However, the expression levels of the 6.0-hMet30 gene in the liver and serum increased after birth to reach adult levels at an age of 4 weeks, while expression of the 0.6-hMet30 gene remained at a low level after birth. These results suggest that the 6-kb upstream sequence contains the cis-elements required for developmental, tissue-specific, and quantitatively normal expression.

Animals↗
PubMed1995

A maternal risk factor for mother-to-child HTLV-I transmission: viral antigen-producing capacities in culture of peripheral blood and breast milk cells.

We examined the relationship between productivity of HTLV-I antigen-positive cells in cultured peripheral blood mononuclear cells (PBMC) and breast milk mononuclear cells (BMMC) and the incidence of mother-to-child transmission of HTLV-I. Among 61 cases of HTLV-I carrier mothers, 17 cases were revealed to produce large numbers of HTLV-I antigen-positive cells (high HTLV-I antigen-producing mothers) whose positive rate was 9.6% in PBMC and 10.2% in BMMC, while the remaining 44 cases produced small numbers of HTLV-I antigen-positive cells (low HTLV-I antigen-producing mothers) whose positive rate was 0.3% in PBMC and 0.5% in BMMC. The HTLV-I transmission rate among children born to the high HTLV-I antigen-producing mothers was 37.5% (6/16 children from 11 mothers), while that of the low HTLV-I antigen-producing mothers was 3.2% (1/31 children from 20 mothers). The transmission rate of HTLV-I was significantly different between high and low HTLV-I antigen-producing mothers (P < 0.05). However, there was no positive relationship between anti-HTLV-I antibody titers and productivity of HTLV-I antigen-positive cells (P = 0.11). These results suggested that mother-to-child transmission of HTLV-I might be influenced by a maternally determined factor to produce HTLV-I antigen-positive cells in PBMC and BMMC of HTLV-I carrier mothers.

Adult↗
PubMed1995

A quantative in vivo method of analyzing human tumor-induced angiogenesis in mice using agarose microencapsulation and hemoglobin enzyme-linked immunosorbent assay.

This study was conducted to develop a quantitative assay system for use in the in vivo evaluation in mice of angiogenesis induced by human tumor cells. The human epidermoid carcinoma cells, A431 cells, were cultured on microcarriers. Microcarrier-attached A431 cells (A431-MC) were microencapsulated with agarose hydrogel to isolate them from the immune system of the C57BL/6 mice after subcutaneous dorsal midline implantation. The agarose hydrogel-microencapsulated A431 cells (Aga-A431 cells; diameter=300 micron) survived for at least 10 days in vitro, and the proliferation profile of the Aga-A431 cells was indistinguishable from that of non-microencapsulated A431 cells. The Aga-A431 cells were subcutaneously injected into mice with an 18-gauge needle. Ten days later, few vessels had formed at the site implanted with cell-free agarose beads, whereas notable angiogenesis was observed at the site implanted with Aga-A431 cells. The degree of angiogenesis was evaluated by measurement of the hemoglobin content in the implanted site using a mouse hemoglobin (mHb) enzyme-linked immunosorbent assay (ELISA) system. This mHb-ELISA system has the advantages of great simplicity and reproducibility. The measured mHb-content of new blood vessels at the site implanted with agarose beads was in good agreement with the amount of angiogenesis observed under a stereoscopic microscope. This assay system enabled us to evaluate the angiogenesis induced by xenogeneic cells, such as human tumor cells. Thus, our novel method may be useful for the study of the angiogenic potential of various human tumor cells and in research on the anti-angiogenic properties of various agents.

Animals↗