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Biomedical subjects

Y Saeki

Publications and source records attributed to Y Saeki.

285 records · Page 16Linked to original sources
PubMedDate not supplied

High-level expression and characterization of hepatitis B virus surface antigen in silkworm using a baculovirus vector.

The Bombyx mori nuclear polyhedrosis virus (BmNPV) in the silkworm was used successfully for mass production of biologically active foreign genes under the control of the polyhedrin promoter. This system was adapted for the production of large amounts of hepatitis B virus surface antigens (HBsAg). The DNA fragments coded for the middle protein, which is composed of the S protein with the pre-S2 region, were cloned, the signal protein gene of beta-IFN was added, and both were inserted into a cloning vector. After co-transfection with wild-type BmNPV, stable recombinant viruses were isolated by the limiting dilution method. Infected silkworm larvae with the recombinants expressed HBsAg at high levels (400-600 micrograms/ml). These products, consisting of two polypeptides with molecular weights of approximately 25,000 (p25) and glycosylated P25 (GP28), were purified as assembled 22-nm particles. We demonstrated that HBsAg from silkworms consists of S protein with 7 amino acids of Pre-S2.

Amino Acid Sequence↗
PubMedDate not supplied

Production of human alpha-interferon in silkworm using a baculovirus vector.

Microorganisms are generally used for mass production of foreign gene products, but multicellular organisms such as plants have been proposed as an economical alternative. The silkworm may be useful in this context as it can be cultured easily and at low cost. We have therefore developed a virus vector to introduce foreign genes, for example, the gene for human alpha-interferon (IFN-alpha), into silkworms. We used the baculovirus Bombyx mori nuclear polyhedrosis virus (BmNPV) which has a large (greater than 100 kilobases, kb) double-stranded circular DNA genome within its rod-shaped capsid. Baculoviruses have been used previously as vectors for expression of beta-interferon and beta-galactosidase in established cell lines. Although BmNPV has not been used previously as an expression vector, it has an advantage over the baculovirus Autographa californica NPV in that it has a narrower host range and will not grow in wild insect pests in the field. In the present study, the polyhedrin gene encoding the major inclusion body protein of BmNPV was identified by hybridization with complementary DNA and cloned in a plasmid. For insertion of foreign genes, we constructed a recombinant plasmid carrying a polylinker linked to the promoter of the polyhedrin gene, and inserted the IFN-alpha gene into this plasmid. The resulting plasmid and the BmNPV genomic DNA were co-transfected into BM-N cells, and stable recombinant viruses isolated by plaque assay on BM-N cells. The recombinant virus replicated in silkworm larvae, which synthesized as much as 5 X 10(7) units (approximately 50 micrograms) of interferon in their haemolymph.

Amino Acid Sequence↗
PubMedDate not supplied

Analysis of secretion and expression of platelet-derived growth factor in cultured endothelial cells from stroke-prone spontaneously hypertensive rat.

1. We characterized the endothelial cell-derived growth factors of SHRSP and Wistar Kyoto rats (WKY), respectively and found that platelet-derived growth factor (PDGF)-B chain related growth factor constituted a major portion of the mitogenic activity of the conditioned media of endothelial cells from both animals. There were no remarkable qualitative differences between the endothelial cell-derived growth factors of SHRSP and WKY. 2. Northern analysis revealed that the expression of PDGF-B chain was 2-4-fold enhanced in cultured aortic endothelial cells of SHRSP. This enhanced expression of PDGF-B chain, which may be induced under chronic hypertensive conditions, is suggested to contribute to the increase in endothelial cell-derived growth factors reported in this animal.

3T3 Cells↗
PubMedDate not supplied

The peroxidatic function of liver microsomal cytochrome P-450: comparison of hydrogen peroxide and NADPH-catalysed N-demethylation reactions.

Fifteen different secondary and tertiary methyl amines have been examined as substrates for the cytochromes P-450 of rat-liver microsomes to determine the similarities or differences between the NADPH and oxygen-dependent N-demethylation reaction and the reaction occurring in the presence of hydrogen peroxide. No apparent correlation of the rates of formaldehyde formation using the two different conditions of oxidation was observed. The types of cytochromes P-450 were altered by using rat-liver microsomes from animals treated with various inducing agents. No obvious predictable dependence on the animals treated with various inducing agents. No obvious predictable dependence on the type of cytochrome P-450 present was obtained for the hydrogen peroxide-supported peroxidatic reaction. It is concluded that the hydrogen peroxide-dependent N-demethylation reaction occurs by a reaction mechanism distinct from that occurring during the mixed-function oxidase activity of cytochrome P-450 obtained in the presence of NADPH and oxygen.

Animals↗
PubMedDate not supplied

The mechanistic plurality of cytochrome P-450 and its biological ramifications.

The mechanistic plurality of the microsomal cytochrome P-450 enzyme system is illustrated by studies of the oxidative metabolism of benzo[a]pyrene, 3-hydroxybenzo[a]pyrene and arachidonic acid. Rat liver microsomal metabolism of benzo[a]pyrene or 3-hydroxy-benzo[a]pyrene, supported by cumene hydroperoxide, generates benzo[a]pyrene quinones via molecular oxygen-dependent and -independent pathways. Arachidonic acid is metabolized by rat liver microsomal fractions to a variety of oxygenated products, including cis-trans diene conjugated monohydroxy-acids, epoxy-acids as well as omega- and omega-1-oxidation products. The chemistry of the different reaction products is discussed in terms of the possible mechanisms responsible for their formation and the role of the haemoprotein during catalysis. An integrated view for the reaction cycle of cytochrome P-450 is presented.

Animals↗
PubMedDate not supplied

Correlation between metastatic potency and the down-regulation of E-cadherin in the mouse hepatoma cell lines G-1 and G-5.

Cell-cell adhesiveness, involving the adherens junction system including homophilic adhesion of cadherin and intracellular catenins, is a critical factor for tumor cell invasion and metastasis. We evaluated the levels of E-cadherin and beta-catenin in hepatoma cell sublines with high and low metastatic capacities. Stimulation of these cells with serum growth factors for more than 3 h after 24 h of starvation caused decreases in levels of E-cadherin and beta-catenin in the subline with high metastatic capacity, G-5. In contrast, no significant changes were observed in the subline with low metastatic capacity, G-1. Concomitantly with the decreases in E-cadherin and beta-catenin levels, G-5 cells were dissociated and detached from the culture dish, although G-1 cells again showed no morphological alterations. These in vitro results reflected the in vivo metastatic potencies of these hepatoma sublines, and further suggested the importance of the adherens junction system in determining metastatic potency of these parenchymal tumor cell lines as in epithelial/endothelial tumors.

Animals↗
PubMedDate not supplied

Effect of Helicobacter pylori culture supernatant on acute reflux esophagitis in a rat model.

BACKGROUND/AIMS: Prevalence of Helicobacter pylori, especially cagA-positive strains is inversely related to gastroesophageal reflux disease. The aim of this study was to examine whether H. pylori culture supernatants affect acute esophagitis induced by acid or mixed reflux in rats. METHODOLOGY: Three different H. pylori strains were used. Acute esophagitis was induced in 59 male Wistar rats by ligation of both the transitional regions between the forestomach and glandular portion, and the pylorus or the lower part of duodenum. After operation, the rats were either left untreated or treated with intragastric injection of either vehicle or H. pylori culture supernatants and were sacrificed 6 or 24 hours later. Esophagitis index, depth of esophageal lesions, esophageal epithelial degeneration, and infiltration of inflammatory cells were examined. RESULTS: Gross esophageal erosions and ulcers were observed up to mid-esophagus in all animals. H. pylori culture supernatants did not affect esophageal mucosal injury and no histologically significant differences were found among rats. CONCLUSIONS: H. pylori culture supernatants, regardless of cagA gene expression, did not affect acute esophagitis induced by acid or mixed reflux. This finding suggests that no powerful protective factors against reflux-induced mucosal injury were produced by H. pylori.

Acute Disease↗
PubMedDate not supplied

Effects of a new calcium channel blocker, TA 3090, on serum lipoprotein levels in mild to moderate essential hypertension.

The 25 hypertensive patients received 20 to 40 mg of TA 3090 daily for 12 weeks. Blood pressures declined significantly during treatment, from a mean of 162/98 to 145/88 mmHg. There were no significant changes in levels of total or very low-density lipoprotein cholesterol or triglyceride, in levels of low-density lipoprotein cholesterol, high-density lipoprotein (HDL) cholesterol, HDL2 cholesterol, HDL3 cholesterol, or in levels of apolipoprotein (apo) B, C-II, C-III, or E. Apo A-I and A-II levels increased significantly from 130 and 29.2 mg/dl before treatment to 152 and 31.4 mg/dl at 12 weeks. Mean serum creatinine levels decreased significantly from 0.92 to 0.80 mg/dl. No other drug-related changes in laboratory test results or side effects were noted. It is concluded that TA 3090 is a safe and effective treatment for mild to moderate hypertension.

Adult↗