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Biomedical subjects

Z Ogita

Publications and source records attributed to Z Ogita.

At least 19 recordsLinked to original sources

Effects of flavonoids isolated from scutellariae radix on fibrinolytic system induced by trypsin in human umbilical vein endothelial cells.

Studies on the effects of flavonoids isolated from the roots of Scutellaria baicalensis on the fibrinolytic system induced by trypsin in cultured human umbilical vein endothelial cells (HUVECs) showed that baicalein (1) strongly inhibited the reduction of t-PA production and the elevation of PAI-1 production induced by trypsin. The IC50 for PAI-1 production was 3.7 microM. In addition, wogonin (3), oroxylin A (5), skullcapflavone II (6), and 2',5,5',7-tetrahydroxy-6',8-dimethoxyflavone (7) inhibited the elevation of PAI-1 induced by trypsin, though less strongly; their IC50 were 105, 61, 110, and 88 microM, respectively. These findings suggest that baicalein prevents the thrombotic tendency induced by trypsin.

Cell Line↗

Cytotoxicity of NaCl, a stomach tumor promoter, and prevention by rice extract in stomach mucosa of F344 rats.

Cytotoxicity of NaCl and its prevention by rice extract were studied in the pyloric mucosa of male F344 rat stomach after oral administration of rice extract and 2.6 M NaCl. Effect were observed histologically by hematoxylin and eosin staining and the bromodeoxyuridine method. Replicative DNA synthesis (RDS) was assayed by liquid scintillation counter with [3H]thymidine. NaCl (2.6 M) induced destruction of the surface mucous cells within 1 min. RDS and S-phase cells increased significantly (p < 0.01) and to a maximum at 17 h, and returned to control levels 48 h after exposure. Administration of aqueous rice extract 3 h before NaCl exposure reduced the morphological damage to the mucosa and prevented the increase in RDS dose dependently by up to 65% (p < 0.01). These results showed that NaCl induced rapid mucosal damage and cell proliferation in rat stomach mucosa and that rice extract prevented the damage and reduced the increase in RDS.

Animals↗

Molecular cloning and chromosomal mapping of a human protein-tyrosine phosphatase LC-PTP.

We isolated cDNA clones encoding a protein-tyrosine phosphatase (PTP) from a human T cell PEER cDNA library. The predicted open reading frame encodes a approximately 40-kDa protein composed of 360 amino acids and has no apparent hydrophobic segments, suggesting that it is a nontransmembrane PTP, which was designated as LC-PTP (leukocyte PTP). Northern blot analysis revealed that the LC-PTP mRNA was preferentially expressed in a variety of hematopoietic cells and the transcriptional sizes were approximately 4.0 kilobases and approximately 2.9 kilobases. Fluorescent in situ hybridization revealed that the human LC-PTP gene is located on the chromosome region 1q32.1, which is known to be a site associated with chromosomal deletion in malignant lymphomas, where candidate tumor suppressor genes might be present.

Amino Acid Sequence↗

Localization of the gene encoding the alpha subunit of human interleukin-5 receptor (IL5RA) to chromosome region 3p24-3p26.

The chromosomal location of the human gene for the alpha subunit of interleukin-5 receptor (IL5RA) has been determined. The human IL5RA gene was localized to the short arm of chromosome 3 by Southern blot analysis of DNA from a panel of mouse-human hybrid somatic cell lines. The IL5RA gene has been further localized to human chromosome region 3p24-3p26 by in situ hybridization of a molecularly cloned IL5RA cDNA fragment to metaphase chromosomes. The results suggest that the IL5RA locus is unlinked to other members of the hematopoietic receptor family. Assignment of the IL5RA gene to chromosome 3 at bands p26-p24 raises the possibility that it may be altered by certain nonrandom chromosomal abnormalities arising in human hematopoietic malignancies and solid tumors.

Animals↗

A new brain glucosensor and its physiological significance.

The concentration of fibroblast growth factor (FGF), which is found in cerebrospinal fluid (CSF), markedly increases after the start of feeding. Food intake was dose-dependently suppressed by picomole doses of FGF and facilitated by anti-FGF antibody. This suppression was caused by activation of protein kinase C in glucose-sensitive neurons in the lateral hypothalamus. In situ hybridization by use of cDNA showed that acidic (a)FGF was produced in ependymal cells. The ependymal cells released aFGF by responding to glucose increase in CSF after feeding. Released aFGF diffused into the brain parenchyma and was taken by neurons. Passive avoidance was significantly more reliable after aFGF infusion into CSF. Clamping cerebral arteries in the gerbil induced ischemia, which damaged neurons in the CA1 layer of the hippocampus. Pretreatment with aFGF prevented this damage. Thus, aFGF is not only the most potent substance yet found for the suppression of feeding, but it is also extremely effective as a neurotrophic and memory facilitating substance.

Animals↗

Establishment of a new Epstein-Barr virus nuclear antigen-positive B-cell line, BALL-2, with t(8;14) (q24;q32) chromosome abnormality from B-cell acute lymphoblastic leukemia, L2.

A new Epstein-Barr virus nuclear antigen (EBNA)-positive B-cell line, designated BALL-2, was spontaneously established from the peripheral blood of a 14-year-old boy with an EBNA-negative B-cell acute lymphoblastic leukemia (B-ALL), L2 in the French-American-British classification. The BALL-2 cell line grew in suspension with or without forming clumps of cells. The cultured cells exhibited lymphoid morphology with indented or lobulated nuclei, prominent nucleoli, and relatively abundant cytoplasm. Immunologic and cytogenetic studies showed that the BALL-2 cell line expressed the B-cell phenotype, CpIg+, SmIg+, CD19+, CD20+, CD38-, Ia+, and had chromosome translocation, t(8;14) (q24;q32). The same phenotypic and chromosome markers were present in original leukemia cells. These results indicated that the cell line was derived from the patient's leukemia cells. Unexpectedly, however, BALL-2 cells were positive for EBNA and EB virus DNA. Gene analysis of the BALL-2 cell line showed biallelic rearrangements in the JH locus. One of the JH rearrangement comigrated with a rearranged c-myc gene, indicating the translocation had occurred between JH and c-myc loci. The t(8;14) abnormality is a known chromosome marker of Burkitt lymphoma and L3 type ALL. Our studies revealed that this translocation and myc gene rearrangement can also be found in L2 type B-ALL.

Adolescent↗

Classification of mouse submaxillary gland esteroproteases by their substrate specificities.

The submaxillary gland esteroproteases were separated by two-dimensional polyacrylamide gel electrophoresis, and their substrate specificities were determined by histochemical staining procedures using cellulose acetate membranes. Twenty-one proteolytically active enzymes were classified into four groups based on substrate specificities on benzoyl-arginine-ethylester (BAEE), N-alpha-benzoyl-DL-arginine-p-nitroanilide (BAPNA), and poly-L-lysine. These types were further divided into eight subgroups by their sensitivity to inhibitors and androgen dependence. These results suggest that eight groups of esteroproteases are present in the submaxillary gland of male mice.

Animals↗

Molecular cloning of a mouse DNA repair gene that complements the defect of group-A xeroderma pigmentosum.

For isolation of the gene responsible for xeroderma pigmentosum (XP) complementation group A, plasmid pSV2gpt and genomic DNA from a mouse embryo were cotransfected into XP2OSSV cells, a group-A XP cell line. Two primary UV-resistant XP transfectants were isolated from about 1.6 X 10(5) pSV2gpt-transformed XP colonies. pSV2gpt and genomic DNA from the primary transfectants were again cotransfected into XP2OSSV cells and a secondary UV-resistant XP transfectant was obtained by screening about 4.8 X 10(5) pSV2gpt-transformed XP colonies. The secondary transfectant retained fewer mouse repetitive sequences. A mouse gene that complements the defect of XP2OSSV cells was cloned into an EMBL3 vector from the genome of a secondary transfectant. Transfections of the cloned DNA also conferred UV resistance on another group-A XP cell line but not on XP cell lines of group C, D, F, or G. Northern blot analysis of poly(A)+ RNA with a subfragment of cloned mouse DNA repair gene as the probe revealed that an approximately 1.0 kilobase mRNA was transcribed in the donor mouse embryo and secondary transfectant, and approximately 1.0- and approximately 1.3-kilobase mRNAs were transcribed in normal human cells, but none of these mRNAs was detected in three strains of group-A XP cells. These results suggest that the cloned DNA repair gene is specific for group-A XP and may be the mouse homologue of the group-A XP human gene.

Animals↗

[Anti-ulcer effect of extract from phellodendri cortex].

In chinese medicine, Phellodendri Cortex (Phellodendron amurense Ruprecht) has been used to treat the patient who suffers from gastroenteritis, abdominal pain or diarrhea. Berberine has been identified as a major component in this plant, and it has biological activities, such as bactericidal activity, anti-cholera toxin effect, anti-inflammatory effect, stimulative effect of bile secretion or bilirubin discharge. In the previous study, we have shown the presence of anti-inflammatory activity in the berberine-free fraction of the extract from this plant. In the present study, we also found anti-ulcer activity in the fraction. The fraction significantly inhibited the formation of ethanol-induced ulcer, aspirin-induced ulcer (s.c., p.o.), pylorus-ligated ulcer (p.o., i.d.) in rats, as well as that of stress ulcer in restrained and water-immersed mice (p.o.). In addition, gastric acid secretion was significantly reduced in pylorus-ligated rats by subcutaneous or intraduodenal administration of the fraction, but not by oral administration. These findings suggest that the suppression of ulcer formation may be due to the additive effect of the cytoprotection effect and the reduction of gastric acid secretion by administration of the berberine-free fraction.

Animals↗

Transgenic mouse carrying a foreign gene on its Y-chromosome.

In an experiment in which a human A gamma/beta-globin construct gene was introduced into the germ line of mice by microinjection, we obtained a transgenic male mouse carrying the construct gene on its Y-chromosome. This mouse carried 13 copies of a 8.9 kb construct gene as a head-to-tail tandem array, and showed a low level of transcription of the human A gamma- and beta-globin gene in the erythroid tissues. This transgenic mouse can be a useful tool in studies of embryology and developmental genetics because it carries a specific DNA marker on only its Y-chromosome.

Animals↗

Experimental production of transgenic mice carrying human beta-globin genes.

To produce transgenic mice carrying human beta-globin genes, we introduced the following two constructs of the genes to male pronuclei of fertilized mouse eggs: 4.4 kb Pst I/Pst I sequences of the human beta-globin gene (experiment 1) and the human beta-globin gene cluster (cosHG 28) containing G gamma, A gamma, delta and beta-globin genes and cosmid vector pJB8 (37.5 kb, experiment 2). In experiment 1, 25 mice were born, and four (one female and three males) carrying the injected gene sequences were identified. One of these mice carried the entire sequence of the human beta-globin gene but three others appeared to carry only a part of the entire sequence. The mouse with the entire sequence showed a slight increase in the minor component of the mouse beta-globin chain in the same position as the human beta-globin chain. In experiment 2, 61 mice were born, and nine (three females and six males) carried the sequences of the injected gene. However, from DNA analysis, no appropriate sequences present within the A gamma- or beta-globin gene were identified in any of the founder mice. In this case, DNA fragments of the gene cluster that were digested in the mouse nucleus after microinjection of the gene might be integrated into host DNA.

Animals↗

Appearance of extrachromosomal circular DNAs during in vivo and in vitro ageing of mammalian cells.

Appearance of extrachromosomal circular DNAs with in vivo and in vitro cellular ageing was examined by using a new technique of mica-press-adsorption for electron microscopy. The size distribution and the copy number of circular DNA complexes varied, depending on the cellular age. Extrachromosomal circular DNA complexes of variable length of more than 0.5 microns or 1.5 kilobase (kb) appeared during in vivo ageing of rat lymphocytes and in vitro ageing of cultured human lung fibroblasts. A restricted size class of circular forms of less than 0.5 microns in contour length was amplified in human skin fibroblasts from aged normal or Werner's syndrome subjects. These circular DNA molecules are suggested to be products of DNA rearrangements or gene amplification occurring in the chromosome.

Aging↗

An improved method for destroying mouse blastomeres electrically inside the zona pellucida and the in vitro development of the surviving blastomeres.

An improved method for isolating mouse blastomeres by electrically destroying the other blastomeres inside the zona pellucida is described. Instruments required are two micromanipulators connected to an inverted microscope and an electronic stimulator equipped with an isolator. The target blastomere in which a glass microelectrode is inserted disintegrates completely within a few seconds after a direct current is applied. Complete destruction of a blastomere requires more than 60 microA delivered at 8 V. Destruction of blastomeres occurred mainly as a result of the medium penetrating the blastomere due to the change of membrane potential in both the zona and the blastomere. Most one-half, two-quarter, and three-quarter embryos obtained in this way developed in vitro into normal blastocysts (76.4--86.2%). By contrast, there was a marked decrease (35.3%) of developmental potential in one-quarter embryos. The results indicate that the present method is an improvement over previous methods of obtaining mammalian blastomeres inside the zona pellucida.

Animals↗

Electrophoretic analysis of pancreatic proteases and zymogen-activating factors in the mouse.

Mouse pancreatic proteases were analyzed by one- and two-dimensional electrophoresis. Active proteases that existed in the luminal fluid were separated into at least eight bands in 8% polyacrylamide gel. Pancreatic proteases activated by intestinal extract were separated into at least seven bands. The mobilities of these bands were exactly the same as those of proteases in the luminal fluid except for those of the most cathodal band. Two kinds of trypsin (Try-I group and Try-II) and one kind of chymotrypsin (Chy-I) were determined by specific and nonspecific protease staining. Try-I group and Try-II were derived from different trypsinogens (Try G-I group and Try G-II), whereas Chy-I was derived from a single chymotrypsinogen (Chy G). Although Try G-II was activated by both intestinal extract and by bovine trypsin, Try G-I group activated only by intestinal extract. Intestinal-activating factors were analyzed by two-dimensional electrophoresis. Mouse enterokinase (enteropeptidase EC 3.4.4.8), which can activate bovine trypsinogen, had a slow mobility. In the intestine of the mouse there are several activating factors in addition to enterokinase. Although it is unclear what intestinal-activating factors can activate Chy G, there is a factor that can convert chymotrypsinogen into chymotrypsin directly. These data suggest that intestinal-activating factors play an important role in the activating mechanisms of mouse pancreatic zymogens.

Animals↗

Two-dimensional gel analysis of zymogen-activating factors in the small intestine of the mouse.

Zymogen-activating factors in the mouse were investigated by two-dimensional electrophoresis. Mouse pancreatic zymogens--trypsinogen-I group (Try G-I group), trypsinogen-II (Try G-II), and chymotrypsinogen (Chy G)--were purified using DEAE-cellulose column chromatography. Analysis by two-dimensional electrophoresis, using the purified zymogens as substrates, revealed enterokinase isozymes and chymotrypsinogen-activating factors in both the intestinal extract and luminal fluid. Mouse enterokinase was separated into at least two bands in the first-dimensional gel, each able to activate both trypsinogens Try G-I group and Try G-II. Chymotrypsinogen-activating factors were separated into several bands in the first-dimensional gel. Some activating factors showed mobilities similar to those of mouse enterokinase isozymes. Moreover, other activating factors that can activate chymotrypsinogen were present only in the more anodal area of the first-dimensional gel. These findings indicate that at least two enterokinases and several chymotrypsinogen-activating factors play an important role in the process of activating digestive enzymes.

Animals↗

Isoelectric focusing of the inbred mouse antibody to bacterial alpha-amylase.

In the IgG antibody response to bacterial alpha-amylase (B alpha A) assayed by the enzymatic procedure, C3H/He (C3) mice were high and C57BL/6 (B6) mice were low responders. High responsiveness was inherited as a dominant characteristic in (B6XC3)F1 hybrid mice. In these strains, the primary antibody response was analyzed for heterogeneity by isoelectric focusing (IEF). The IEF spectra were visualized with the use of the capacity of antibody to inhibit the amylase activity of antigen. Increases in the antigen dose and in the time interval between immunization and bleeding resulted in increases in antibody titers accompanied by strong staining of focused antibodies and by the expansion of the pH range where antibodies were focused. High responsiveness in C3 and F1 hybrid mice was also associated with the increase in intensity of stain and the rapid expansion of pH range of focused antibodies. Another strain difference was noted in the isoelectric point (pI) values of antibodies taken early in the primary response. B6 antisera contained those fractions of antibodies focusing over a more alkaline area than C3 antibodies. A similar strain difference in the pI values of antibodies occurred in the response to an irrelevant antigen, Taka-amylase A (TAA), suggesting that the hypervariable regions of antibody molecules play no major part in the strain difference observed. Antisera from F1 hybrid mice displayed bands covering the combined pH ranges of B6 and C3 spectrotypes.

Animals↗