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11.
Noriko Usui Kouji Matsushima Anne M. Pilaro Dan L. Longo Robert H. Wiltrout 《Biotherapy》1996,9(4):199-208
Recombinant human interleukin 1α (rh IL-1α) and etoposide (VP-16) synergize for direct growth inhibition of several human
tumor cell linesin vitro. Our previous studies demonstrated that VP-16 increased the number of membrane-associated IL-1 receptors (IL-1Rs) and also
enhanced the internalization of receptor-bound rh IL-1α. The purposes of this study were to test our hypothess that these
events were critical to the synergy between rhIL-1α and VP-16, to determine whether rhIL-1α and VP-16 synergize to increase
superoxide (SO) anion radical productionin vitro since SO anion has been implicated in the toxic effects of IL-1, and to investigate the antitumor efficacy of the combinaton
against tumors in vivo. A375/C6 melanoma cells and OVCAR-3 ovarian carcinoma cells were tested with IL-1 receptor antagonist
(IL-1ra) before exposure to rhIL-1α, VP-16 and rhIL-1α plus VP-16. The synergistic or antagonistic effects were assessed by
MTT assay. SO production was measured by reduction of cytochrome C. Athymic female mice bearing the A375/C6 melanoma were
treated by rhIL-1α, VP-16, and rhIL-1α+VP-16. The antitumor effects were evaluated by quantitating tumor growth and survival
time. Pretreatment with the IL-1ra abrogated the synergistic effects of rhIL-1α and VP-16. The production of SO radical by
A375/C6 cells was increased 2.5 fold by the combination of rhIL-1α and VP-16, and the addition of exogenous SOD blocked the
synergy between rhIL-1α and VP-16. However, when A375/S0D15 cells which over-expressed manganese superoxide dismutase (MnSOD)
after MnSOD cDNA transfecton were exposed to rhIL-1α and VP-16, in vitro antagonism was observed. In vivo studies demonstrated
that the combination of rhIL-1α and VP-16 delayed tumor growth better than either agent alone, although long-term survival
was not improved because of substantial toxicity. Our results suggest that the synergistic antitumor effects of IL-1α and
VP-16 may be due to IL-1R modulation and increased internalization of IL-1-IL-1R complex by VP-16 treatment, as well as to
a subsequent increase in SO anion radical production from the tumor cells exposed to both drugs. Thus, the combnation of IL-1α
and VP-16 might prove useful for the treatment of malignant diseasein vivo, if the increased toxicity can be reduced or managed.
The US Government’s right to retain a non-exclusive royalty-free license on and to any copyright is acknowledged. 相似文献
12.
Feasibility of the combination of the immunogold-silver staining method (IGSS) and computer-aided image analysis was assessed for the detection of antigen in an immunostained, paraffin-embedded section. Using low-temperature IGSS, we stained a specimen of human oral squamous cell carcinoma with a monoclonal antibody, PC 10, against a proliferating cell nuclear antigen (PCNA/cyclin), and the section was analyzed by ACAS 570 interactive laser cytometry. The PCNA-positive cells, exhibiting a heteromorphic texture, were contrasted by the dark staining of their nuclei, but showed heterogeneity in staining intensity from cell to cell. Using a conventional microscope light source rather than a laser, and by employing the COMPLEMENT DATA program (which permits inversion of the data values) installed in the ACAS 570 software system, we were able to obtain a complemental image which replicated the real immunohisto-morphology. Approximately 30–35 cells from three different areas in the same section were selected by DEFINE CELL and MARK AREA programs, and quantitative image analysis was performed in terms of cell integrated value, area, perimeter, and shape factor indicated in histogram form. The combined utilization of IGSS with computer-aided image analysis was demonstrated to offer a crucial advantage for the quantitative assessment of immunostained sections. 相似文献
13.
Noriko Kato Sachiko Karaki Kouichi Kashiwase Claudia Müller Tatsuya Akaza Takeo Juji Kyoichi Kano Masafumi Takiguchi 《Immunogenetics》1993,37(3):212-216
Serological studies have suggested the presence of a new HLA-B39 subtype (B39.2) in the Japanese population. To identify the new HLA-B39 subtype and compare it with an other HLA-B39 subtype (B39.1), the genes encoding HLA-B39.1 (B
*
39013) and B39.2 (B
*
3902) have been cloned from Japanese. We have sequenced these genes and completed the sequence of HLA-B39.1 (B
*39011
) gene from a Caucasian that was partially sequenced. Comparison of the sequence data revealed that B
*
3902 and B
*
39013 differ by three nucleotide substitutions which result in a two amino acids change at residues 63 and 67, while one silent substitution at codon 312 is found between B
*
39011 and B
*
39013. These results suggest that B
*
3902 has evolved from B
*
39013 rather than B
*
39011.The nucleotide sequence data reported in this paper have been submitted to the GenBank nucleotide sequence database and have been assigned the accession numbers M94051 (HLA-B*39013), M94052 (HLA-B*39011), and M94053 (HLA-B*3902). 相似文献
14.
Noriko Katayama Emi Iwata Hiroaki Sakurai Tomofusa Tsuchiya Masaaki Tsuda 《Journal of neurochemistry》1993,60(3):902-907
Abstract: Administration of carbachol, noradrenaline, and bradykinin induced Egr-1 mRNA expression within 1 h in mouse neuroblastoma × rat gliomahybrid NG108–15 cells. With specific receptor antagonists, the Egr-1 inductions by carbachol and noradrenaline were shown to be mediated via cholinergic muscarinic and α2 -adrenergic receptors, respectively. At their saturation levels for Egr-1 induction, the two agonists had additive effects when added together, but no prolongation of the effect on Egr-1 induction was observed. Addition of carbachol or noradrenaline 6 h after primary stimulation with carbachol or noradrenaline did not result in secondary Egr-1 induction, probably because of receptor desensitization. On the other hand, bradykinin consistently had an additive effect on Egr-1 induction, irrespective of the time of its addition, suggesting that the signal pathways for Egr-1 induction by carbachol or noradrenaline and by bradykinin are different. Treatment of cells with pertussis toxin or cholera toxin strongly inhibited Egr-1 induction by carbachol or noradrenaline but only partially inhibited the induction by bradykinin. Thus, the signals transduced in NG108–15 cells by different neurotransmitter receptors appear to have different effects on Egr-1 induction, depending on the times of stimulation and the combinations of receptors stimulated. 相似文献
15.
Noriko Takahashi Katsuyoshi Masuda Kenji Hiraki Kazuo Yoshihara Hung-Hsiang Huang Kay-Hooi Khoo Koichi Kato 《European journal of biochemistry》2003,270(12):2627-2632
To determine the glycoforms of squid rhodopsin, N-glycans were released by glycoamidase A digestion, reductively aminated with 2-aminopyridine, and then subjected to 2D HPLC analysis [Takahashi, N., Nakagawa, H., Fujikawa, K., Kawamura, Y. & Tomiya, N. (1995) Anal. Biochem.226, 139-146]. The major glycans of squid rhodopsin were shown to possess the alpha1-3 and alpha1-6 difucosylated innermost GlcNAc residue found in glycoproteins produced by insects and helminths. By combined use of 2D HPLC, electrospray ionization-mass spectrometry and permethylation and gas chromatography-electron ionization mass spectrometry analyses, it was revealed that most (85%) of the N-glycans exhibit the novel structure Manalpha1-6(Manalpha1-3)Manbeta1-4GlcNAcbeta1-4(Galbeta1-4Fucalpha1-6)(Fucalpha1-3)GlcNAc. 相似文献
16.
The Expression of Two Splice Variants of Metabotropic Glutamate Receptor Subtype 5 in the Rat Brain and Neuronal Cells During Development 总被引:3,自引:0,他引:3
Reiko Minakami Ken-ichiro Iida Noriko Hirakawa Hiroyuki Sugiyama 《Journal of neurochemistry》1995,65(4):1536-1542
Abstract: We previously reported that a variant with extra amino acid residues exists in the metabotropic glutamate receptor subtype 5 (mGluR5). Either of the two isoforms, named mGluR5b and mGluR5a for the isoforms with and without the inserted sequence, respectively, generated Ca2+ -activated Cl− current when expressed in Xenopus oocytes. We herein report that these two isoforms are produced by the alternative splicing of the exon skipping type. When examined during the course of postnatal development, the major mGluR5 isotype mRNA was observed to switch from mGluR5a to mGluR5b in the rat hippocampus and the cerebral cortex. We also investigated two cell lines that could be differentiated into neuron-like cells in vitro. Whereas the mGluR5b mRNA was hardly detectable in either undifferentiated or differentiated NG108-15 cells, the relative amounts of the two variant mRNAs changed after the induction of differentiation in the P19 cells. An extracellular application of trans - d,l -1-amino-1,3-cyclopentanedicarboxylate on the neuron-like P19 cells induced intracellular Ca2+ mobilization, thus suggesting that the cells could express functional mGluR(s) coupled to phospholipase C and other components that could mediate the signal transduction pathway. This cell line may thus provide a model system for studying both mGluR5 expression and other mGluR-induced phenomena at the molecular level. 相似文献
17.
Rika Morishita Shinsuke Saga Noriko Kawamura †Yoshio Hashizume ‡Toshiaki Inagaki Kanefusa Kato Tomiko Asano 《Journal of neurochemistry》1997,68(2):820-827
Abstract: The localization of two forms of the γ subunit of G proteins, γ3 and γ12, was examined in the mammalian brain. Concentrations of these two γ subunits increased markedly, as did those of glial fibrillary acidic protein, during postnatal development in the rat cerebral cortex. In aged human brains, by contrast, the concentration of γ3 tended to decrease with age, whereas that of γ12 in the temporal cortex increased slightly. An immunohistochemical study of human brains revealed that γ3 was abundant in the neuropil, whereas γ12 was localized in glial cells. In the hippocampal formation of aged human brains, levels of γ12-positive cells, as well as levels of glial fibrillary acidic protein- and vimentin-positive astrocytes, increased, in particular in the CA1 subfield and the prosubiculum, in which there was a decrease in the number of pyramidal cells. The appearance of γ12-positive cells associated with the loss of pyramidal cells was also observed in the hippocampus of rats that had been treated with kainic acid. These results indicate that γ12 is strongly expressed in reactive astrocytes. In a study of cultured neural cells, we found that γ12 was predominant in glioma cells, such as C6 and GA-1 cells, in contrast with the specific localization of γ3 in PC12 pheochromocytoma cells, which are neuron-like cells. Taken together, the results indicate that γ3 and γ12 are selectively expressed in neuronal and glial cells, respectively, and that concentrations of γ3 and γ12 in the brain are related to the numbers and/or extent of maturation of these cells. 相似文献
18.
Shinji Hosoi Mitsuo Satoh Hiromasa Miyaji Tatsunari Nishi Tamio Mizukami Mamoru Hasegawa Seiga Itoh Tatsuya tamaoki 《Cytotechnology》1995,19(1):1-10
Pro-UKS1 was designed as a thrombin-resistant derivative of pro-urokinase (pro-UK) by introducing a glycosylation site using site-directed mutagenesis. An expression plasmid for pro-UKS1, pMo1UKS1SEd1-5, was constructed and introduced into Namalwa KJM-1 cells (Hosoiet al., 1988), and cells resistant to G418 and Methotrexate (MTX) were obtained. Amongst them, the highest pro-UKS1 producer (resistant to 500 nM of MTX), clone 41-8, was selected and further characterized. Clone 41-8 was cultured in serum-free ITPSGF medium (Hosoiet al., 1988). Under the conventional conditions, the concentration of pro-UKS1 reached 26 g ml–1. Addition of glucose and tri-iodothyronine (T3) improved productivity, and the maximal productivity of pro-UKS1 was 67 g ml–1 day–1. In this conditioned medium, content of pro-UKS1 was above 80% of total proteins.Abbreviations BSA
bovine serum albumin
- dhfr
dihydrofolate reductase
- HEPES
4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid
- kb
kilobase pairs
- kDa
kilodaltons
- MTX
Methotrexate
- PBS
phosphate buffered saline
- pro-UK
pro-urokinase
- SDS-PAGE
sodium dodecyl sulfate-polyacrylamide gel electrophoresis
- T3
tri-iodothyronine
- Tween-PBS
phosphate buffered saline containing 0.05% Tween 80 相似文献
19.
A sulfite-dependent ATPase [EC 3.6.1.3
[EC]
] of Thiobacillus thiooxidanswas activated and solubilized by treatment with trypsin [EC3.4.4.4
[EC]
], and purified 84-fold with a 32% recovery. It requiredboth Mg2+ and SO32 for full activity, and its optimumpH was found at 7.58.0. Mn2+, Co2+, and Ca2+ could partiallysubstitute for Mg2+, while SeO32 and CrO42 couldpartially substitute for SO32. The enzyme hydrolyzed ATP and deoxy-ATP most rapidly and otherphosphate esters were poorer substrates. The apparent Km valuefor ATP was 0.33 mM. The enzyme activity was strongly inhibitedby 0.2 mM NaN3 and 10 mM NaF. (Received July 27, 1977; ) 相似文献
20.
Noriko Murakami Therese M. Quattrociocchi Shaun P. Healy Virinder K. Moudgil 《Archives of biochemistry and biophysics》1982,214(1):326-334
Effects of sodium tungstate on the nuclear uptake of rat liver cytosolic glucocorticoidreceptor complex were examined at pH 7. The nuclear uptake of heat-activated [3H]triamcinolone acetonide-receptor complex was blocked completely in the presence of 1 mm tungstate. A preincubation of nuclear preparation with tungstate (>0.1 mm) blocked the subsequent uptake of [3H]triamcinolone acetonide-receptor complex. When the tungstate-treated nuclear preparation was washed with 0.3 M KCl, its [3H]triamcinolone acetonide-receptor complex binding capacity recovered to 50% of that of control samples with no tungstate treatment. A preincubation of chromatin with tungstate yielded similar results. The nuclear-bound [3H]triamcinolone acetonide-receptor complex, formed either by an in vivo administration of [3H]triamcinolone acetonide or by an in vitro incubation of glucocorticoid-receptor complex with isolated nuclei, was extracted by tungstate in a concentration-dependent manner. The majority of nuclear-bound [3H]triamcinolone acetonide could be extracted with 0.1 and 1 mm tungstate from in vitro- and in vivo-labeled nuclei, respectively. The tungstate-extracted steroid-receptor complexes sedimented in 4–5 S and 3.3–3.5 S region in 10 mm KCl- and 0.3 mm KCl-containing sucrose gradients, respectively. Tungstate treatment caused an irreversible loss of the nuclear binding capacity of [3H]triamcinolone acetonide-receptor complex which could not be recovered after dialysis. These studies indicate that tungstate affects both glucocorticoidreceptor complex and certain nuclear or chromatin proteins. 相似文献