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91.
Ariyuki Kagaya Yosuke Uchitomi Akira Kugaya Minoru Takebayashi Ikuo Nagaoka Mitsutaro Muraoka Norio Yokota Shigeto Yamawaki 《Journal of neurochemistry》1996,66(4):1483-1488
Abstract: We investigated the rapid and slow effects of NaF on intracellular signaling systems such as Ca2+ homeostasis and cyclic GMP (cGMP) generation in rat glioma C6 cells, using the Ca2+ -sensitive dye fura-2 and cGMP enzyme immunoassay. We found that the following: (a) NaF enhanced cGMP generation in a concentration-dependent manner. This enhancement was abolished by pretreatment with 100 µ M BAPTA tetraacetoxymethyl ester or in the presence of W-7 in a concentration-dependent manner. N G -Monomethyl- l -arginine (NMMA), a competitive inhibitor of nitric oxide synthase (NOS), also inhibited the NaF-induced generation of cGMP. These results suggest that NaF-induced cGMP generation occurs via a calcium/calmodulin- and NOS-dependent pathway. (b) The basal intracellular Ca2+ concentration ([Ca2+ ]i ) was transiently greater at 1 and 3 h after pretreatment with NaF. W-7 and W-13 antagonized the increase in [Ca2+ ]i , whereas NMMA had little effect. This suggests that the NaF-induced change in basal [Ca2+ ]i was mediated by a calmodulin-dependent pathway but was independent of a NOS-sensitive pathway. (c) The serotonin (5-HT)-induced intracellular mobilization of Ca2+ was reduced by pretreating the cells with NaF. The reduction in Ca2+ mobilization was antagonized by genistein, a tyrosine kinase inhibitor. W-7, W-5, and H-8 had no effect. Results suggest that NaF differentially regulates the cGMP generation, basal [Ca2+ ]i , and 5-HT2A receptor function in C6 glioma cells. 相似文献
92.
Akira Nomura Yuko Yamazaki Takashi Tsuji Yukishige Kawasaki Shoji Tanaka 《Experimental cell research》1996,227(2):256
For all biological particles such as cells or cellular organelles, there are three-dimensional coordinates representing the centroid or center of gravity. These coordinates and other numerical parameters such as volume, fluorescence intensity, surface area, and shape are referred to in this paper as geometric properties, which may provide critical information for the clarification ofin situmechanisms of molecular and cellular functions in living organisms. We have established a method for the elucidation of these properties, designated the three-dimensional labeling program (3DLP). Algorithms of 3DLP are so simple that this method can be carried out through the use of software combinations in image analysis on a personal computer. To evaluate 3DLP, it was applied to a 32-cell-stage sea urchin embryo, double stained with FITC for cellular protein of blastomeres and propidium iodide for nuclear DNA. A stack of optical serial section images was obtained by confocal laser scanning microscopy. The method was found effective for determining geometric properties and should prove applicable to the study of many different kinds of biological particles in three-dimensional space. 相似文献
93.
Prosaposin Facilitates Sciatic Nerve Regeneration In Vivo 总被引:3,自引:0,他引:3
Yasunori Kotani Seiji Matsuda Masahiro Sakanaka Keiji Kondoh Shu-ichi Ueno Akira Sano 《Journal of neurochemistry》1996,66(5):2019-2025
Abstract: Prosaposin, a multifunctional protein, is the precursor of saposins, which activate sphingolipid hydrolases. In addition to acting as a precursor for saposins, prosaposin has been shown to rescue hippocampal CA1 neurons from lethal ischemic damage in vivo and to promote neurite extension of neuroblastoma cells in vitro. Here we show that prosaposin, when added to a collagen-filled nerve guide after sciatic nerve transection in guinea pigs, increased dramatically the number of regenerating nerve fibers within the guide. To identify the target neurons of prosaposin during peripheral nerve regeneration, we determined the degree of atrophy and chromatolysis of neurons in the spinal anterior horn and dorsal root ganglia on the prosaposin-treated and untreated side. The effect of prosaposin on large spinal neurons and small neurons of the dorsal root ganglion was more conspicuous. Subsequent immunohistochemistry demonstrated that the atrophy of cholinergic large neurons in the anterior horn is prevented to significant extent by prosaposin treatment. These findings suggest that prosaposin promotes peripheral nerve regeneration by acting on α-motor neurons in the anterior horn and on small sensory neurons in the dorsal root ganglion. The present study raises the possibility of using prosaposin as a tool for the treatment of peripheral nerve injuries. 相似文献
94.
Junko Komura Ikumi Tamai Mizuho Senmaru Tetsuya Terasaki Yoshimichi Sai Akira Tsuji 《Journal of neurochemistry》1996,67(1):330-335
Abstract: The characteristics of β-alanine transport at the blood-brain barrier were studied by using primary cultured bovine brain capillary endothelial cells. Kinetic analysis of the β-[3 H]alanine transport indicated that the transporter for β-alanine functions with Kt of 25.3 ± 2.5 µ M and J max of 6.90 ± 0.48 nmol/30 min/mg of protein in the brain capillary endothelial cells. β-[3 H]Alanine uptake is mediated by an active transporter, because metabolic inhibitors (2,4-dinitrophenol and NaN3 ) and low temperature reduced the uptake significantly. Furthermore, the uptake of β-[3 H]alanine required Na+ and Cl− in the external medium. Stoichiometric analysis of the transport demonstrated that two sodium ions and one chloride ion are associated with one β-alanine molecule. The Na+ and Cl− -dependent uptake of β-[3 H]alanine was stimulated by a valinomycin-induced inside-negative K+ -diffusion potential. β-Amino acids (β-alanine, taurine, and hypotaurine) inhibited strongly the uptake of β-[3 H]alanine, whereas α- and γ-amino acids had little or no inhibitory effect. In ATP-depleted cells, the uptake of β-[3 H]alanine was stimulated by preloading of β-alanine or taurine but not l -leucine. These results show that β-alanine is taken up by brain capillary endothelial cells, via the secondary active transport mechanism that is common to β-amino acids. 相似文献
95.
Makiko Kawagishi-Kobayashi Masayuki Yamamoto Akira Ishihama 《Molecular & general genetics : MGG》1996,250(1):1-6
Local sequence similarity exists between the subunit 2 of eukaryotic RNA polymerases II and the barnase-type bacterial RNases. The RNase-like domain from the Rpb2 ofSchizosaccharomyces pombe was expressed inEscherichia coli as a GST fusion protein and examined for its RNase activity. When the GST fusion protein was incubated in vitro with32P-labeled RNA, the RNA degradation activity was less than 0.1%, if any, of the level of synthetic barnase. In order to check the in vivo function of this region, we constructed two mutantrpb2 alleles,rpb2
E357A
andrpb2
H3a6L
, each carrying a single amino acid substitution at the site correponding to one of the three essential amino acid residues forming the catalytic site in barnase (mutation of barnase at the corresponding sites results in complete loss of RNase activity) and five other mutantrpb2 alleles, each carrying a single mutation at various positions within the RNase-like domain but outside the putative catalytic site for RNase activity. When these mutantrpb2 alleles were expressed in anrpb2-disruptedS. pombe strain, all the mutants grew as well as the wild-type parent and did not show any clear defective phenotypes. These results suggest either that the RNase-like domain in Rpb2 does not function as an RNase in vivo or that the RNase activity of this domain, if present at all, is not essential for cell growth. 相似文献
96.
Effects of interleukin-12 on in vitro culture with interleukin-2 of regional lymph node lymphocytes from lung cancer patients 总被引:2,自引:0,他引:2
T. Hanagiri Mitsuhiro Takenoyama Takashi Yoshimatsu Chikashi Hirashima Ichiro Yoshino Kozo Nakanishi Akira Nagashima Kikuo Nomoto Kosei Yasumoto 《Cancer immunology, immunotherapy : CII》1996,43(2):87-93
In the present study, we carried out a functional analysis of regional lymph node lymphocytes (RLNL) from patients with lung
cancer after in vitro activation by interleukin-2 (IL-2) and interleukin-12 (IL-12). IL-12 (100 U/ml) enhanced both the proliferation
and cytotoxic activity of RLNL in a culture with low doses of IL-2 (5 – 10 JRU/ml). After comparing an RLNL culture with a
low dose of IL-2 alone, a higher proportion of CD8+ cells and CD56+ cells and a lower proportion of CD4+ cells were found in the culture with both IL-12 and a low dose of IL-2. Such a combination of the cytokines effectively activated
RLNL in terms of the expression of IL-2 receptors. In the culture condition of IL-12 and a low dose of IL-2, a synergistic
effect was observed in the production of such cytokines as interferon γ, tumor necrosis factor α (TNFα), and TNFβ, as well
as in tumor cytotoxicity. However, the addition of IL-12 inhibited the cytotoxicity of RLNL in the culture with a high dose
of IL-2 (100 JRU/ml). This inhibition is considered to be partially due to the endogenous production of TNFα by lymphocytes,
because the neutralization of TNFα bioactivity partially restored the cytotoxic activities of RLNL. Furthermore, in the presence
of hydrocortisone, IL-12 synergistically enhanced the cytotoxic activity of RLNL cultured with a high dose of IL-2. These
results provide useful information about the improvement of adoptive immunotherapy against cancer using RLNL.
Received: 2 February 1996 / Accepted: 30 July 1996 相似文献
97.
Norio Ohashi Yoko Koyama Hiroshi Urakami Masahiro Fukuhara Akira Tamura Fumihiko Kawamori Seigo Yamamoto Shiro Kasuya Kentaro Yoshimura 《Microbiology and immunology》1996,40(9):627-638
A total of 40 strains of Orientia tsutsugamushi (34 isolates from patients and trombiculid mites in Japan, and 6 prototype strains of antigenic variants) were examined for classification based on the reactivities with type-specific monoclonal antibodies in indirect immunofluorescence tests, and on the restriction fragment length polymorphism of a polymerase chain reaction (PCR)-amplified 56-kilodalton type-specific antigenic protein gene. By these methods, several antigenic and genotypic variants were found among the strains, and these variants were classified into types and further into subtypes. These results suggest that there are many variants in O. tsutsugamushi, and the methods used here seem to be useful for the systematic classification of the numerous variants. A strain which may be a new type distinguishable from those identified previously was also found in this study. Furthermore, variety in the degree of pathogenicity in mice related to type and/or subtype classification were observed. 相似文献
98.
Akira Yamada Masaaki Murakami Kenichi Ijima Hideo Yagita Ko Okumura Sakuzo Komatsu Toshimitsu Uede 《Microbiology and immunology》1996,40(7):513-518
The immunosuppressant FK506 prolongs allograft survival. However, at therapeutic doses it has significant side effects. A fusion protein consisting of the extracellular portion of CTLA4 and the Fc portion of human IgG (CTLA4IgG) also prolongs allograft survival, but large doses of CTLA4IgG are required for the induction of cardiac allograft acceptance. Therefore, we constructed a pentameric form of a new CTLA4 fusion protein, CTLA4IgM. We tested whether low doses of CTLA4IgG or CTLA4IgM in combination with subtherapeutic doses of FK506 can prolong allograft survival in a synergistic fashion. C57BL/6 (H-2b) neonatal hearts were transplanted to CBA/J (H-2k) mice in a heterotopic, nonvascularized cardiac allograft model. The findings demonstrate that a combination of low doses of FK506 plus a pentameric form of CTLA4Ig, CTLA4IgM, leads to significant graft survival, while a combination of FK506 plus CTLA4IgG does not. 相似文献
99.
Takako S. Kaneko Mamiko Sato Masako Osumi Makoto Muroi Akira Takatsuki 《Plant cell reports》1996,15(6):409-413
Summary The effects of brefeldin A (BFA) on the secretion of acid phosphatase (APase) by tobacco protoplasts were investigated. Secretion of APase was inhibited by BFA in a dose-dependent manner, with a concomitant intracellular accumulation of the enzyme. The secreted APase was composed of two isoforms. BFA (10/ g/ml) inhibited the secretion of one of the isoforms without inhibiting that of the other, and this phenomenon explains the partial inhibition of APase secretion as a whole. The inhibition of APase secretion was accompanied by changes in the morphology of the Golgi apparatus and also by an increment in massdensity of cells.Abbreviations APase
acid phosphatase
- BFA
brefeldin A
- CHX
cycloheximide
- PAGE
polyacrylamide gel electrophoresis 相似文献
100.