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E Wada

Publications and source records attributed to E Wada.

At least 73 records · Page 4Linked to original sources
PubMed1993

Mechanisms of suppression of X-ray-induced transformation of 10T1/2 cells by lipopolysaccharide.

X-ray induced neoplastic transformation of C3H/10T1/2 cells was suppressed equally by Lipid A and LPS. We examined the effects of several modifiers of arachidonic acid (AA) metabolism in our investigation on the suppression mechanism of LPS. Dexamethasone (DM), an inhibitor of phospholipase A2 (PLA2), abolished the effect of LPS, whereas bromophenacyl bromide (BPB), another inhibitor of PLA2 did not. AA and aspirin neither changed the transformation frequency nor blocked the effect of LPS. TPA, 12-O-tetradecanoyl-phorbol-13-acetate, slightly enhanced radiation transformation, which was diminished by LPS. We also examined the effects of these modifiers on the release of radioactivity from 3H-AA-labeled 10T1/2 cells. No definite correlation was found between the change of AA metabolism and suppression of the transformation frequency by LPS. LPS, however, enhanced dose-dependent myristoylation of the 22 and 67 kDa proteins of 10T1/2 cells.

Animals↗
PubMed1993

[Cross-allergenicity between rice and buckwheat antigens and immediate hypersensitive reactions induced by buckwheat ingestion].

Immediate hypersensitive reactions (IHR) induced by buckwheat ingestion are considered to be IgE-mediated. However we found 28 subjects without IHR to buckwheat ingestion out of 46 subjects who had positive RAST values for both buckwheat and rice antigens. The IHR-positive group showed significantly higher RAST values for buckwheat antigens (p < 0.01) but lower RAST values for rice antigens (p < 0.01) than did the IHR-negative group. RAST values for buckwheat and rice were significantly correlated with each other (p < 0.01) in the IHR-negative group, but not in the IHR-positive group. An effective dose-dependent inhibition was obtained in a RAST inhibition assay between homologous combinations of inhibitor and disc antigens such as rice and rice or buckwheat and buckwheat. The IHR-positive group showed no significant RAST inhibition between heterogeneous combinations of rice and buckwheat antigens. In contrast, the IHR-negative group showed a significant decrease in IgE binding even in the RAST inhibition assay between heterogenous combinations. These results led to the conclusion that there is cross-reactivity with IgE antibodies between buckwheat and rice and that IgE antibodies from IHR-negative subjects might recognize the epitopes on buckwheat antigens which cross react with rice antigens, whereas IgE antibodies from IHR-positive subjects might bind to buckwheat-specific epitopes.

Adolescent↗
PubMed1993

[Allergen analysis of buckwheat by the immunoblotting method].

Inhalation or ingestion of very small amounts of buckwheat allergen can initiate severe symptoms including wheezing, rhinorrhea, urticaria, vomiting and anaphylactic shock in patients with hypersensitivity against buckwheat. However, few studies of the immunological properties of buckwheat allergen have been made. The aim of the present investigation was to analyze the allergen in buckwheat antigen using the immunoblotting method, radioallergosorbent test (RAST) and RAST inhibition assay. Buckwheat 24 kilodalton protein (BW24KD) was shown by immunoblotting analysis to be the most frequently recognized allergenic component, binding to IgE antibodies from 100% of the patients' sera. There was a significant positive correlation (p < 0.001) between % bindings for BW24KD and buckwheat. In the RAST inhibition assay using patients' sera which showed positive IgE antibodies to buckwheat and BW24KD, there was effective inhibition in a dose-dependent manner between inhibition in a dose-dependent manner between BW24KD and buckwheat. These data indicated that BW24KD was a major allergen in buckwheat antigens. The experiments of SDS-PAGE using digestion buffer with or without 2-mercaptoethanol suggested that BW24KD was a component of heterodimer and the paired components had different molecular weights.

Adolescent↗
PubMed1992

Neuronal cdc2-like kinase: a cdc2-related protein kinase with predominantly neuronal expression.

Recent studies have shown that there exists a family of protein kinases structurally and functionally related to the yeast cell cycle regulatory kinase cdc2 [Meyerson, M., Faha, B., Su, L.-K., Harlow, E. & Tsai, L.-H. (1991) Cold Spring Harbor Symp. Quant. Biol. 56, 177-186 and Meyerson, M., Enders, G. H., Wu, C.-L., Su, L.-K., Gorka, C., Nelson, C., Harlow, E. & Tsai, L.-H. (1992) EMBO J. 11, 2909-2917]. Two members of cdc2 family, p34cdc2 (also named cdk1) and cdk2, have been identified in mammalian cells. cdk1 kinase regulates the progression from G2 to M phase, and cdk2 kinase has been proposed to regulate the progression from G1 to S phase. In this work, we have cloned and structurally characterized a third member of the cdc2 kinase family with 58% amino acid sequence identity to mouse cdk1 and 61% identity to human cdk2. We call this kinase neuronal cdc2-like kinase (nclk) because, in contrast to either cdk1 or cdk2, nclk is expressed at high levels in terminally differentiated neurons no longer in the cell cycle. Previous studies have shown [Hisanaga, S., Kusubata, M., Okumura, E. & Kishimoto, T. (1991) J. Biol. Chem. 266, 21798-21803 and Guan, R. J., Hall, F. L. & Cohlberg, J. A. (1992) J. Neurochem. 58, 1365-1371] that cdk1 kinase, but not other structurally defined protein kinases, could phosphorylate the repeated Lys-Ser-Pro (KSP) motifs found in mammalian high and middle molecular mass neurofilament subunits in vitro, but the precise molecular nature of the endogenous neuronal KSP kinase has remained undefined. The structural similarity of nclk to cdk1 kinase and its high level of expression in terminally differentiated neurons suggest that nclk may play a role in the phosphorylation of the neurofilament KSP repeats in vivo, a function distinct from cell cycle regulation.

Amino Acid Sequence↗
PubMed1992

Activation of neuromedin B-preferring bombesin receptors on rat glioblastoma C-6 cells increases cellular Ca2+ and phosphoinositides.

Recent cloning studies confirm the presence of two subtypes of bombesin (Bn) receptors. In contrast to the gastrin-releasing peptide (GRP)-preferring subtype, which has been widely studied, nothing is known about the cellular mechanisms of the neuromedin B (NMB)-preferring subtype, which occurs widely in the central nervous system and gastrointestinal tissues, partially because of the lack of a cell line with functional receptors. In the present study we have investigated Bn receptors on the rat glioblastoma cell line C-6, reported to contain mRNA of the NMB receptor subtype. Binding of 125I-[D-Tyr0]NMB to these cells was time- and temperature-dependent, saturable, reversible, and only inhibited by Bn receptor agonists or antagonists. For Bn receptor agonists the relative potencies were: NMB (1.7 nM) approximately equal to litorin (3 nM) greater than ranatensin (8 nM) greater than Bn (19 nM) greater than neuromedin C (NMC) (210 nM) greater than GRP (500 nM). These relative affinities were almost identical to those for the NMB receptor subtype on rat oesophageal tissue and for Balb 3T3 cells stably transfected with the NMB receptor subtype. These potencies differed from those for the GRP receptor subtype on rat pancreatic acini [Bn approximately equal to litorin (4 nM) greater than ranatensin, NMC, GRP (15-20 nM) much greater than NMB (351 nM)]. The relative potencies of four different classes of Bn receptor antagonists were compared. Results from C-6 tumour cells agreed closely with those for binding to the NMB receptor subtype on rat oesophageal tissue and in Balb 3T3 cells stably transfected with this receptor, and differed markedly from those for binding to the GRP receptor subtype on rat pancreatic acini. Four Bn receptor antagonists had a higher affinity for the GRP subtype ([D-Phe6]Bn-(6-13)ethyl ester (500 x), [D-Phe6][psi 13-14,Cpa14]Bn- (6-14) (70 x) (where psi 13-14 refers to the replacement of the -CONH- peptide bond between Leu13 and Met14 by -CH2NH2) [psi 13-14,Leu14]Bn, [D-Phe6]Bn-(6-13) propylamide (30 x)] and two had a higher affinity for the NMB subtype on C-6 cells and transfected cells ([D-Pro4,D-Trp7,9,10] substance P-(4-11) (9 x) and [Tyr4,D-Phe12]Bn (18 x)]. In C-6 tumour cells, Bn receptor agonists caused an increase in cytosolic Ca2+ and the generation of inositol phosphates. For both responses, NMB was more than 50-fold more potent than GRP. Neither NMB nor GRP increased cyclic AMP. These results demonstrate that the rat glioblastoma cell line C-6 possesses functional NMB-preferring Bn receptors, and agonist occupation activates phospholipase C, thus increasing cytosolic Ca2+ and inositol phosphate formation. Because the interaction of Bn-related peptides with C-6 cell receptors is identical with that reported in other tissues containing the mRNA for the NMB subtype, this cell line should prove useful in exploring further the cellular basis of action of the peptides that interact with this receptor in the central nervous system and various other tissues.

3T3 Cells↗
PubMed1992

Experimental spondylosis in the rabbit spine. Overuse could accelerate the spondylosis.

It has been suggested that overuse can accelerate the progression of cervical spondylosis. This assumption was derived from a study examining premature onset of cervical spondylosis in patients with athetoid cerebral palsy. To clarify the relationship between overuse and structural abnormalities of the spine, repetitive extension-flexion movement was located onto young rabbit spines through electric stimulation of the trapezius muscle. Repetitive loads of 200,000 cycles caused more severe delamination of the anulus fibrosus than control at the lower cervical spine. In addition, it was associated with early osteophyte formation at the same disc level. No severe degeneration of the nucleus pulposus, however, occurred through repetitive loading. It was found that repetitive movement could accelerate the progression of structural abnormalities, such as cervical spondylosis. This result suggests that overuse is an important factor in the pathogenesis of spondylosis.

Animals↗
PubMed1992

Distribution of type I interleukin-1 receptor messenger RNA in testis: an in situ histochemical study in the mouse.

The cytokine interleukin-1 (IL-1) has been reported to inhibit the hypothalamic-pituitary-gonadal axis, both through actions in brain and at the gonadal level. Recently, high affinity binding sites for 125I-recombinant human IL-1 alpha have been identified in the mouse testis with characteristics similar to those of type I IL-1 receptors on T lymphocytes and fibroblasts. The present study employed in situ hybridization histochemistry with 35S-labeled antisense cRNA probes derived from a murine type I IL-1 receptor cDNA to identify type I IL-1 receptor mRNA in the mouse testis. An intense signal was observed over interstitial cells, and over the cytoplasm of the epithelium of epididymal ducts, most prominently in the head region. The signal over seminiferous tubules, and over sperm cells within tubules and epididymal ducts, was comparable to background. This distribution of type I IL-1 receptor mRNA was similar to that recently reported for 125(I)I-IL-1-alpha binding sites, and supports evidence implicating IL-1 as a direct regulator of gonadal function.

Animals↗
PubMed1992

Molecular genetic analysis of two distinct receptors for mammalian bombesin-like peptides.

The mammalian bombesin-like peptides are known to be growth factors for certain cells with high-affinity bombesin receptors and have been implicated as autocrine growth factors influencing the pathogenesis and progression of a subset of human small-cell lung carcinomas. Thus, antagonists that interfere with bombesin receptor-ligand interaction might prove to be of value in treatment of gastrin-releasing peptide (GRP)-responsive tumors. A precise definition of the structure and properties of the bombesin receptors found on human lung cancer cells would provide important information for the design and rational application of such antagonists. Recently, we isolated cDNA clones encoding two distinct receptors for the mammalian bombesin-like peptides, GRP, and neuromedian B (NMB). The two receptors show 56% amino acid identity, encode seven putative transmembrane domains, and are members of the G-protein-coupled receptor superfamily. Ligand-binding studies show that while both receptors can be activated by either GRP or NMB, one receptor has a higher affinity for GRP than for NMB (GRP-R), while the other has a higher affinity for NMB than for GRP (NMB-R). A different spectrum of antagonists is needed to block responses from the two different receptors. These studies indicate that it will be critical in future studies to define which bombesin receptor subtypes are present on a given tumor to optimize the potential therapeutic benefit of antagonists in blocking growth.

Amino Acid Sequence↗
PubMed1992

Neuromedin B receptors retain functional expression when transfected into BALB 3T3 fibroblasts: analysis of binding, kinetics, stoichiometry, modulation by guanine nucleotide-binding proteins, and signal transduction and comparison with natively expressed receptors.

The receptor that interacts with the mammalian bombesin-related peptide neuromedin B (NMB) is ubiquitous in the gastrointestinal tract and central nervous system. However, little is known regarding its cellular mechanisms of action. This receptor has been recently cloned, sequenced, and stably transfected into BALB 3T3 fibroblasts, permitting detailed study of the pharmacology and coupled biological activities of this receptor. In the present study, we compare the ability of transfected receptors to alter cell function with that of receptors natively expressed in small numbers by the rat glioblastoma cell line C6. NMB inhibited binding of 125I-[D-Tyro]NMB with high affinity in transfected cells (Ki = 3.08 +/- 0.14 nM) and in C6 cells (Ki = 1.90 +/- 1.10 nM), whereas the bombesin-related agonists gastrin-releasing peptide (GRP) and [D-Phe6, D-Ala11, Leu14]bombesin(6-16) (GRP analogue) had 100- and 300-fold lower affinities, respectively, for NMB receptors in either cell type. For both cell systems, maximal binding was observed between 5 and 15 min at 22 degrees. Both cell types internalized NMB at similar rates, with > 70% of bound ligand being internalized by 60 min at 22 degrees. The nonhydrolyzable guanosine analogue guanosine 5'-(beta,gamma-imido)triphosphate was equipotent in causing a decrease in binding of 125I-[D-Tyro]NMB due to decreased receptor affinity in both cell types, without a change in receptor number, demonstrating that the NMB receptor remained coupled to a guanine nucleotide-binding protein in both native and transfected cells. In both cell systems, NMB increased inositol monophosphate, inositol bisphosphate, and inositol trisphosphate in a time-dependent fashion. Inositol phosphates were increased in a dose-dependent fashion, with similar half-maximal values being obtained for NMB in both cell types (transfected, 1.01 +/- 0.09 nM; C6, 2.09 +/- 0.15 nM) and for the GRP analogue (transfected, 1855 +/- 140 nM; C6, 2129 +/- 250 nM). NMB mobilized intracellular Ca2+ in both cell systems, and the dose-response curves were superimposible (EC50 for transfected, 0.10 +/- 0.08 nM; C6, 0.11 +/- 0.02 nM). These data demonstrate that activation of the receptor for NMB stimulates phospholipase C and increases intracellular Ca2+. These results also demonstrate that transfected and native NMB receptors behave similarly, suggesting that the transfected cell line will be useful in future studies investigating ligand-receptor interactions, as well as in molecular biological studies of the structure-function relationship of the receptor.

3T3 Cells↗
PubMed1992

In situ histochemical localization of type I interleukin-1 receptor messenger RNA in the central nervous system, pituitary, and adrenal gland of the mouse.

The cytokine interleukin-1 (IL-1) has a number of biologic activities, including pronounced effects on the nervous and neuroendocrine systems. In this study, in situ histochemical techniques were used to investigate the distribution of cells expressing type I IL-1 receptor mRNA in the CNS, pituitary, and adrenal gland of the mouse. Hybridization of 35S-labeled antisense cRNA probes derived from a murine T-cell IL-1 receptor cDNA revealed a distinct regional distribution of the type I IL-1 receptor, both in brain and in the pituitary gland. In the brain, an intense signal was observed over the granule cell layer of the dentate gyrus, over the entire midline raphe system, over the choroid plexus, and over endothelial cells of postcapillary venules throughout the neuraxis. A weak to moderate signal was observed over the pyramidal cell layer of the hilus and CA3 region of the hippocampus, over the anterodorsal thalamic nucleus, over Purkinje cells of the cerebellar cortex, and in scattered clusters over the external-most layer of the median eminence. In the pituitary gland, a dense and homogeneously distributed signal was observed over the entire anterior lobe. No autoradiographic signal above background was observed over the posterior and intermediate lobes of the pituitary, or over the adrenal gland. This study therefore provides evidence for discrete receptor substrates subserving the central effects of IL-1, thus supporting the notion that IL-1 acts as a neurotransmitter/neuromodulator in brain. It also supports studies suggesting that IL-1-mediated activation of the hypothalamic-pituitary-adrenal axis occurs primarily at the level of the brain and/or pituitary gland.

Adrenal Glands↗
PubMed1992

Elemental distribution in frozen-hydrated mouse lenses with hereditary cataract.

Nakano mice (nct/nct) and its congenic strain (Balb/c-nct/nct) develop different types of cataract: the former develop intense nuclear cataract but the latter develop milder opacity at the cortical region of lenses more later in life. The present study was undertaken to compare the elemental compositions at the localized region of these lenses in order to examine the possible role of ions in the opacification of the hereditary cataracts. Eight lenses from Nakano mice (age: 39-40 days) and ten lenses from congenic mice (age: 73 days) were prepared for the X-ray analysis with a high energy ion microprobe at a resolution of 50 microns. The age of mice was two weeks after 50% cataract formation for the respective strain. The maps of the concentrations of S, Cl, K and Ca in the central planes of the frozen-hydrated lenses were obtained. In all of the congenic lenses analyzed, the normal elemental composition, low Cl and Ca, and high K, was retained only in the narrow anterial and equatorial peripheral region and the other region underwent a K loss with a gain of Cl and Ca. In the Nakano lenses, the altered elemental composition was localized in central and in posterial cortical regions. There was no distinct difference in the concentration levels of Cl, K and Ca in the central region of the lenses between the two strains. The level of S was lower in Nakano lenses. High calcium containing spots were detected in low K regions of some of the lenses examined.

Animals↗
PubMed1991

Distinguishing bombesin receptor subtypes using the oocyte assay.

Physiological responses to mammalian bombesin-like peptides were studied in Xenopus oocytes injected with mRNA isolated from Swiss 3T3 cells and rat esophagus in order to identify and characterize bombesin receptor subtypes. Both groups respond similarly to either gastrin releasing peptide or neuromedin B, but only the response to neuromedin B in oocytes expressing the esophagus mRNA is not blocked by a specific gastrin releasing peptide receptor antagonist, des-Met-[D-Phe6]Bn(6-13) ethyl ester. Complete desensitization of gastrin releasing peptide-evoked responses in oocytes expressing esophagus mRNA does not abolish neuromedin B-evoked responses. A single application of neuromedin B abolishes responses to subsequently applied gastrin releasing peptide in oocytes expressing esophagus, but not Swiss 3T3, mRNA. RNA blot hybridization studies using a Swiss 3T3 gastrin releasing peptide receptor cDNA probe show no detectable hybridization in esophagus mRNA samples. These data suggest that a gastrin releasing peptide receptor is expressed in the esophagus and that it is distinct from that expressed in Swiss 3T3 cells and may represent a third subtype of mammalian bombesin receptor.

Animals↗
PubMed1991

Biochemical evidence for conversion to milder form of hereditary mouse cataract by different genetic background.

Congenic hereditary cataract mice, BALB/c-nct/nct, were established by introducing the nct gene from Nakano into BALB/c mice. These mice developed a milder cortical form of cataract which developed sporadically and later in life than in Nakano mice. Combined use of BALB/c and BALB/c-nct/nct mice enables biochemical comparison of normal clear lenses, congenic clear lenses which are destined to be opacified some time later, and opacified lenses in the same genetic and aging statuses. We compared the age-related changes in water content and water-soluble and -insoluble fractions among these three types of lenses. Congenic clear lenses and opaque lenses were more similar to BALB/c normal clear lenses and Nakano opaque ones, respectively, in these parameters. These results suggest, in addition to formation of aggregated crystallins and their accumulation in water-insoluble fractions, that decreased protein synthesis, increased protein degradation and augmented leakage of crystallin might have a significant role in the nct-induced lens opacification.

Aging↗
PubMed1991

Two distinct receptor subtypes for mammalian bombesin-like peptides.

The mammalian bombesin-like peptides, gastrin-releasing peptide (GRP) and neuromedin B (NMB), are structurally related neuropeptides that elicit a wide spectrum of biological activities including regulation of smooth muscle contraction, stimulation of secretion, modulation of neural activity, and growth regulation. Earlier studies have shown that GRP and NMB are expressed in different regions of both the CNS and peripheral organs. Recent ligand-binding and molecular-cloning studies have revealed two pharmacologically distinct G-protein-coupled receptor subtypes for mammalian bombesin-like peptides that have different relative affinities for GRP, NMB and bombesin receptor antagonists. Similar to the peptide ligands, the two receptor subtypes are expressed in a distinct but overlapping set of CNS regions, some of which have been identified in functional studies as sites where bombesin peptides elicit defined biological responses. Delineation of these peptide ligands and receptor subtypes will be important in future studies that explore the molecular basis for the heterogeneous nature of the responses to bombesin observed in mammalian systems.

Amino Acid Sequence↗
PubMed1991

cDNA cloning, characterization, and brain region-specific expression of a neuromedin-B-preferring bombesin receptor.

Recent binding studies in the central nervous system and other tissues provide evidence that the mammalian bombesin-like peptides, gastrin-releasing peptide (GRP) and neuromedin-B (NMB), exert their numerous physiological effects through at least two different receptors. We describe the structure and expression of a cloned NMB-preferring bombesin receptor (NMB-R) with properties distinct from a GRP-preferring bombesin receptor (GRP-R) reported previously. In particular, the NMB-R shows higher affinity binding to NMB than to GRP in BALB 3T3 fibroblasts expressing the cloned NMB-R. The distinct regional distribution of NMB-R and GRP-R mRNA in the brain suggests that both bombesin receptor subtypes play independent roles in mediating many of the dramatic effects of bombesin-like peptides in the central nervous system.

Amino Acid Sequence↗
PubMed1991

The use of stable isotopes for food web analysis.

General aspects in isotope biogeochemistry was summarized with emphasis on delta 15N and delta 13C contents in plants and animals in natural ecosystems. In the estuary, the variation of isotope ratios were principally governed by the mixing of land-derived organic matter, marine phytoplankton, and seagrasses. A clear cut linear relationship between animal delta 15N and its trophic level was obtained in the Antarctic food chain system. Several current efforts to use the stable isotopes for food web analysis were demonstrated for some terrestrial and marine systems as well as human food web.

Animals↗