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Biomedical subjects

A Miyake

Publications and source records attributed to A Miyake.

At least 91 records · Page 5Linked to original sources
PubMed1996

Transforming growth factor-alpha regulates subpopulation of giant cells which secrete mouse placental lactogen-I (mPL-I) and/or mPL-II at midpregnancy.

The aim of this study was to determine whether transforming growth factor-alpha (TGF-alpha) regulates expression of mouse placental lactogen-I (mPL-I) and mPL-II at midpregnancy in vitro. Treatment of placental cells from day 9 of pregnancy with TGF-alpha resulted in stimulation of mPL-I secretion and inhibition of mPL-II secretion in a time- and dose-dependent manner without changing the amount of newly synthesized trichloroacetic acid precipitable proteins and cell viability. Moreover, TGF-alpha increased the intra-cellular mPL-I concentration and immuno-precipitable newly synthesized mPL-I concentration both in the medium and cells. TGF-alpha increased the number of cells containing only mPL-I and cells containing both mPL-I and mPL-II, but decreased that of the cells containing only mPL-II assessed by double immunocytochemistry. TGF-alpha increased the number of cells releasing only mPL-I but decreased the number of cells releasing only mPL-II, however TGF-alpha did not affect the number of cells releasing both mPL-I and mPL-II assessed by sequential reverse hemolytic plaque assay (RHPA). TGF-alpha decreased the expression of mPL-II mRNA, but did not change the expression of mPL-I mRNA. In situ hybridization for epidermal growth factor receptor (EGF-R) after RHPA for mPL-I indicated that giant cells releasing mPL-I express EGF-R mRNA. These findings suggest that TGF-alpha regulates the subpopulation of giant cells which produces and releases mPL-I and mPL-II and it results in an increase of mPL-I secretion and decrease of mPL-II secretion, and suggests possible post-transcriptional stimulation of mPL-I secretion and transcriptional inhibition of mPL-II secretion by TGF-alpha. Giant cells releasing mPL-I express EGF-R mRNA, suggesting direct regulation of giant cell differentiation by EGF and TGF-alpha at midpregnancy.

Animals↗
PubMed1996

A stress-sensitive chemokinergic neuronal pathway in the hypothalamo-pituitary system.

Recently we found that cytokine-induced neutrophil chemoattractant influenced anterior pituitary hormone release in vitro. These observations prompted us to investigate the possibility of the existence of cytokine-induced neutrophil chemoattractant in the hypothalamus. Immunohistochemistry showed that cytokine-induced neutrophil chemoattractant-like immunoreactivity existed in the paraventricular hypothalamic nucleus, the supraoptic nucleus, both the internal and the external layers of the median eminence and the posterior pituitary. Since the paraventricular hypothalamic nucleus plays a pivotal role in response to stressful stimuli, we examined the effect of a single episode of immobilization stress on cytokine-induced neutrophil chemoattractant messenger RNA expression in the paraventricular hypothalamic nucleus. Immobilization stress induced strong hybridization signals of cytokine-induced neutrophil chemoattractant messenger RNA in the parvocellular and magnocellular subdivision of the paraventricular hypothalamic nucleus within 15 min, and cytokine-induced neutrophil chemoattractant-like immunostaining intensity in the posterior pituitary started to increase around the periphery of the posterior lobe at 30 min after stress and extended to the whole lobe at 1 h after stress. The increase in the serum cytokine-induced neutrophil chemoattractant in response to stress showed a kinetically biphasic pattern. A first phase occurred within 15 min which may be due to an immediate release of stored cytokine-induced neutrophil chemoattractant in the neurohypophysis, since hypophysectomy completely blocked this phase. A second phase may reflect the release of newly synthesized cytokine-induced neutrophil chemoattractant in the paraventricular hypothalamic nucleus and/or peripheral cytokine-induced neutrophil chemoattractant, since hypophysectomy could not reduce this phase. These data suggest that cytokine-induced neutrophil chemoattractant in the paraventricular hypothalamic nucleus was immediately synthesized in response to stress, and then released into the peripheral blood via the hypothalamo-neurohypophysial system, revealing the presence of a stress-sensitive chemokinergic neuronal pathway in the hypothalamo-pituitary system.

Animals↗
PubMed1996

Changes in levels of mRNAs of transforming growth factor (TGF)-beta1, -beta2, -beta3, TGF-beta type II receptor and sulfated glycoprotein-2 during apoptosis of mouse uterine epithelium.

To examine the roles played by transforming growth factors (TGF)-beta1, -beta2, -beta3, and TGF-beta type II receptors in the induction of apoptosis in the mouse uterine epithelium after estrogen deprivation, we investigated the expression of their mRNAs and the mRNA of sulfated glycoprotein-2 (SGP-2). Pellets containing 100 microg estradiol-17beta (E2) were implanted into ovariectomized mice and removed four days later. Apoptotic indices (percentage of apoptotic cells) of both luminal and glandular epithelia increased after E2 pellets were removed, but administration of progesterone (P), 5alpha-dihydrotestosterone (DHT), or continued implantation of E2 pellets suppressed this increase. Levels of mRNAs of TGF-beta1, -beta2, and -beta3, and SGP-2 did not increase after estrogen deprivation. However, estrogen deprivation caused a gradual increase in the level of TGF-beta type II receptor mRNA, and its level increased about six-fold six days later. Moreover, E2, P, and DHT markedly decreased the level of TGF-beta type II receptor mRNA. In situ hybridization demonstrated that mRNAs of TGF-beta1, -beta2, -beta3 and TGF-beta type II receptor were localized to the epithelium. Exogenous administration of TGF-beta1 into the uterine stroma induced apoptosis in the epithelium, a finding that suggests that signals produced by TGF-betas can induce apoptosis. Therefore, the present results suggest that increased sensitivity of uterine epithelial cells to TGF-betas, as demonstrated by an increase in TGF-beta type II receptor mRNA, is involved in the induction of apoptosis after estrogen deprivation, although signals produced by TGF-betas do not appear sufficient to induce apoptosis.

Animals↗
PubMed1996

The separability of working memory resources for spatial thinking and language processing: an individual differences approach.

The current study demonstrates the separability of spatial and verbal working memory resources among college students. In Experiment 1, we developed a spatial span task that taxes both the processing and storage components of spatial working memory. This measure correlates with spatial ability (spatial visualization) measures, but not with verbal ability measures. In contrast, the reading span test, a common test of verbal working memory, correlates with verbal ability measures, but not with spatial ability measures. Experiment 2, which uses an interference paradigm to cross the processing and storage demands of span tasks, replicates this dissociation and further demonstrates that both the processing and storage components of working memory tasks are important for predicting performance on spatial thinking and language processing tasks.

Adult↗
PubMed1996

Involvement of Rabphilin-3A in cortical granule exocytosis in mouse eggs.

Rabphilin-3A is a putative target protein for Rab3A, a member of the small GTP-binding protein superfamily that has been suggested to play a role in regulated exocytosis in presynapses. In this study we determined the expression and the function of Rabphilin-3A in mouse eggs at fertilization. Rabphilin-3A mRNA and protein were detected by reverse transcriptase-PCR and immunoblot analysis, respectively, in metaphase II mouse eggs. Immunofluorescence analysis showed that Rabphilin-3A protein was distributed in the cortical region in eggs. Sperm induces cortical granule (CG) exocytosis via an increase in cytosolic Ca2+ at fertilization. We microinjected the NH2- or COOH-terminal fragment of recombinant Rabphilin-3A into metaphase II eggs. Neither treatments altered the sperm-induced cytosolic Ca2+ increase, but both inhibited CG exocytosis in a dose-dependent manner. The NH2-terminal fragment was more effective than the COOH-terminal fragment. Full-length Rabphilin-3A did not affect CG exocytosis, but it attenuated the inhibition of CG exocytosis by the NH2-terminal fragment. These results show that Rabphilin-3A is involved in Ca(2+)-dependent CG exocytosis at fertilization in mouse eggs.

Adaptor Proteins, Signal Transducing↗
PubMed1996

Reactive oxygen species block sperm-egg fusion via oxidation of sperm sulfhydryl proteins in mice.

The effects of reactive oxygen species (ROS) on mouse sperm-egg fusion were determined. Sperm were treated with superoxide and hydrogen peroxide generated by addition of xanthine oxidase (XO: 10-200 mlU/ml) to hypoxanthine (HX: 1 mM). While XO at concentrations higher than 100 mlU/ml decreased the motility and lipid peroxidation of sperm, XO at less than 50 mlU/ml had no such effect. However, 20-50 mlU/ml XO significantly suppressed sperm fusion with zona-free eggs. Two ROS scavengers, superoxide dismutase and catalase, attenuated the inhibition of sperm-egg fusion by HX-XO. The sulfhydryl (SH) reductant, dithiothreitol, also reversed the inhibition. The sperm SH-rich fusion-related proteins were highly sensitive to ROS. These results suggest that ROS at low concentrations may inhibit sperm-egg fusion via oxidation of the SH-proteins in the sperm membrane, without causing loss of motility.

Animals↗
PubMed1996

Dynamics of cortical granule exocytosis at fertilization in living mouse eggs.

Sperm-egg fusion induces an intracellular free calcium concentration ([Ca2+]i) increase and exocytosis of cortical granules (CGs). Recently we used an impermeable fluorescent membrane probe, 1-[4-(trimethylammonio)phenyl]-6-phenyl-1,3,5-hexatriene (TMA-DPH), to develop a method to evaluate the kinetics of exocytosis in single living cells. In this study we used digital imaging and confocal laser scanning microscopy to evaluate CG exocytosis in living mouse eggs with TMA-DPH. Time-related changes of CG exocytosis were estimated as the percent increase of TMA-DPH fluorescence. The increase of fluorescence in the egg started after sperm attachment, continued at an almost uniform rate, and ceased at 45-60 min. Whereas the [Ca2+]i increase at fertilization was transient or oscillatory, exocytosis was not always induced concomitantly with each [Ca2+]i peak. Next we used this method to determine some intracellular mediators of exocytosis in the egg. An intracellular calcium chelator, 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-acetoxymethyl ester, and a microfilament inhibitor, cytochalasin B, blocked sperm-induced exocytosis. A guanosine 5'-triphosphate-binding protein activator, AlF4-, induced exocytosis. These results suggest that [Ca2+]i, microfilament, and guanosine 5'-triphosphate-binding proteins may be involved in CG exocytosis. In conclusion, this method has significant advantages for studying exocytosis in living eggs.

Animals↗
PubMed1996

Endotoxin activates a chemokinergic neuronal pathway in the hypothalamo-pituitary system.

Cytokines play a critical role in the cascade of events that cause septic shok. One regulatory system suggested to be important in controlling inflammatory response is the neuroendocrine axis. One of the chemokines is cytokine induced neutrophil chemoattractant (CINC), which was first described as an immuno-modulator of peripheral tissue in inflammatory responses. To assess further the contribution of the chemokine to the central nervous system, we performed immunohistochemistry on rat brains and found strong CINC-like immunoreactivity in the posterior pituitary gland. Treatment with bacterial endotoxin (lipopolysaccharide; LPS) markedly enhanced CINC-like immunoreactivity in the posterior pituitary. Before the LPS challenge, signal for CINC mRNA was undetectable in the paraventricular hypothalamic nucleus (PVN). The LPS challenge induced strong hybridization signals of CINC mRNA in the parvocellular and magnocellular subdivision of the PVN within 15 minutes (min) and peaked at 30 min. The LPS challenge provoked no observable change in the supraoptic nucleus. These studies demonstrate the presence of an endotoxin-sensitive chemokinergic neuronal pathway in the hypothalamo-neurohypophysial system and this newly-described pathway will provide a novel information to understand another possible neuralimmune mechanism.

Animals↗
PubMed1996

Synthetic studies on condensed-azole derivatives. IV. Synthesis and anti-asthmatic activities of omega-sulfamoylalkyloxyimidazo[1,2-b]pyridazines.

A series of novel (imidazo[1,2-b]pyridazin-6-yl)oxyalkylsulfonamides was synthesized and evaluated for the ability to inhibit platelet activating factor (PAF)-induced bronchoconstriction in guinea pigs. The compounds bearing a gem-dialkyl or a cycloalkylidene group at the 2 position of the sulfamoylpropyloxy group in the side chain were found to have potent activity. Among them, 3-(imidazo[1,2-b]pyridazin-6-yl)oxy-2,2-dimethylpropanesulfonam ide (6) showed excellent anti-asthmatic activity and the longest duration of action. The compounds bearing a methyl group at the 7 or 8 position of the imidazo[1,2-b]pyridazine ring were found to have enhanced activity. Among them, 3-(7-methylimidazo[1,2-b]pyridazin-6-yl)oxy-2,2- dimethylpropanesulfonamide (25) showed the most potent inhibitory effect, and its anti-asthmatic effect in an experimental model of allergic asthma was superior to that of theophylline. The structure-activity relationships in this series of compounds are discussed.

Animals↗
PubMed1996

Antagonistic effect of YM022, an antiulcer agent in rats, on human cholecystokinin (CCK)B/gastrin receptor.

We recently isolated a cDNA clone for the human cholecystokinin (CCK)B/gastrin receptor and permanently expressed this receptor cDNA in NIH-3T3 cells. [125I]CCK-8 specifically bound to the membrane of the transfectant, and this binding was displaced by unlabeled CCK-8 with an IC50 of 0.32 nM. Treatment of these cells with CCK-8 increased the intracellular Ca2+ concentration with an EC50 of 0.30 nM. Using these cells expressing functional human CCKB/gastrin receptors, we investigated the pharmacological properties of (R)-1-[2,3-dihydro-1-(2'-methylphenacyl)-2-oxo-5-phenyl-1H-1, 4-benzodiazepin-3-yl]-3-(3-methylphenyl) urea (YM022), a potent and selective CCKB/gastrin receptor antagonist in rats. YM022 potently inhibited [125I]CCK-8 binding to the membrane with an IC50 of 55 pM and CCK-8-induced Ca2+ mobilization with that of 7.4 nM. On the other hand, its racemate and enantiomer more weakly inhibited this binding (IC50 of 110 pM and 11 nM, respectively) and Ca2+ mobilization (IC50 of 18 nM and 94 nM, respectively). These results indicate that YM022 stereoselectively recognizes the human CCKB/gastrin receptor as a potent antagonist and that the established transfectant is useful for characterization of human CCKB/gastrin-receptor ligands.

Animals↗
PubMed1996

Regulation of mouse placental lactogen secretion by factors secreted from the pituitary in vitro.

The effect of factors secreted from the pituitary on mouse placental lactogen I (mPL-I) and mPL-II secretion in vitro was examined. Co-culture of mouse placental cells from day 7 of pregnancy with the pituitary cells of the mother significantly stimulated mPL-I secretion but did not regulate mPL-II secretion. The effect on mPL-I secretion was dependent on the number of pituitary cells. The conditioned medium of pituitary cells also significantly stimulated mPL-I secretion but did not regulate mPL-II secretion. The stimulatory effect of mPL-I secretion was dependent on the volume of the conditioned medium. The number of cells containing mPL-I assessed by immunocytochemistry was increased by the co-culture in a cell number-dependent manner. Northern blot analysis for mPL-I indicated that treatment of placental cells with the pituitary-conditioned medium results in an increase of mPL-I gene expression. These findings suggest that factors secreted from the pituitary directly regulate mPL-I secretion, but not mPL-II secretion, before midpregnancy in vivo.

Animals↗
PubMed1996

Cyclic AMP regulates expression of activin and inhibin subunit mRNAs in the mouse placenta and decidua: a short communication.

We examined whether cAMP regulates expression of activin/inhibin subunit mRNAs in the mouse placenta and decidua. 8-Bromo-cAMP inhibited expression of inhibin alpha-subunit mRNA but stimulated expression of activin/inhibin beta A- and beta B-subunit mRNAs in placenta and decidua by the 3rd day of culture. Forskolin and cholera toxin also regulated expression of activin/inhibin subunit mRNAs in the same manner. However, activin-A did not affect expression of activin/inhibin subunit mRNAs. These findings suggest that cAMP inhibits inhibin production but stimulates activin production in the mouse placenta and decidua.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1996

8-Bromo-cAMP inhibits glucose transport activity in mouse placental cells in culture.

Glucose plays an important role in fetal development and energy metabolism. Facilitative glucose transporter-1 (GLUT1) has been found in placenta. However, little is known about GLUT1 modulation in placental cells. To examine changes in mouse placental GLUT1 levels caused by 8-bromo-cAMP, we performed 2-deoxyglucose uptake experiments, Northern blot analysis and immunoblot analysis using a primary mouse placental cell culture. Immunohistochemical analysis showed that GLUT1 was localized to the ectoplacental cone and the labyrinth zone of mouse placentas on days 7 and 11 of pregnancy respectively. Treatment of mouse placental cells with 250 mumol/l 8-bromo-cAMP resulted in a significant (P < 0.01) decrease in glucose uptake on days 2-5 of culture. The inhibitory effect of 8-bromo-cAMP on glucose uptake was concentration-dependent. Glucose uptake was also inhibited by 100 micrograms/l cholera toxin and by 0.1 mmol/l forskolin. Northern blot and immunoblot analysis revealed that both GLUT1 mRNA and protein levels were also decreased by 8-bromo-cAMP. These findings suggest that 8-bromo-cAMP inhibits glucose transport activity in mouse placental cells in culture.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1996

Localization of human placental glucose transporter 1 during pregnancy. An immunohistochemical study.

To elucidate the potential roles of glucose transporter 1 (GLUT1) in human placenta during pregnancy, we examined the localization of GLUT1 in human placenta at various stages by immunohistochemistry with an anti-GLUT1 antibody by use of both light and electron microscopy. Specific staining for GLUT1 was localized on the apical brush border and along the basal plasma membrane of the syncytiotrophoblasts. The staining at the apical side was more intense than that at the basal side during the early stages of gestation. In later gestational stages, however, the staining pattern at the apical side became blurred and the staining intensity at the basal side increased. The cytotrophoblasts, seen embedded in the basal part of the syncytiotrophoblasts, seemed to show immunoreactivity for GLUT1 along the plasma membranes at the light-microscopic level. However, immuno-electron microscopic analysis with either pre- or post-embedding methods revealed that specific staining for GLUT1 was hardly observed on the cytotrophoblasts, but the cytotrophoblasts were often surrounded by immunoreactive processes of syncytiotrophoblasts. The blood capillaries and erythrocytes in the stroma of placental villi were always immunoreactive for GLUT1 throughout pregnancy. These findings suggest that GLUT1 may play a vital role in human pregnancy.

Capillaries↗
PubMed1995

Expression and localization of glucose transporter 1 (GLUT1) in the rat oviduct: a possible supplier of glucose to embryo during early embryonic development.

The oviduct fluid mainly derived from the oviduct epithelium is reported to provide the environment necessary for embryonic development. To elucidate the origin of glucose in the oviduct fluid, we examined the expression and localization of glucose transporter 1 (GLUT1) in the rat oviduct by Northern blot analysis, immunoblot analysis and immunohistochemistry using both light and electron microscopy. Northern blot and immunoblot analyses both showed the presence of the GLUT1 mRNA and protein. Specific staining for GLUT1 was observed in the ampulla and the isthmus, but only slightly in the fimbria and the utero-tubal junction. Staining was confined to the luminal surface of the epithelial mucosa. Immunoelectron microscopic analysis revealed that GLUT1 was observed only on the surface of the microvilli in non ciliated secretory cells, but not in ciliated cells. These findings suggest that GLUT1 plays an important role in the glucose transfer from the oviduct epithelium into the lumen and in maintaining the adequate glucose concentration of the oviduct fluid for embryonic development in rat oviduct.

Animals↗
PubMed1995

Regulation of mouse soluble interleukin-6 receptor secretion by decidua.

Regulation of msIL-6R secretion by mIL-6 and 8-bromo cAMP was examined using primarily cell culture of mouse decidua. Mouse decidua on day 11 of pregnancy was digested by collagenase and decidual cells were cultured for up to 3 days. Addition of mIL-6 and 8-bromo cAMP resulted in significant inhibition of msIL-6R secretion from cultured mouse decidual cells by the 2nd day in culture. The effects were dose-dependent and the lowest concentrations of mIL-6 and 8-bromo cAMP which cause significant inhibition of msIL-6R secretion were 250 pM and 50 microM, respectively. Northern blot analysis indicated that treatment of decidual cells with 8-bromo cAMP significantly decreased the steady-state level of mIL-6R mRNA. However, treatment of decidual cells with mIL-6 did not affect the steady-state level of mIL-6R mRNA. These results suggest that mIL-6 decreases msIL-6R secretion without changing of the steady-state level of mIL-6R mRNA, and that cAMP is one of the second messengers in mouse decidual cells involved in this down regulation of msIL-6R secretion.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1995

A truncated isoform of human CCK-B/gastrin receptor generated by alternative usage of a novel exon.

An isoform cDNA of CCK-B/gastrin receptor was isolated from human stomach. This cDNA differed from initially cloned cDNA only in the 5'-end region and encoded a truncated isoform (delta CCK-B) in which the putative N-terminal extracellular domain of the CCK-B/gastrin receptor was completely lost. Isolation of genomic CCK-B/gastrin receptor DNA revealed that this transcript is generated by alternative usage of a novel exon, termed exon 1b. Human stomach expressed both transcripts of delta CCK-B and entire CCK-B/gastrin receptor (CCK-BR), whereas human stomach cancer cell line AGS exclusively expressed delta CCK-B transcripts. Transfection of COS-7 with delta CCK-B cDNA led to the appearance of binding sites for 125I-CCK-8. Its ligand selectivity was different from that of CCK-BR. These results suggest the molecular diversity in CCK-B/gastrin receptor subtypes.

Alternative Splicing↗