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ABSTRACT: Asthma prevalence and severity are higher in females than in males after puberty. The underlying mechanisms of this gender difference are not fully understood. More severe airway inflammation in female mice has been reported to be associated with higher levels of T helper type 2 (Th2) cytokines in asthma models. The aim of this study was to investigate sex differences in CD4+ and CD8+ T cell functions in Th2 cytokine production.
Splenocytes from naive mice were stimulated with anti-CD3/CD28 antibodies and the proportions of CD4+ and CD8+ T cells were analyzed. CD4+ T cells were stimulated in the presence of CD8+ T cells. The concentrations of interleukin (IL)-5, IL-10 and interferon (IFN)-γ in the cultures were measured.
The concentration of IL-5, but not IFN-γ, was significantly higher in female splenocytes than in male splenocytes. There were no sex differences in the proportions of CD4+ and CD8+ T cells in the splenocytes. Although the IL-5 production levels in male and female CD4+ T cells were similar, IL-5 production in male CD4+ T cells, but not female CD4+ T cells, was suppressed by both male and female CD8+ T cells. While IL-5 and IL-10 were not detected in the cultures from both male and female CD8+ T cells, IFN-γ concentration in female CD8+ T cells was significantly higher than in male CD8+ T cells.
The sex difference in the sensitivity of CD4+ T cells to CD8+ T cell suppression might contribute to the sex difference in IL-5 production by splenocytes.
International Archives of Allergy and Immunology 01/2012; 158 Suppl 1:35-41. · 2.40 Impact Factor
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ABSTRACT: Psychological stress has a recognized association with asthma symptoms. Using a murine model of allergic asthma, we recently demonstrated the involvement of μ-opioid receptors (MORs) in the central nervous system in the stress-induced exacerbation of airway inflammation. However, the involvement of MORs on neurons and immunological alterations in the stress asthma model remain unclear.
MOR-knockout (MORKO) mice that express MORs only on noradrenergic and adrenergic neurons (MORKO/Tg mice) were produced and characterized for stress responses. Sensitized mice inhaled antigen and were then subjected to restraint stress. After a second antigen inhalation, bronchoalveolar lavage cells were counted. Before the second inhalation, bronchial lymph node (BLN) cells and splenocytes from stressed and non-stressed mice were cultured with antigen, and cytokine levels and the proportions of T cell subsets were measured.
Stress-induced worsening of allergic airway inflammation was observed in wild-type and MORKO/Tg mice but not MORKO mice. In wild-type stressed mice, IFN-γ/IL-4 ratios in cell culture supernatants and the proportion of regulatory T cells in BLN cell populations were significantly lower than those in non-stressed mice. These differences in BLN cells were not observed between the stressed and non-stressed MORKO mice. Restraint stress had no effect on cytokine production or T cell subsets in splenocytes.
Restraint stress aggravated allergic airway inflammation in association with alterations in local immunity characterized by greater Th2-associated cytokine production and a reduced development of regulatory T cells, mediated by MORs.
Allergology International 12/2011; 61(2):245-58.
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Yuki Fujii,
Shigeki Sugawara,
Daisuke Araki,
Tasuku Kawano,
Takeo Tatsuta,
Kohta Takahashi,
Sarkar M A Kawsar,
Ryo Matsumoto,
Robert A Kanaly,
Hidetaro Yasumitsu,
Yasuhiro Ozeki,
Masahiro Hosono,
Taeko Miyagi,
Sen-Itiroh Hakomori, Motoaki Takayanagi,
Kazuo Nitta
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ABSTRACT: A novel anticancer mechanism of catfish (Silurus asotus) egg lectin (SAL) was found to occur via the down-regulation of the membrane transopter protein, MRP1 (multidrug resistance associate protein-1) on Burkitt's lymphoma cells through Gb3(Galα1-4Galβ1-4Glc)-glycosphingolipid. Although SAL did not influence the viability of the cells directly, only 10 and 100 ng/mL of vincristine and etoposide, respectively induced anticancer effects when the lectin was applied in conjunction with these drugs. These phenomena were specifically inhibited by the co-presence of the α-galactoside, melibiose, which is a strong haptenic sugar of SAL that mimicks Gb3. The degree of expression regulation of the transporter proteins on the cells surface was investigated through the examination of the binding between SAL and Gb3-glycosphingolipid by immunological and molecular biological procedures. PCR data showed that MRP1 was more highly expressed when compared to another ATP-binding cassette family, multi-drug resistant protein and the expression levels of MRP1 on the cells were specifically dose- and time-dependently depleted by the addition of SAL. These results were also evaluated by immunological procedures using FACS and western-blotting. Small interfering RNA coding a part of MRP1 was transfected to Raji cells to knock down the protein, and cell death was increased by 10% when vincristine was administered at a concentration as low as 10 ng/mL compared to non-transfected cells. These results indicated that SAL possesses the potential to enhance the anticancer activites of low-concentrations of vincristine by the down-regulating the MRP1 gene expression to inhibit the multidrug resistance by binding to the target ligand Gb3-glycosphingolipid on Burkitt's lymphoma cells.
The Protein Journal 11/2011; 31(1):15-26. · 1.04 Impact Factor
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ABSTRACT: Before puberty, the prevalence and severity of asthma are higher in boys than in girls, but this pattern is reversed after puberty. The underlying mechanisms of these gender differences in asthma are not fully understood. Using murine models of allergic asthma, a sex difference in Th2 cytokine production has been suggested to contribute to the gender differences in asthma. Therefore, we determined which subsets of T cells are involved in the sex difference in Th2 cytokine production.
Splenocytes from wild-type mice and CD4+ T cell-, CD8+ T cell-, and iNKT cell-deficient mice were stimulated with anti-CD3/CD28 antibodies for 3 days, and the concentrations of IL-4, IL-5, IL-13, and IFN-γ in the cultures were measured by ELISA.
IL-5, but not IL-4 and IL-13, concentrations in culture derived from female wild-type mice were significantly higher than those in male wild-type mice. The sex difference in IL-5 concentrations was not observed in the cultures of splenocytes from CD4+ and CD8+ T cell-deficient mice. The disappearance of the sex differences in CD4+ and CD8+ T cell-deficient mice was attributable to a decrease in IL-5 concentration in female mice and an increase in IL-5 concentration in male mice. In iNKT cell-deficient mice, the sex difference was still observed. There was no significant difference between the sexes in any type of mice with respect to IFN-γ production.
There was a sex difference in IL-5 production by splenocytes stimulated by TCR activation. The difference might be attributable to sex differences in CD4+ and CD8+ T cell functions.
International Archives of Allergy and Immunology 01/2011; 155 Suppl 1:21-6. · 2.40 Impact Factor
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ABSTRACT: The severity of asthma after puberty is higher in women than in men. Increased numbers of eosinophils in the airways of female mice after antigen challenge was associated with increased levels of T helper (Th)2 cytokines at the site of inflammation, and in human and mouse studies, the profile of cytokines produced by immune cells from women showed greater Th2 predominance. The aim of this study was to investigate gender differences in the development of Th2 immune responses.
Male and female C57BL/6 mice were sensitized with ovalbumin. Cells prepared from bronchial lymph nodes were cultured in the absence or presence of ovalbumin. Cytokine concentrations in the culture supernatants were measured, and IL-5 and GATA-binding protein 3 (GATA-3) gene expression were evaluated. T-cell subsets were analysed using specific surface markers.
The concentrations of IL-4, IL-5, IL-13 and IL-10, but not interferon-gamma or transforming growth factor-beta(1), were higher in cell supernatants from female mice than in those from male mice. IL-5 and GATA-3 gene expressions were higher in cells from women than in cells from men. The numbers of CD3(+)CD4(+)T1/ST2(+) cells, but not CD3(+)CD4(+) or CD4(+)CD25(+) cells, were significantly higher in cells from women than in cells from men.
Greater antigen-induced Th2 cytokine production by bronchial lymph node cells from female mice was associated with enhanced Th2 cell differentiation and increased expression of the Th2-specific transcription factor, GATA-3.
Respirology 03/2010; 15(4):629-35. · 2.42 Impact Factor
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ABSTRACT: The apoptosis and cell cycle effect of doxorubicin were evaluated in the presence and absence of ivermectin in mouse doxorubicin-resistant P388 leukaemia cells.Ivermectin (2 μM) increased the sensitivity to doxorubicin of multidrug resistant (MDR) mouse leukemic P388 cells and significantly enhanced the apoptosis and intracellular accumulation of doxorubicin in resistant cells, but had no effect on parent cells. Using the fluorescent potential probe, 3,3′-dihexyl-oxacarbocyanine, we found that ivermectin induced a plasma membrane potential increase in resistant cells. Ivermectin also enhanced doxorubicin-induced G2/M blockade of the cell cycle in resistant cells. It is possible that ivermectin could reverse resistance by direct interaction with the P-glycoprotein or other components of the altered MDR cell membrane.
Pharmacy and Pharmacology Communications. 02/2010; 6(3):129 - 134.
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ABSTRACT: Psychological stress has a recognized association with asthma symptoms. However, the mechanisms linking stress to the exacerbation of asthma are not well defined. mu-Opioid receptors (MOR) have been shown to be involved in the shift of the immune system toward a Th2-predominant response caused by psychological stress.
To test the hypothesis that MOR play a role in the worsening of allergic airway inflammation evoked by psychological stress.
Sensitized mice were exposed to restraint stress followed by antigen challenge. The levels of corticosterone and ovalbumin (OVA)-specific IgE in the blood and the levels of inflammatory cells and cytokine contents in bronchoalveolar lavage fluid were compared between stressed and nonstressed mice. The effects of MOR gene deletion and MOR antagonists/agonists were also investigated.
Stress exposure was confirmed by an increase in corticosterone levels. Although OVA-specific IgE levels were not significantly different, the numbers of inflammatory cells and Th2 cytokine levels after antigen challenge in stressed mice were significantly higher than in nonstressed mice. MOR gene deletion ameliorated the stress-induced worsening of antigen-induced airway inflammation, and the administration of morphine, a MOR agonist, reproduced the stress-induced antigen-induced airway inflammation. Selective blocking of MOR in the central nervous system (CNS) significantly reduced stress-induced inflammatory exacerbation, but the blocking of peripheral MOR did not.
MOR in the CNS are involved in psychological stress-induced aggravation of allergic airway inflammation.
International Archives of Allergy and Immunology 02/2010; 152(4):342-52. · 2.40 Impact Factor
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Kazumasa Suzuki,
Katsushi Nishimaki,
Kaori Okuyama,
Tadashi Katoh,
Minoru Yasujima,
Junichi Chihara,
Akira Suwabe,
Yoko Shibata,
Choichiro Takahashi,
Hiroaki Takeda,
Shiro Ida,
Mitsuo Kaku,
Akira Watanabe,
Toshihiro Nukiwa,
Kazunao Niitsuma,
Keiji Kanemitsu, Motoaki Takayanagi,
Isao Ohno
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ABSTRACT: Streptococcus pneumoniae is a common cause of respiratory tract infections (RTIs). The prevalence of Streptococcus pneumoniae strains with reduced susceptibility to antimicrobial agents has dramatically increased worldwide. Susceptibility to nine antimicrobial agents and serotypes were determined among 1,644 Streptococcus pneumoniae strains isolated from patients with RTIs in the Tohoku district of Japan from October to December every year from 1998 to 2007. The prevalence of penicillin G-nonsusceptible Streptococcus pneumoniae (PNSP) strains increased gradually from 48.5% in 1998, reached a statistical peak in 2004 (65.1%) and then decreased to 51.5% in 2007. Streptococcus pneumoniae strains with each serotype 3, 6, 19 and 23 were constantly detected, and the distribution of these serotypes in PNSP strains did not significantly change during the study period. A trend of Streptococcus pneumoniae strains nonsusceptible to other beta-lactams tested was similar to that of PNSP strains, except for cefditoren, to which the resistance rate was < 20% throughout the analysis period. The prevalence of strains nonsusceptible to erythromycin and minocycline were consistently > 60%. Almost all penicillin G-resistant Streptococcus pneumoniae (PRSP) strains were resistant to both erythromycin and minocycline throughout the analysis period. The prevalence of strains resistant to fluoroquinolones tested were < 3% over the study period. Our longitudinal surveillance demonstrated for the first time that decreased prevalence of both beta-lactam- and multidrug-resistant strains has been occurring since 2004 in a region of Japan. Careful monitoring of antimicrobial susceptibility of Streptococcus pneumoniae should be continued.
The Tohoku Journal of Experimental Medicine 01/2010; 220(1):47-57. · 1.24 Impact Factor
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ABSTRACT: The pathogenesis of airway inflammation in diffuse panbronchiolitis (DPB) is unknown. Neutrophil survival-enhancing activity, partially mediated by granulocyte-macrophage colony-stimulating factor (GM-CSF), has been shown in the sputum from DPB patients. This study investigated the mechanisms of GM-CSF expression in the airways of DPB patients. This involved examining the effects of mucoid Pseudomonas aeruginosa strains derived from chronically colonized patients with DPB on neutrophil survival and GM-CSF expression.
Neutrophils from healthy subjects were cultured with the culture supernatants of P. aeruginosa isolated from sputum of DPB patients in the presence or absence of anti-GM-CSF and anti-GM-CSF receptor (alpha chain) antibodies, and viable neutrophils were counted daily. GM-CSF gene expression in neutrophils was evaluated by RT-PCR.
Neutrophils cultured with the culture supernatants showed significantly prolonged survival, compared with neutrophils cultured with the control broth. The neutrophil survival-enhancing activity in the culture supernatants was lost by heating. The enhanced survival of neutrophils was abolished in the presence of anti-GM-CSF and anti-GM-CSF receptor (alpha chain) antibodies. GM-CSF mRNA was detected in neutrophils cultured with the bacterial supernatants, but not in those with the control broth.
P. aeruginosa-derived factors (rich in proteins) stimulated neutrophils to synthesize GM-CSF, which enhanced neutrophil survival in an autocrine/paracrine fashion.
Respirology 10/2007; 12(5):664-9. · 2.42 Impact Factor
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ABSTRACT: Psychological stress has long been recognized to be associated with asthma symptoms. There appear to be individual differences in the susceptibility to even the same kind of stress, and furthermore, stress responses are different between the types of the stress, acute and chronic, even in the same person. However, the mechanisms linking stress to asthma are not well defined. Psychological stress upregulates the expression of endogenous opioids. The opioids stimulate the hypothalamus-pituitary-adrenal axis and sympathetic and adrenomedullary system, through the activation of mu-opioid receptor (MOR) to release stress hormones, such as cortisol and catecholamines, respectively. These hormones can modulate immune responses via the induction of Th1 immunity.
Female BALB/c and C57BL/6, wild and MOR-deficient, mice sensitized with ovalbumin (OVA) were exposed to OVA with or without either acute or chronic restraint stress. Airway inflammation was evaluated by the measurement of the number of inflammatory cells and cytokine contents in bronchoalveolar lavage fluids.
In BALB/c mice, but not in C57BL/6 mice, the number of total cells, eosinophils and lymphocytes in the acute stress group were significantly decreased compared with those in the non-acute stress group. In contrast, chronic stress significantly increased the cell numbers and the contents of IL-4 and IL-5 in both mouse strains. Furthermore, these exacerbations were abolished in MOR-deficient mice.
These results suggest that acute stress modifies the allergic airway responses distinctively depending on the genetic background, and MOR is involved in the chronic psychological stress-induced exacerbation of allergic airway inflammation.
Allergology International 04/2007; 56(1):29-35.
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ABSTRACT: Peroxisome prolifelator-activated receptor gamma (PPARgamma) is a ligand-activated transcription factor, through which PPARgamma agonists have been demonstrated to down-regulate inflammatory cell functions. Recently, the agonists are reported to exert, in some conditions, their inhibitory actions independently of PPARgamma. Previously, we showed that a PPARgamma agonist, troglitazone, inhibited cysteinyl (Cys)-leukotrienes production in RBL-2H3 cells after IgE receptor triggering. Here we examined whether the inhibition of cycteinyl-leukotrienes production in the cells was dependent on the activation of PPARgamma. A PPARgamma agonist, ciglitazone, significantly inhibited Cys-leukotrienes, but not prostaglandin D2, production. The inhibition was not attenuated by the pretreatment with a PPARgamma antagonist. Ciglitazone did not alter the mRNA expression of acyl-coenzyme A binding protein, the gene expression of which is up-regulated by PPARgamma, nor induce the nucleus translocation of PPARgamma. These results suggest that the inhibition by PPARgamma agonists of Cys-leukotrienes production in RBL-2H3 cells after IgE receptor triggering is not through the activation of PPARgamma.
European Journal of Pharmacology 11/2005; 521(1-3):21-8. · 2.52 Impact Factor
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ABSTRACT: A novel C-type lectin (OLABL) was isolated from the eggs of shishamo smelt [Osmerus (Spirinchus) lanceolatus] by affinity chromatography on asialofetuin-Sepharose. OLABL had a molecular mass of 29 kDa on SDS-PAGE under nonreducing conditions and two subunits with masses of 15 kDa (OLABL-H) and 14 kDa (OLABL-L) under reducing conditions. Thus, OLABL is a heterodimeric protein. cDNA sequence analysis revealed that the H- and L-subunits of OLABL were composed of 137 and 136 amino acid residues, respectively, and showed almost identical (95%) sequences, with slight differences in the N-terminal and C-terminal regions. Since each subunit contained only the characteristic motif of C-type lectin-like domain (CTLD), EPN-E-WND, OLABL is a member of group VII of the CTLD-containing protein family. Although OLABL had an EPN sequence that is known as a mannose-specific motif found in the collectin family, OLABL agglutinated rabbit erythrocytes without the addition of Ca(2+) ion, and this activity was inhibited by l-rhamnose and d-galactose derivatives, but not by d-mannose and d-glucose. These results indicate that OLABL has similar characteristics to AJL-2, a calcium-independent lactose specific lectin isolated from Japanese eel skin mucus. Recombinant OLABLs (rHisOLABLs), His-tagged homodimers of the H- and L-subunits, were refolded from inclusion bodies expressed by Escherichia coli. rHisOLABL-L was recovered as a soluble form, but rHisOLABL-H was hardly dissolved in a renaturing buffer. The specific activities of rHisOLABL-L, rHisOLABL-H, and native OLABL were 500, 36, and 20, respectively. These findings suggest that the combination of subunits may affect the solubility and activity of these dimeric form lectins.
Biochimica et Biophysica Acta 10/2005; 1725(2):160-73. · 4.66 Impact Factor
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ABSTRACT: Two C-type lectins (OLLafs and OLLafl) were isolated from Osmerus (Spirinchus) lanceolatus eggs using asialofetuin-Sepharose column. OLLafs and OLLafl were eluted with 0.2 M sucrose and 0.2 M lactose from the same column, respectively. OLLafl has been estimated to be a heterodimeric protein composed of H- and L-subunit and involved C-type lectin like domain (CTLD). In this study we revealed that OLLafs was a homodimeric protein composed of L-subunit of OLLafl. Although adding EDTA diminished the hemagglutinating activity of OLLafs, the activity of OLLafl was not influenced. Recombinant lectins (rOLLafl-H and -L) and mutant lectins replaced Cys(123, 131 and 136) with Ala (mOLLafl-L(123, 131 and 136)) were established. The activity of mOLLafl-L(136) was comparable to rOLLafl-L, and rOLLafl-H was 15 times lower than rOLLafl-L. On the other hand, the activity of mOLLafl-L(123) and mOLLafl-L(131) were lower than that of rOLLafl-H. Therefore, Cys(136) may not participate in hemagglutinating activity of rOLLafl-L. In contrast, Cys(123) and Cys(131) may partially contribute this activity. Although hemagglutination inhibition profiles of rOLLafl-L, rOLLafl-H and mOLLafl-L(136) were similar, m-OLLafl-L(131)-induced hemagglutination was not inhibited by any sugars tested even at a concentration of 150 mM. Then, Cys(131) may directly contribute to the sugar-binding capacity of OLLafl. Affinities of mOLLafl-L(123) for these sugars were lower than the others. These results suggest that Cys(136) might contribute to the intermolecular disulfide bond in the rOLLafl-L dimer, and that the intramolecular disulfide bond concerning Cys(131) might important for lectin activity.
Biological & Pharmaceutical Bulletin 06/2005; 28(5):791-6. · 1.66 Impact Factor
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ABSTRACT: Selective inhibitors of phosphodiesterases (PDEs) have been suggested to have anti-inflammatory effects on bronchial asthma through the inhibition of chemotaxis, adhesion, degranulation, the respiratory burst, and survival prolongation of eosinophils. However, the mechanisms by which these agents inhibit eosinophil survival remain unclear. We therefore investigated the possible mechanisms of inhibitory effects of selective inhibitors of PDE 3 (cilostazol) and PDE 4 (rolipram) on granulocyte-macrophage colony-stimulating factor (GM-CSF)-mediated eosinophil survival. Purified blood eosinophils were cultured with medium alone or GM-CSF (0.01 ng/ml) in the presence or absence of the agents for up to 6 d. DNA was extracted from freshly isolated eosinophils and eosinophils cultured for 2 d with medium alone, GM-CSF, or GM-CSF in the presence of the agents, and analyzed using agarose gel electrophoresis. The presence of rolipram (10(-4), 10(-5), 10(-6) M), but not cilostazol, significantly inhibited eosinophil survival at days 2, 4, and 6. A laddering pattern was observed in the DNA of eosinophils cultured with medium alone and with GM-CSF in the presence of rolipram. The results reveal that selective PDE 4 inhibitors inhibit GM-CSF-mediated eosinophil survival through the induction of apoptosis.
Biological & Pharmaceutical Bulletin 04/2005; 28(3):515-9. · 1.66 Impact Factor
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ABSTRACT: Rhamnose-binding lectin from catfish (Silurus asotus) eggs (SAL) has the ability to induce externalization of phosphatidylserine (PS), followed by cell shrinkage in globotriaosylceramide (Gb3)-expressing Burkitt's lymphoma Raji cells. Because phospholipid scramblase and aminophospholipid translocase did not participate in SAL-induced PS externalization, we examined the relationship of ATP-binding cassette (ABC) transporters, such as multidrug resistance (MDR) 1 P-glycoprotein (MDR1 P-gp) and MDR-associated protein 1 (MRP1), for translocation of PS. Since cyclosporin A (MDR1 P-gp inhibitor) but not MK571 (MRP1 inhibitor) inhibited SAL-induced PS externalization, it was suggested that MDR1 P-gp is involved in this phenomenon. On the other hand, SAL activated both of the ABC transporters for efflux of rhodamine123 (MDR1 P-gp substrate, Rho123) and 5-carboxyfluorescein diacetate (MRP1 substrate, 5-CFDA) in Raji cells. In contrast, SAL did not activate these two transporters in Gb3-negative cell lines, such as K562 and doxorubicin-resistant K562 cells, involving not only PS externalization but also efflux of Rho123 or 5-CFDA. Since Gb3 and both transporters in Raji cells are located in the glycosphingolipid-enriched microdomain (GEM), it is suggested that the binding of SAL to Gb3 localized in the GEM specifically induces MDR1 P-gp activation in Raji cells.
Biological & Pharmaceutical Bulletin 04/2005; 28(3):434-41. · 1.66 Impact Factor
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ABSTRACT: Hemagglutinating activity for glutaraldehyde-fixed, trypsinized human and rabbit erythrocytes was obtained from carp (Ciprinus capio) egg extract. This activity was inhibited by sulfated glycans such as heparin and fucoidan, and salmon testis DNA (stDNA), but not by mono- and oligosaccharides. The active fraction from hydroxyapatite column, HA400, caused aggregation of stDNA at a concentration of 125 microg/ml. HA400-induced aggregation was inhibited by heparin and fucoidan at the concentrations of 25 and 50 microg/ml, respectively. HA400 also bound to short size single- or double-stranded oligonucleotide in dot blot hybridization analysis using 32P-labeled probe. HA400 was further purified by heparin-Sepharose column chromatography, and consequently, active fractions (HS2, HS3 and HS4) were obtained. Hemagglutinating activities of HS2 and HS4 were 16 and 32 times higher than that of HS3, respectively. In contrast, HS3 showed the highest DNA aggregating activity among these fractions, indicating that the activities of DNA aggregation and hemagglutination were separated by heparin-Sepharose column chromatography. HS3-induced stDNA aggregation was inhibited by 0.5 M NaCl or above. Southwestern blot analysis revealed that the DNA aggregating protein (DAP) possessing oligonucleotide-binding capacity was the 18 kDa protein. The N-terminal amino acid sequence of DAP was LKEGECEVCVGFLGR having partial homology with DNA-binding protein HU-alpha, but showed no homology with nucleohistone proteins. These results indicate that there is a novel, non-histone DAP in carp eggs.
Biological & Pharmaceutical Bulletin 02/2005; 28(1):13-8. · 1.66 Impact Factor
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ABSTRACT: Calcitonin gene-related peptide (CGRP) and adrenomedullin (ADM) belong to a calcitonin-family of regulatory peptides. Receptors for CGRP and ADM have been suggested to be present on both mucosal (MMC) and connective tissue (CTMC) type of mast cells, based on histamine release by these peptides. Recently, it was reported that mRNA for ADM receptors, but not for CGRP receptors, was expressed in rat peritoneal mast cells, a representative of type CTMC. However, mRNA expression for the receptors in MMC has not been studied yet. Therefore, we examined whether mRNAs encoding CGRP or ADM receptor subunit, RDC-1, calcitonin receptor-like receptor (CRLR), and receptor activity-modifying proteins (RAMPs) are present, and if so, whether their expression is modified by IgE receptor triggering, in a mucosal type mast cell line, rat basophilic leukemia (RBL-2H3) cells using RT-PCR. RBL-2H3 cells constitutively express mRNA for RDC-1, CRLR, RAMP3 but not that for RAMP1 and RAMP2, and IgE receptor triggering was shown neither to induce the gene expression of RAMP1 and RAMP2, nor to enhance that of RDC-1, CRLR or RAMP3. These results indicate that RBL-2H3 cells posses receptors for both CGRP and ADM, suggesting various functions of these peptides in physiological and pathophysiological conditions where mast cells of the mucosal type are involved.
Biological & Pharmaceutical Bulletin 07/2004; 27(6):896-8. · 1.66 Impact Factor
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ABSTRACT: The role of p44/42 mitogen-activated protein kinase (MAPK) in the expression of intercellular adhesion molecule-1 (ICAM-1) in NCI-H292 cells, a human bronchial epithelial cell line, was analyzed. Treatment with the protein kinase C (PKC) activator 12-O-tetradecanoylphorbol 13-acetate (TPA) (16.2 nM) or interferon-gamma (IFN-gamma) (100 U/ml) induced phosphorylation of p44/42 MAPK. The MEK inhibitor U0126 (0.1 to 10 microM) enhanced the TPA-induced ICAM-1 expression but not the IFN-gamma-induced one. U0126 also enhanced the ICAM-1 expression induced by two other PKC activators teleocidin (22.5 nM) and aplysiatoxin (14.9 nM). Furthermore, PD98059 (0.5 to 50 microM), another MEK inhibitor, enhanced the TPA-induced ICAM-1 expression as well. The inhibitor of p38 MAPK SB203580 did not affect the TPA-induced ICAM-1 expression. BAY11-7082, an inhibitor of nuclear factor kappaB (NF-kappaB) activation, and MG132, a 26S proteasome inhibitor, reduced the TPA-induced ICAM-1 expression but not the IFN-gamma-induced one. TPA partially decreased the level of IkappaB-alpha and the reduction was further augmented by U0126 in a concentration-dependent manner. These findings suggested that, in NCI-H292 cells, p44/42 MAPK suppresses PKC activator-induced NF-kappaB activation, thus negatively regulating the PKC activator-induced ICAM-1 expression but not the IFN-gamma-induced one.
Life Sciences 07/2004; 75(4):435-46. · 2.53 Impact Factor
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ABSTRACT: The mechanism of action of anti-rheumatic gold compounds on 12-O-tetradecanoylphorbol 13-acetate (TPA)-induced prostaglandin E(2) (PGE(2)) production in rat peritoneal macrophages were examined. Auranofin (AF) at 3-10 muM inhibited TPA-induced PGE(2) production in a concentration-dependent manner. In the pharmacological experiments, prostaglandin G/H synthase (PGHS)-2-dependent PGE(2) production was inhibited by 10 muM of AF. The enzyme activities of both PGHS-1 and PGHS-2 were not affected by the 10 muM AF. Other gold compounds, aurothioglucose (ATG) and aurothiomalate (ATM) did not inhibit PGE2 production at 10 muM. AF decreased the PGHS-2 protein content, but had no effect on the PGHS-1 protein content. AF at 3-10 muM decreased the PGHS-2 messenger RNA (mRNA) level by RT-PCR determination. Then, the effect of AF on nuclear factor kappa B (NF-kappaB), one of the transcription factors known to regulate transcription of a group of proinflammatory proteins, was determined. AF at 1-10 muM inhibited nuclear translocation of NF-kappaB in a concentration-dependent manner. ATG and ATM at 10 muM did not inhibit NF-kappaB nuclear translocation, but with 20 h preincubation, ATG and ATM inhibited PGE(2) production and NF-kappaB nuclear translocation. AF, ATG, and ATM did not affect the binding of NF-kappaB to its specific DNA. These observations may suggest that the effects of gold compounds on the inhibition of NF-kappaB nuclear translocation plays one of the major role in its anti-inflammatory effects in rat peritoneal macrophages.
Current Drug Targets - Inflammation & Allergy 10/2003; 2(3):216-23.
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ABSTRACT: Auranofin, aurothioglucose and aurothiomalate (10 microM each) inhibited 12-O-tetradecanoylphorbol 13-acetate (TPA, 16.2 nM)-induced nuclear translocation of nuclear factor-kappa B (NF-kappaB), and production of nitric oxide (NO) and prostaglandin E(2) (PGE(2)) in rat peritoneal macrophages when the cells were pre-incubated with each gold compound for 20 h. Without pre-incubation for 20 h, aurothioglucose and aurothiomalate, but not auranofin, failed to inhibit the TPA-induced NF-kappaB nuclear translocation and production of NO and PGE(2). Auranofin, aurothioglucose and aurothiomalate did not affect the direct binding of NF-kappaB to the DNA probe. It was suggested that these gold compounds inhibit the TPA-induced production of NO and PGE(2) by inhibiting the NF-kappaB nuclear translocation.
Journal of Pharmacy and Pharmacology 03/2003; 55(2):245-51. · 2.17 Impact Factor
