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John Girdlestone - One of the best experts on this subject based on the ideXlab platform.
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ifn α super induction of hla class i expression by a variant Thymoma Cell Line involves nuclear translocation of rel complexes
European Journal of Immunology, 1998Co-Authors: Helen Travers, John GirdlestoneAbstract:: Variant Thymoma Lines have been described which exhibit a substantially increased level of HLA class I induction by IFN-alpha, but not by IFN-gamma, and an unchanged response of other IFN-alpha-stimulated genes (Burrone et al., EMBO J. 1985. 4: 2855-2860). We report that their amplified response correlates with the nuclear translocation of Rel transcription factors upon prolonged treatment with IFN-alpha. The variant Cells contain an IkappaBalpha subset with a significantly shortened half-life, and a constitutively active form of IkappaBalpha efficiently blocks HLA class I induction. Therefore, in addition to STAT-mediated induction, prolonged exposure to IFN-alpha can affect transcription involving Rel factors, which are implicated in the regulation of numerous immune response and viral genes.
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IFN‐α super‐induction of HLA class I expression by a variant Thymoma Cell Line involves nuclear translocation of Rel complexes
European Journal of Immunology, 1998Co-Authors: Helen Travers, John GirdlestoneAbstract:: Variant Thymoma Lines have been described which exhibit a substantially increased level of HLA class I induction by IFN-alpha, but not by IFN-gamma, and an unchanged response of other IFN-alpha-stimulated genes (Burrone et al., EMBO J. 1985. 4: 2855-2860). We report that their amplified response correlates with the nuclear translocation of Rel transcription factors upon prolonged treatment with IFN-alpha. The variant Cells contain an IkappaBalpha subset with a significantly shortened half-life, and a constitutively active form of IkappaBalpha efficiently blocks HLA class I induction. Therefore, in addition to STAT-mediated induction, prolonged exposure to IFN-alpha can affect transcription involving Rel factors, which are implicated in the regulation of numerous immune response and viral genes.
J.m. Bidlack - One of the best experts on this subject based on the ideXlab platform.
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The synthetic κ-opioid agonist (−)U50,488 does not affect calcium transport into R1.1 mouse Thymoma Cell Line
International Journal of Immunopharmacology, 1999Co-Authors: I. Martin-kleiner, J.m. BidlackAbstract:Abstract In this paper, the effect of the synthetic κ-opioid agonist (−)U50,488 on 45 Ca 2 transport into R1.1 mouse Thymoma Cells is presented. This Thymoma Cell Line expresses selectively the κ -opioid class of receptors. 45 Ca 2 transport into R1.1 Cells was not affected by the κ -opioid agonist (−)U50,488 (10 −10 M–10 −4 M) alone, or in the presence of the plant lectins: PHA (250 μ g/ml) and Con A (800 μ g/ml), after a 60 min treatment. The plant lectins PHA and Con A stimulated 45 Ca 2 transport into R1.1 Cells, in high concentrations (100–800 μ g/ml) and (200–1000 μ g/ml) respectively, after a 60 min treatment. Thus, 45 Ca 2 transport was not affected in R1.1 Cells by the κ -opioid agonist (−) U50,488 alone, or in the presence of mitogens after a 60 min treatment. This negative result does not indicate the lack of calcium channels on R1.1 Cells, since the plant lectins PHA and Con A were able to stimulate 45 Ca 2 transport into R1.1 Cells.
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the synthetic κ opioid agonist u50 488 does not affect calcium transport into r1 1 mouse Thymoma Cell Line
International Journal of Immunopharmacology, 1999Co-Authors: I Martinkleiner, J.m. BidlackAbstract:Abstract In this paper, the effect of the synthetic κ-opioid agonist (−)U50,488 on 45 Ca 2 transport into R1.1 mouse Thymoma Cells is presented. This Thymoma Cell Line expresses selectively the κ -opioid class of receptors. 45 Ca 2 transport into R1.1 Cells was not affected by the κ -opioid agonist (−)U50,488 (10 −10 M–10 −4 M) alone, or in the presence of the plant lectins: PHA (250 μ g/ml) and Con A (800 μ g/ml), after a 60 min treatment. The plant lectins PHA and Con A stimulated 45 Ca 2 transport into R1.1 Cells, in high concentrations (100–800 μ g/ml) and (200–1000 μ g/ml) respectively, after a 60 min treatment. Thus, 45 Ca 2 transport was not affected in R1.1 Cells by the κ -opioid agonist (−) U50,488 alone, or in the presence of mitogens after a 60 min treatment. This negative result does not indicate the lack of calcium channels on R1.1 Cells, since the plant lectins PHA and Con A were able to stimulate 45 Ca 2 transport into R1.1 Cells.
I Martinkleiner - One of the best experts on this subject based on the ideXlab platform.
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the κ opioid agonist u 69593 affects intraCellular calcium level in r1 1 mouse Thymoma Cell Line
International Immunopharmacology, 2002Co-Authors: I Martinkleiner, Jelka GabrilovacAbstract:In this study, the effect of the κ-opioid agonist U-69593 on the intraCellular calcium level in R1.1. Cells was investigated using FURA 2-AM dye. In the previous study, calcium transport into R1.1 Cells was not affected by the κ-opioid agonist (−)U50,488 [Int. J. Immunopharmacol. 21 (1999) 133]. In this study, the κ-opioid agonist U-69593 (10−10–10−6 M), decreased intraCellular calcium level in unstimulated Cells. This decrease could not be reversed by the κ-opioid antagonist NBI (10−5 or 10−6 M). Ionophore A23187 was used to increase intraCellular calcium level. Stimulation of intraCellular calcium level by Ionophore A23187 was potentiated by the κ-opioid agonist U-69593. Thus, we have shown that basal intraCellular calcium level was decreased in R1.1 by the κ-opioid agonist U-69593 and increased in R1.1 Cells stimulated by Ionophore A23187.
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the synthetic κ opioid agonist u50 488 does not affect calcium transport into r1 1 mouse Thymoma Cell Line
International Journal of Immunopharmacology, 1999Co-Authors: I Martinkleiner, J.m. BidlackAbstract:Abstract In this paper, the effect of the synthetic κ-opioid agonist (−)U50,488 on 45 Ca 2 transport into R1.1 mouse Thymoma Cells is presented. This Thymoma Cell Line expresses selectively the κ -opioid class of receptors. 45 Ca 2 transport into R1.1 Cells was not affected by the κ -opioid agonist (−)U50,488 (10 −10 M–10 −4 M) alone, or in the presence of the plant lectins: PHA (250 μ g/ml) and Con A (800 μ g/ml), after a 60 min treatment. The plant lectins PHA and Con A stimulated 45 Ca 2 transport into R1.1 Cells, in high concentrations (100–800 μ g/ml) and (200–1000 μ g/ml) respectively, after a 60 min treatment. Thus, 45 Ca 2 transport was not affected in R1.1 Cells by the κ -opioid agonist (−) U50,488 alone, or in the presence of mitogens after a 60 min treatment. This negative result does not indicate the lack of calcium channels on R1.1 Cells, since the plant lectins PHA and Con A were able to stimulate 45 Ca 2 transport into R1.1 Cells.
Jean M Bidlack - One of the best experts on this subject based on the ideXlab platform.
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Kappa Opioid Receptors on Immune Cells as Studied by Fluorescent Ligands
Advances in Experimental Medicine and Biology, 1996Co-Authors: Jean M Bidlack, Diane M P Lawrence, Tracey A. IgnatowskiAbstract:Despite both functional (1–4) and molecular biological (5–8) evidence for the presence of opioid receptors on lymphocytes, the search for opioid binding sites on leukocytes has been difficult. Opioid binding sites possessing all the characteristics of the classical brain opioid receptors had not been detected until the identification of the κ opioid receptor on the mouse Rl.l Thymoma Cell Line (9,10). Possible reasons why opioid receptors have been difficult to detect on mixed Cell populations from the immune system include: 1) opioid receptors may be expressed on only a small population of lymphocytes; 2) the expression of opioid receptors on lymphocyte changes during lymphocyte maturation; and 3) radioreceptor binding methodology is not sufficiently sensitive to detect opioid receptors present at a low density. While the R1.1 Cell Line was useful in establishing that a lymphocyte could express an opioid receptor, interest still lies in determining if lymphocytes from a mixed Cell population, such as thymus and spleen, express opioid receptors. To address this question, we developed an indirect immunofluorescent method that is more sensitive than radioreceptor binding assays (11,12).
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the kappa opioid receptor expressed on the mouse r1 1 Thymoma Cell Line down regulates without desensitizing during chronic opioid exposure
Journal of Pharmacology and Experimental Therapeutics, 1995Co-Authors: David B Joseph, Jean M BidlackAbstract:The R1.1 mouse Thymoma Cell Line expresses a single class of kappa opioid receptors that is negatively coupled to adenylyl cyclase through a Bordetella pertussis toxin-sensitive inhibitory guanine nucleotide-binding protein. The aim of the present study was to determine whether chronic opioid treatment of R1.1 Cells altered either the binding properties or the functional response associated with the kappa opioid receptor. Culturing of R1.1 Cells with the kappa-selective agonist (trans)-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)-cyclohexyl] benzeneacetamide methane-sulfonate hydrate (U50,488) for 3 hr and longer, followed by extensive washing of R1.1 Cell membranes, produced a concentration- and time-dependent reduction in the binding of the kappa-selective ligand (5 alpha,7 alpha,8 beta)-(-)-N-methyl-N-(7-(1-pyrrolidinyl)-1- oxaspiro(4,5)dec-8-yl) benzeneacetamide ([3H]U69,593). Culturing of R1.1 Cells with 100 nM U50,488 for 24 hr produced approximately a 50% reduction in the Bmax value for [3H]U69,593 and [3H]naloxone binding. In contrast to the reduction in binding, there was no change in the inhibition of adenylyl cyclase activity by (-)-U50,488. To determine whether kappa opioid receptor function was maintained by spare receptors after agonist-induced down-regulation, membranes from untreated R1.1 Cells were incubated with 400 nM of the irreversible opioid antagonist beta-chlornaltrexamine (beta-CNA) followed by extensive washing. beta-CNA produced a 50% reduction in the [3H]U69,593 binding and a 6-fold increase in the IC50 value for (-)-U50,488 inhibition of adenylyl cyclase activity, with no change in the maximal inhibition of cyclic AMP levels.(ABSTRACT TRUNCATED AT 250 WORDS)
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The κ-opioid receptor expressed on the mouse lymphoma Cell Line R1.1 contains a sulfhydryl group at the binding site
European Journal of Pharmacology, 1994Co-Authors: David B Joseph, Jean M BidlackAbstract:Abstract Studies were directed at determining whether the κ-opioid receptor expressed on the mouse R1.1 Thymoma Cell Line contained either a disulfide bond or a sulfhydryl group at the opioid binding site. The binding of the κ-opioid receptor agonist [3H](−)-bremazocine to R1.1 Cell membranes was unchanged following treatment with the disulfide bond-reducing reagent dithiothreitol at concentrations up to 130 mM. However, treatment of membranes with the sulfhydryl-alkylating reagent N-ethylmaleimide, followed by extensive washing, reduced [3H](−)-bremazocine binding by as much as 90%. Inhibition of [3H](−)-bremazocine binding by N-ethylmaleimide was concentration- and time-dependent. When R1.1 Cell membranes were treated with 1 mM N-ethylmaleimide for 10 min at 24°C, the Bmax value for [3H](−)-bremazocine binding decreased by 50%, with no change in receptor affinity. N-Ethylmaleimide-induced reduction of [3H](−)-bremazocine binding was attenuated by pretreatment of membranes with the κ-selective opioids U50,488 and U69,593. The results indicate that a sulfhydryl group is present at or near the binding site on the κ-opioid receptor expressed by the R1.1 Thymoma Cell Line.
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the kappa opioid receptor expressed on the mouse r1 1 Thymoma Cell Line is coupled to adenylyl cyclase through a pertussis toxin sensitive guanine nucleotide binding regulatory protein
Journal of Pharmacology and Experimental Therapeutics, 1993Co-Authors: Diane M P Lawrence, Jean M BidlackAbstract:The R1.1 mouse Thymoma Cell Line expresses a high-affinity kappa opioid binding site. Opioid binding to this site is inhibited by guanine nucleotides, suggesting that the receptor is coupled to a guanine nucleotide-binding protein. Here, we present evidence that the kappa opioid binding site on R1.1 Cell membranes is negatively coupled to adenylyl cyclase. The kappa-selective agonists (trans)-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)- cyclohexyl]benzeneacetamide methane-sulfonate hydrate [(-)-U50,488], (5 alpha,7 alpha, 8 beta)-(-)-N-methyl-N-(7-(1-pyrrolidinyl)-1-oxas- piro(4,5)dec-8-yl)benzeneacetamide (U69,593) and several dynorphin peptides inhibited basal and forskolin-stimulated cyclic AMP production by up to 40% in R1.1 Cell membranes. The order of potency for the inhibition of adenylyl cyclase activity by opioid agonists correlated with their Ki values for the inhibition of [3H]U69,593 binding. Opioid-mediated inhibition of adenylyl cyclase activity was stereoselective, as (-)-U50,488 was more potent than the (+) isomer, and the inhibition was blocked by the kappa-selective antagonist nor-binaltorphimine. The opioid-mediated inhibition of adenylyl cyclase activity was also completely blocked by incubating R1.1 Cells with Bordetella pertussis toxin (PTX). Incubation of R1.1 Cell membranes with PTX and [adenylate-32P]NAD+ resulted in the exclusive labeling of a 41-kDa protein, as determined by separating the membrane proteins under reducing conditions on a SDS polyacrylamide gel, followed by autoradiography. These results suggest that a PTX-sensitive inhibitory guanine nucleotide-binding protein mediates the link between the Thymoma kappa opioid receptor and adenylyl cyclase.
John W Kappler - One of the best experts on this subject based on the ideXlab platform.
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ikaros promotes rearrangement of tcr α genes in an ikaros null Thymoma Cell Line
European Journal of Immunology, 2013Co-Authors: Bernard Collins, Eric T Clambey, James Scottbrowne, Janice White, Philippa Marrack, James Hagman, John W KapplerAbstract:Ikaros is important in the development and maintenance of the lymphoid system, functioning in part by associating with chromatin-remodeling complexes. We have studied the functions of Ikaros in the transition from pre-T Cell to the CD4+CD8+ thymocyte using an Ikaros null CD4−CD8− mouse Thymoma Cell Line (JE131). We demonstrate that this Cell Line carries a single functional TCR β gene rearrangement and expresses a surface pre-TCR. JE131 Cells also carry nonfunctional rearrangements on both alleles of their TCR α loci. Retroviral reintroduction of Ikaros dramatically increased the rate of transcription in the α locus and TCR Vα/Jα recombination resulting in the appearance of many new αβTCR+ Cells. The process is RAG dependent, requires switch/sucrose nonfermentable chromatin-remodeling complexes and is coincident with the binding of Ikaros to the TCR α enhancer. Furthermore, knockdown of Mi2/nucleosome remodeling and deacetylase complexes increased the frequency of TCR α rearrangement. Our data suggest that Ikaros controls Vα/Jα recombination in T Cells by controlling access of the transcription and recombination machinery to the TCR α loci. The JE131 Cell Line should prove to be a very useful tool for studying the molecular details of this and other processes involved in the pre-T Cell to αβTCR+ CD4+CD8+ thymocyte transition.