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The dominant role of exogenous or endogenous interleukin-1 beta on expression and activity of inducible nitric oxide synthase in rat microvascular brain endothelial cells.

Abstract

In the brain large amounts of nitric oxide are produced in response to various pathological stimuli such as infectious agents, ischemia and trauma. Although it is known that endothelial cells can express the inducible isoform of nitric oxide synthase (iNOS) upon activation, the impact of different cytokines on iNOS expression in rat microvascular endothelial cells remains unclear. We now investigated iNOS mRNA expression and enzyme activity in primary cell cultures of rat microvascular brain endothelial cells after treatment with the proinflammatory cytokines interleukin-1beta (IL-1beta), Tumor necrosis factor-alpha (TNF-alpha), and interferon-gamma (IFN-gamma) alone or in combination. Cells were characterized by immunocytochemistry staining for von-Willebrand-factor and the rat brain endothelial antigen recognized by monoclonal antibody Ox2. iNOS-enzyme activity was determined by measurement of nitrite in the supernatants of cell culture using the Griess-reaction. In addition mRNA expression was analysed by RT-PCR with iNOS and IL-1beta specific primers. All cells in the endothelial cell culture were found to express the antigenic phenotype vWF+/Ox2+/Ox43-, thus identifying the cells as rat brain endothelial cells of microvascular origin. IL-1beta was the only cytokine that as a single stimulus induced iNOS mRNA expression and iNOS-enzyme activity in these endothelial cells. All combinations of two cytokines, including that of TNF-alpha and IFN-gamma--or the triple combination led to expression of iNOS-mRNA and active protein. Cell activation by the combination of TNF-alpha + IFN-gamma led to an early expression of IL-1beta by the endothelial cells suggesting iNOS induction as a consequence of endogenous IL-1beta production under this challenge. The experiments prove that rat brain microvascular endothelial cells express iNOS and produce large amounts of NO under inflammatory conditions. Furthermore, our results indicate a decisive role of IL-1beta in iNOS expression and NO generation.

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  • Authors+Show Affiliations

    ,

    Department of Neurology, Ruprecht-Karls-University, Heidelberg, Germany. EckhardBonmann@krzmail.krz.uniheidelberg.de

    , ,

    Source

    Neuroscience letters 230:2 1997 Jul 18 pg 109-12

    MeSH

    Animals
    Cells, Cultured
    Cerebrovascular Circulation
    Endothelium, Vascular
    Humans
    Interferon-gamma
    Interleukin-1
    Microcirculation
    Nitric Oxide Synthase
    Polymerase Chain Reaction
    RNA, Messenger
    Rats
    Rats, Wistar
    Recombinant Proteins
    Transcription, Genetic
    Tumor Necrosis Factor-alpha

    Pub Type(s)

    Journal Article
    Research Support, Non-U.S. Gov't

    Language

    eng

    PubMed ID

    9259476

    Citation

    Bonmann, E, et al. "The Dominant Role of Exogenous or Endogenous Interleukin-1 Beta On Expression and Activity of Inducible Nitric Oxide Synthase in Rat Microvascular Brain Endothelial Cells." Neuroscience Letters, vol. 230, no. 2, 1997, pp. 109-12.
    Bonmann E, Suschek C, Spranger M, et al. The dominant role of exogenous or endogenous interleukin-1 beta on expression and activity of inducible nitric oxide synthase in rat microvascular brain endothelial cells. Neurosci Lett. 1997;230(2):109-12.
    Bonmann, E., Suschek, C., Spranger, M., & Kolb-Bachofen, V. (1997). The dominant role of exogenous or endogenous interleukin-1 beta on expression and activity of inducible nitric oxide synthase in rat microvascular brain endothelial cells. Neuroscience Letters, 230(2), pp. 109-12.
    Bonmann E, et al. The Dominant Role of Exogenous or Endogenous Interleukin-1 Beta On Expression and Activity of Inducible Nitric Oxide Synthase in Rat Microvascular Brain Endothelial Cells. Neurosci Lett. 1997 Jul 18;230(2):109-12. PubMed PMID: 9259476.
    * Article titles in AMA citation format should be in sentence-case
    TY - JOUR T1 - The dominant role of exogenous or endogenous interleukin-1 beta on expression and activity of inducible nitric oxide synthase in rat microvascular brain endothelial cells. AU - Bonmann,E, AU - Suschek,C, AU - Spranger,M, AU - Kolb-Bachofen,V, PY - 1997/7/18/pubmed PY - 1997/7/18/medline PY - 1997/7/18/entrez SP - 109 EP - 12 JF - Neuroscience letters JO - Neurosci. Lett. VL - 230 IS - 2 N2 - In the brain large amounts of nitric oxide are produced in response to various pathological stimuli such as infectious agents, ischemia and trauma. Although it is known that endothelial cells can express the inducible isoform of nitric oxide synthase (iNOS) upon activation, the impact of different cytokines on iNOS expression in rat microvascular endothelial cells remains unclear. We now investigated iNOS mRNA expression and enzyme activity in primary cell cultures of rat microvascular brain endothelial cells after treatment with the proinflammatory cytokines interleukin-1beta (IL-1beta), Tumor necrosis factor-alpha (TNF-alpha), and interferon-gamma (IFN-gamma) alone or in combination. Cells were characterized by immunocytochemistry staining for von-Willebrand-factor and the rat brain endothelial antigen recognized by monoclonal antibody Ox2. iNOS-enzyme activity was determined by measurement of nitrite in the supernatants of cell culture using the Griess-reaction. In addition mRNA expression was analysed by RT-PCR with iNOS and IL-1beta specific primers. All cells in the endothelial cell culture were found to express the antigenic phenotype vWF+/Ox2+/Ox43-, thus identifying the cells as rat brain endothelial cells of microvascular origin. IL-1beta was the only cytokine that as a single stimulus induced iNOS mRNA expression and iNOS-enzyme activity in these endothelial cells. All combinations of two cytokines, including that of TNF-alpha and IFN-gamma--or the triple combination led to expression of iNOS-mRNA and active protein. Cell activation by the combination of TNF-alpha + IFN-gamma led to an early expression of IL-1beta by the endothelial cells suggesting iNOS induction as a consequence of endogenous IL-1beta production under this challenge. The experiments prove that rat brain microvascular endothelial cells express iNOS and produce large amounts of NO under inflammatory conditions. Furthermore, our results indicate a decisive role of IL-1beta in iNOS expression and NO generation. SN - 0304-3940 UR - https://www.unboundmedicine.com/medline/citation/9259476/The_dominant_role_of_exogenous_or_endogenous_interleukin_1_beta_on_expression_and_activity_of_inducible_nitric_oxide_synthase_in_rat_microvascular_brain_endothelial_cells_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0304-3940(97)00485-0 DB - PRIME DP - Unbound Medicine ER -