Nuclear Factor-κB Mediates Simultaneous Induction of Inducible Nitric-Oxide Synthase and Up-Regulation of the Cationic Amino Acid Transporter CAT-2B in Rat Alveolar Macrophages

  1. Rainer Hammermann,
  2. Maria Donata Messeri Dreißig,
  3. Jutta Mössner,
  4. Margarita Fuhrmann,
  5. Liberato Berrino1,
  6. Manfred Göthert and
  7. Kurt Racké
  1. Institute of Pharmacology and Toxicology, Rheinische Friedrich Wilhelms University, Bonn, Germany

    Abstract

    The connection between the regulation of l-arginine transport and nitric oxide (NO) synthesis was studied in rat alveolar macrophages. Lipopolysaccharides (LPSs) and interferon-γ stimulated in the same concentration- and time-dependent manner NO synthesis (measured by nitrite accumulation) andl-[3H]arginine uptake. This correlated with an increased mRNA expression for iNOS and the cationic amino acid transporter CAT-2B (analyzed by reverse transcription-polymerase chain reaction), with the same kinetics observed for the up-regulation of both mRNAs. Because nuclear factor-κB (NF-κB) is essential for induction of iNOS its role for the regulation of CAT-2B expression andl-arginine transport was investigated. The NF-κB inhibitors pyrrolidine dithiocarbamate andN α-p-tosyl-l-lysine chloromethyl ketone abrogated LPS- and interferon-γ-induced increase of nitrite accumulation and l-[3H]arginine uptake as well as up-regulation of iNOS and CAT-2B mRNA. LPS-induced increase in iNOS and CAT-2B mRNA was also suppressed by specific NF-κB decoy oligodesoxynucleotides, confirming the essential role of NF-κB for iNOS and CAT-2B expression. Dexamethasone did not affect the initial (5 h) LPS-induced increase of iNOS and CAT-2B mRNA, but down-regulated both mRNAs after prolonged (20 h) exposure and this was accompanied by partial inhibition of LPS-stimulated nitrite accumulation and l-[3H]arginine uptake. These findings demonstrate parallel regulation of the expression of iNOS and CAT-2B, and of NO synthesis and l-arginine uptake in rat alveolar macrophages. NF-κB is an essential transcription factor not only for the induction of iNOS, but also for the up-regulation of CAT-2B. The simultaneous up-regulation of CAT-2B with iNOS is considered as a mechanism to ensure a high substrate supply for iNOS.

    Footnotes

    • Send reprint requests to: Dr. Rainer Hammermann, Institute of Pharmacology and Toxicology, University of Bonn, Reuterstr. 2b, D-53113 Bonn, Germany. E-mail:r.hammermann{at}uni-bonn.de

    • ↵1 Current address: Institute of Pharmacology and Toxicology, II University of Naples, Naples, Italy.

    • This work was supported by Deutsche Forschungsgemeinschaft (Ra 400/9-1 and 9-2). M.D.M.D. and L.B. were supported by travel grants of the “Vigoni Programm” of the Deutsche Akademische Auslandsdienst and the Conferenza Permanente dei Rettori delle UniversitàItaliane, respectively.

    • Abbreviations:
      MΦ
      macrophages
      NO
      nitric oxide
      NOS
      nitric-oxide synthase
      iNOS
      inducible nitric-oxide synthase
      AMΦ
      alveolar macrophages
      LPS
      lipopolysaccharide
      IFN-γ
      interferon-γ
      NF-κB
      nuclear factor-κB
      CAT
      cationic amino acid transporter
      ODN
      oligodesoxynucleotide
      PDTC
      pyrrolidine dithiocarbamate
      TLCK
      Nα-p-tosyl-l-lysine chloromethyl ketone
      RT
      reverse transcription
      PCR
      polymerase chain reaction
      bp
      base pair
      • Received March 23, 2000.
      • Accepted September 7, 2000.
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    1. Molecular Pharmacology December 1, 2000 vol. 58 no. 6 1294-1302
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