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

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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 andNα-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.

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