Nuclear factor-kappa B mediates simultaneous induction of inducible nitric-oxide synthase and up-regulation of the cationic amino acid transporter CAT-2B in rat alveolar macrophages

The connection between the regulation of L-arginine transport and nitric ox ide (NO) synthesis was studied in rat alveolar macrophages. Lipopolysacchar ides (LPSs) and interferon-gamma stimulated in the same concentration- and time-dependent manner NO synthesis (measured by nitrite accumulation) and L -[H-3] arginine uptake. This correlated with an increased mRNA expression f or 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-kappaB (NF-kapp aB) is essential for induction of iNOS its role for the regulation of CAT-2 B expression and L- arginine transport was investigated. The NF-kappaB inhi bitors pyrrolidine dithiocarbamate and N-alpha-p-tosyl- L-lysine chlorometh yl ketone abrogated LPS- and interferon-gamma -induced increase of nitrite accumulation and L-[H-3] arginine uptake as well as up-regulation of iNOS a nd CAT-2B mRNA. LPS-induced increase in iNOS and CAT-2B mRNA was also suppr essed by specific NF-kappaB decoy oligodesoxynucleotides, confirming the es sential role of NF-kappaB 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 a nd L-[H-3] arginine uptake. These findings demonstrate parallel regulation of the expression of iNOS and CAT-2B, and of NO synthesis and L- arginine u ptake in rat alveolar macrophages. NF-kappaB is an essential transcription factor not only for the induction of iNOS, but also for the up-regulation o f CAT-2B. The simultaneous up-regulation of CAT-2B with iNOS is considered as a mechanism to ensure a high substrate supply for iNOS.