Nuclear factor-kappaB 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 oxide (NO) synthesis was studied in rat alveolar macrophages. Lipopolysaccharides (LPSs) and interferon-gamma stimulated in the same concentration- and time-dependent manner NO synthesis (measured by nitrite accumulation) and L-[(3)H]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-kappaB (NF-kappaB) is essential for induction of iNOS its role for the regulation of CAT-2B expression and L-arginine transport was investigated. The NF-kappaB inhibitors pyrrolidine dithiocarbamate and N(alpha)-p-tosyl-L-lysine chloromethyl ketone abrogated LPS- and interferon-gamma-induced increase of nitrite accumulation and L-[(3)H]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-kappaB decoy oligodesoxynucleotides, confirming the essential 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 and L-[(3)H]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-kappaB 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.
Hammermann
R
R
Institute of Pharmacology and Toxicology, Rheinische Friedrich Wilhelms University, Bonn, Germany. r.hammermann@uni-bonn.de
Dreissig
M D
MD
Mössner
J
J
Fuhrmann
M
M
Berrino
L
L
Göthert
M
M
Racké
K
K
D026923:Amino Acid Transport Systems, Basic; D000818:Animals; D001120:Arginine / Q000378:metabolism; D002352:Carrier Proteins / Q000235:genetics*; D002478:Cells, Cultured; D003513:Cycloheximide / Q000494:pharmacology; D003907:Dexamethasone / Q000494:pharmacology; D004347:Drug Interactions; D004790:Enzyme Induction; D005260:Female; D058851:GPI-Linked Proteins; D005786:Gene Expression Regulation; D007371:Interferon-gamma / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology; D016676:Macrophages, Alveolar / Q000201:enzymology / Q000502:physiology*; D008297:Male; D008565:Membrane Proteins / Q000235:genetics*; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000502:physiology*; D009569:Nitric Oxide / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000096:biosynthesis* / Q000235:genetics; D052247:Nitric Oxide Synthase Type II; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D012333:RNA, Messenger / Q000096:biosynthesis / Q000187:drug effects; D051381:Rats; D017207:Rats, Sprague-Dawley; D018124:Receptors, Tumor Necrosis Factor / Q000378:metabolism; D053320:Receptors, Tumor Necrosis Factor, Member 10c; D015398:Signal Transduction / Q000187:drug effects; D053319:Tumor Necrosis Factor Decoy Receptors; D015854:Up-Regulation
D026923:Amino Acid Transport Systems, Basic; D002352:Carrier Proteins; D058851:GPI-Linked Proteins; D008070:Lipopolysaccharides; D008565:Membrane Proteins; D016328:NF-kappa B; D011500:Protein Synthesis Inhibitors; D012333:RNA, Messenger; D018124:Receptors, Tumor Necrosis Factor; D053320:Receptors, Tumor Necrosis Factor, Member 10c; C108661:TNFRSF10C protein, human; D053319:Tumor Necrosis Factor Decoy Receptors; D009569:Nitric Oxide; D003907:Dexamethasone; D007371:Interferon-gamma; D001120:Arginine; D003513:Cycloheximide; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496318:Nos2 protein, rat