cAMP and purinergic P2y receptors upregulate and enhance inducible NO synthase mRNA and protein in vivo.
Adenosine 3',5'-cyclic monophosphate (cAMP) and purinergic P2y receptor agonists upregulate inducible nitric oxide (NO) synthase (iNOS) but inhibit Escherichia coli endotoxin lipopolysaccharide (LPS)- and cytokine-mediated upregulation of iNOS in cultured cells. We examined the effects of cAMP and P2y receptor agonists on the iNOS system in vivo. Intratracheal administration of dibutyryl-cAMP (DBcAMP, 0.1 and 1 mg/kg), a P2y receptor agonist [2-methylthioadenosine 5'-triphosphate (MeS-ATP), 5 mg/kg], or LPS (0.6 mg/kg) to rats 2 h before bronchoalveolar lavage (BAL) increased iNOS mRNA (competitor-equalized reverse transcription-polymerase chain reaction) and iNOS protein (Western blot) in rat alveolar macrophages compared with the effects of sterile phosphate-buffered saline (0.5 ml it). At equal levels of upregulation of iNOS mRNA, 1) LPS, but not DBcAMP or MeS-ATP, upregulated nuclear transcription factor-kappa B (NF-kappa B) and 2) iNOS protein and formation of NO were greater in alveolar macrophages from LPS- and MeS-ATP-treated rats than from DBcAMP-treated rats. Administration of DBcAMP or MeS-AMP 15 min before LPS did not inhibit LPS-induced alveolar macrophage-derived iNOS mRNA, iNOS protein, and NO. Diethyldithiocarbamate (DETC, 5 mg/kg it) inhibited LPS-induced iNOS mRNA but did not affect upregulation of iNOS mRNA produced by the other agonists. We conclude that an LPS-dependent and -independent pathway of iNOS mRNA induction exists in vivo. The former is activated by IPS and most cytokines, is associated with upregulation of NF-kappa B and inhibited by DETC, and elicits an inflammatory response. The latter, activated by DBcAMP and MeS-ATP, is not associated with upregulation of NF-kappa B, inhibition by DETC, or activation of inflammation. The two systems are additive in vivo rather than antagonistic. Speculatively, if the LPS-independent iNOS pathway exists in humans, the iNOS in tissues from patients taking drugs affecting cAMP or P2y receptors may be iatrogenic rather than pathogenetic in origin.
Greenberg
S S
SS
Department of Medicine, Louisiana State University Medical Center, New Orleans 70112, USA.
Zhao
X
X
Wang
J F
JF
Hua
L
L
Ouyang
J
J
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives* / Q000494:pharmacology; D000420:Albuterol / Q000494:pharmacology; D000818:Animals; D001992:Bronchoalveolar Lavage Fluid / Q000737:chemistry; D003994:Bucladesine / Q000494:pharmacology*; D002634:Chemotaxis, Leukocyte / Q000187:drug effects; D000242:Cyclic AMP / Q000378:metabolism*; D004050:Ditiocarb / Q000494:pharmacology; D004790:Enzyme Induction; D007545:Isoproterenol / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology; D008168:Lung / Q000187:drug effects / Q000502:physiology*; D016676:Macrophages, Alveolar / Q000187:drug effects / Q000201:enzymology* / Q000276:immunology; D008297:Male; D016328:NF-kappa B / Q000096:biosynthesis; D019001:Nitric Oxide Synthase / Q000096:biosynthesis*; D016133:Polymerase Chain Reaction; D058910:Purinergic P2 Receptor Agonists; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D017207:Rats, Sprague-Dawley; D018048:Receptors, Purinergic P2 / Q000502:physiology*; D013873:Thionucleotides / Q000494:pharmacology*; D014158:Transcription, Genetic / Q000187:drug effects / Q000502:physiology*; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis
D008070:Lipopolysaccharides; D016328:NF-kappa B; D058910:Purinergic P2 Receptor Agonists; D012333:RNA, Messenger; D018048:Receptors, Purinergic P2; D013873:Thionucleotides; D014409:Tumor Necrosis Factor-alpha; D003994:Bucladesine; D000255:Adenosine Triphosphate; D004050:Ditiocarb; D000242:Cyclic AMP; D019001:Nitric Oxide Synthase; D007545:Isoproterenol; D000420:Albuterol; C008501:2-methylthio-ATP